Annex 1 : Manufacture of Sterile Products Document map Section

Annex 1 : Manufacture of Sterile Products 1

Document map 3

Section Number General overview

1. Scope Includes additional areas (other than sterile products) where the

general principles of the annex can be applied.

2. Principle General principles as applied to the manufacture of sterile

products.

3. Pharmaceutical Quality

System (PQS)

Highlights the specific requirements of the PQS when applied

to sterile products.

4. Premises General guidance regarding the specific needs for premises

design and also guidance on the qualification of premises

including the use of Barrier Technology.

5. Equipment General guidance on the design and operation of equipment.

6. Utilities Guidance with regards to the special requirements of utilities

such as water, gas and vacuum.

7. Personnel Guidance on the requirements for specific training, knowledge

and skills. Also gives guidance to the qualification of

personnel.

8. Production and specific

technologies

Discusses the approaches to be taken with regards to aseptic

and terminal sterilization processes. Discusses approaches to

sterilization of products, equipment and packaging

components. Also discusses different technologies such as

lyophilization and Form-Fill-Seal where specific requirements

apply.

9. Viable and non-viable

environmental and process

monitoring

This section differs from guidance given in section 4 in that the

guidance here applies to ongoing routine monitoring with

regards to the design of systems and setting of action limits

alert levels and reviewing trend data.

The section also gives guidance on the requirements of Aseptic

Process Simulation (APS).

10. Quality control (QC) Gives guidance on some of the specific Quality Control

requirements relating to sterile products.

11. Glossary Explanation of specific terminology.

1 Scope 6

The manufacture of sterile products covers a wide range of sterile product types (active substance, 8

sterile excipient, primary packaging material and finished dosage form), packed sizes (single unit to 9

multiple units), processes (from highly automated systems to manual processes) and technologies (e.g. 10

biotechnology, classical small molecule manufacturing and closed systems). This Annex provides 11

general guidance that should be used for the manufacture of all sterile products using the principles of 12

Quality Risk Management (QRM), to ensure that microbial, particulate and pyrogen contamination is 13

prevented in the final product. 14

QRM applies to this document in its entirety and will not be referred to in specific paragraphs. Where 16

specific limits or frequencies are written, these should be considered as a minimum requirement. They 17

are stated due to regulatory historical experience of issues that have previously been identified and 18

have impacted the safety of patients. 19

The intent of the Annex is to provide guidance for the manufacture of sterile products. However, 21

some of the principles and guidance, such as contamination control strategy, design of premises, 22

cleanroom classification, qualification, monitoring and personnel gowning, may be used to support 23

the manufacture of other products that are not intended to be sterile such as certain liquids, creams, 24

ointments and low bioburden biological intermediates but where the control and reduction of 25

microbial, particulate and pyrogen contamination is considered important. Where a manufacturer 26

elects to apply guidance herein to non-sterile products, the manufacturer should clearly document 27

which principles have been applied and acknowledge that compliance with those principles should be 28

demonstrated. 29

2 Principle 31

2.1 The manufacture of sterile products is subject to special requirements in order to minimize risks of 33

microbial, particulate and pyrogen contamination. The following key areas should be considered: 34

i. Facility, equipment and process design should be optimized, qualified and validated 36

according to the relevant sections of the Good Manufacturing Practices (GMP) guide. 37

The use of appropriate technologies (e.g. Restricted Access Barriers Systems (RABS), 38

isolators, robotic systems, rapid microbial testing and monitoring systems) should be 39

considered to increase the protection of the product from potential extraneous sources of 40

particulate and microbial contamination such as personnel, materials and the surrounding 41

environment, and assist in the rapid detection of potential contaminants in the 42

environment and product. 43

ii. Personnel should have adequate qualifications and experience, training and attitude with a 45

specific focus on the principles involved in the protection of sterile product during the 46

manufacturing, packaging and distribution processes. 47

iii. Processes and monitoring systems for sterile product manufacture should be designed, 49

commissioned, qualified and monitored by personnel with appropriate process, engineering 50

and microbiological knowledge. 51

2.2 Processes, equipment, facilities and manufacturing activities should be managed in accordance 53

with QRM principles to provide a proactive means of identifying, scientifically evaluating and 54

controlling potential risks to quality. Where alternative approaches are used, these should be 55

supported by appropriate rationales and risk assessment and should meet the intent of this Annex. 56

QRM priorities should include good design of the facility, equipment and process in the first instance, 57

then implementation of well-designed procedures, with monitoring systems as the final element that 58

demonstrate that the design and procedures have been correctly implemented and continue to perform 59

in line with expectations. Exclusively monitoring or testing does not give assurance of sterility. 60

2.3 Quality Assurance is particularly important, and manufacture of sterile products must strictly 62

follow carefully established and validated methods of manufacture and control. A Contamination 63

Control Strategy (CCS) should be implemented across the facility in order to define all critical control 64

points and assess the effectiveness of all the controls (design, procedural, technical and 65

organisational) and monitoring measures employed to manage risks associated with contamination. 66

The CCS should be actively updated and should drive continuous improvement of the manufacturing 67

and control methods. 68

2.4 Contamination control and steps taken to minimize the risk of contamination from microbial and 70

particulate sources are a series of successively linked events and measures. These are typically 71

assessed, controlled and monitored individually but their collective effectiveness should be considered 72

altogether. 73

2.5 The development of the CCS requires thorough technical and process knowledge. Potential 75

sources of contamination are attributable to microbial and cellular debris (e.g. pyrogen, endotoxins) as 76

well as particulate matter (e.g. glass and other visible and sub-visible particulates). 77

Elements to be considered within a documented CCS should include (but are not limited to): 78

i. Design of both the plant and processes. 80

ii. Premises and equipment. 82

iv. Personnel. 84

v. Utilities. 86

vi. Raw material controls – including in-process controls. 88

vii. Product containers and closures. 90

viii. Vendor approval – such as key component suppliers, sterilization of components and single 92

use systems (SUS), and services. 93

ix. For outsourced services, such as sterilization, sufficient evidence should be provided to the 95

contract giver to ensure the process is operating correctly. 96

x. Process risk assessment. 98

xi. Process validation. 100

101

xii. Preventative maintenance – maintaining equipment, utilities and premises (planned and 102

unplanned maintenance) to a standard that will not add significant risk of contamination. 103

104

xiii. Cleaning and disinfection. 105

106

xiv. Monitoring systems - including an assessment of the feasibility of the introduction of 107

scientifically sound, modern methods that optimize the detection of environmental 108

contamination. 109

110

xv. Prevention – trending, investigation, corrective and preventive actions (CAPA), root cause 111

determination and the need for more comprehensive investigational tools. 112

113

xvi. Continuous improvement based on information derived from the above. 114

115

2.6 The CCS should consider all aspects of contamination control and its life cycle with ongoing and 116

periodic review resulting in updates within the quality system as appropriate. 117

118

2.7 The manufacturer should take all steps and precautions necessary to assure the sterility of the 119

products manufactured within its facilities. Sole reliance for sterility or other quality aspects should 120

not be placed on any terminal process or finished product test. 121

122

3 Pharmaceutical Quality System (PQS) 123

3.1 The manufacture of sterile products is a complex activity that requires specific controls and 124

measures to ensure the quality of products manufactured. Accordingly, the manufacturer’s PQS 125

should encompass and address the specific requirements of sterile product manufacture and ensure 126

that all activities are effectively controlled so that microbial, particulate and pyrogen contamination is 127

minimized in sterile products. In addition to the PQS requirements detailed in Chapter 1 of the GMPs, 128

the PQS for sterile product manufacture should also ensure that: 129

130

i. An effective risk management system is integrated into all areas of the product life cycle 131

with the aim to minimize microbial contamination and to ensure the quality of sterile 132

products manufactured. 133

134

ii. The manufacturer has sufficient knowledge and expertise in relation to the products 135

manufactured and the equipment, engineering and manufacturing methods employed that 136

have an impact on product quality. 137

138

iii. Root cause analysis of procedural, process or equipment failure is performed in such a 139

way that the risk to product is correctly understood and suitable corrective and 140

preventative actions (CAPA) are implemented. 141

142

iv. Risk management is applied in the development and maintenance of the CCS, to identify, 143

assess, reduce/eliminate (where applicable) and control contamination risks. Risk 144

management should be documented and should include the rationale for decisions taken 145

in relation to risk reduction and acceptance of residual risk. 146

147

v. The risk management outcome should be reviewed regularly as part of on-going quality 148

management, during change control and during the periodic product quality review. 149

150

vi. Processes associated with the finishing and transport of sterile products should not 151

compromise the sterile product. Aspects that should be considered include: container 152

integrity, risks of contamination and avoidance of degradation by ensuring that products 153

are stored and maintained in accordance with the registered storage conditions. 154

155

vii. Persons responsible for the quality release of sterile products have appropriate access to 156

manufacturing and quality information and possess adequate knowledge and experience 157

in the manufacture of sterile products and their critical quality attributes. This is in order 158

to allow such persons to ascertain that the sterile products have been manufactured in 159

accordance with the registered specifications and are of the required quality. 160

161

3.2 All non-conformities, such as sterility test failures, environmental monitoring excursions or 162

deviations from established procedures should be investigated. The investigation should determine the 163

potential impact upon process and product quality and whether any other processes or batches are 164

potentially impacted. The reason for including or excluding a product or batch from the scope of the 165

investigation should be clearly justified and recorded. 166

167

4 Premises 168

169

4.1 The manufacture of sterile products should be carried out in appropriate cleanrooms, entry to 170

which should be through changing rooms that act as airlocks for personnel and airlocks for 171

equipment and materials. Cleanrooms should be maintained to an appropriate cleanliness standard 172

and supplied with air which has passed through filters of an appropriate efficiency. Controls and 173

monitoring should be scientifically justified and capable of evaluating the state of environmental 174

conditions for cleanrooms, airlocks and pass-throughs used for material and equipment transfer. 175

176

4.2 The various operations of component preparation, product preparation and filling should be 177

carried out with appropriate technical and operational separation measures within the cleanroom or 178

facility to prevent mix up and contamination. 179

180

4.3 Restricted Access Barrier Systems (RABS) and isolators are beneficial in assuring the required 181

conditions and minimizing the microbial contamination associated with direct human interventions 182

in the critical zone. Their use should be considered in the CCS. Any alternative approaches to the use 183

of RABS or isolators should be justified. 184

185

4.4 For the manufacture of sterile products there are four grades of cleanroom. 186

187

Grade A zone: The critical zone for high risk operations or for making aseptic connections by 188

ensuring protection by first air (e.g. aseptic processing line, filling zone, stopper bowl, open 189

ampoules and vials). Normally, such conditions are provided by a localised airflow protection, 190

such as unidirectional airflow work stations, RABS or isolators. The maintenance of 191

unidirectional airflow should be demonstrated and qualified across the whole of the Grade A 192

zone. Direct intervention (e.g. without the protection of barrier and glove port technology) into 193

the Grade A zone by operators should be minimized by premises, equipment, process and 194

procedural design. 195

196

Grade B area: For aseptic preparation and filling, this is the background cleanroom for the 197

Grade

A zone (where it is not an isolator). When transfer holes are used to transfer filled, 198

closed products to an adjacent cleanrooms of a lower grade, airflow visualization studies should 199

demonstrate that air does not ingress from the lower grade cleanrooms to the Grade B. Pressure 200

differentials should be continuously monitored. Cleanrooms of lower grade than Grade B can 201

be considered where isolator technology is used (refer to paragraph 4.22). 202

203

Grade C and D area: These are cleanrooms used for carrying out less critical stages in the 204

manufacture of aseptically filled sterile

products but can be used for the preparation /filling 205

of terminally sterilized products. (See section 8 for the specific details on terminal sterilization 206

activities). 207

208

4.5 In cleanrooms, all exposed surfaces should be smooth, impervious and unbroken in order

to 209

minimize the shedding or accumulation of particulates or micro-organisms and to permit the

210

repeated application of cleaning, disinfectant and sporicidal agents where used. 211

212

4.6 To reduce accumulation of dust and to facilitate cleaning there should be no recesses that are 213

difficult to clean effectively therefore projecting ledges, shelves, cupboards and equipment should be 214

kept to a minimum. Doors should be designed to avoid recesses that cannot be cleaned. 215

216

4.7 Materials used in cleanrooms should be selected to minimize generation of particles. 217

218

4.8 Ceilings should be designed and sealed to prevent contamination from the space above them. 219

220

4.9 Sinks and drains are prohibited in Grade A zone and Grade B area. In other cleanrooms, air 221

breaks should be fitted between the machine or sink and the drains. Floor drains in lower grade 222

cleanrooms should be fitted with traps or water seals designed to prevent back flow and should be 223

regularly cleaned, disinfected and maintained. 224

225

4.10 The transfer of equipment and materials into and out of the cleanrooms and critical zones is one 226

of the greatest potential sources of contamination. Any activities with the potential to compromise 227

the cleanliness of cleanrooms or the critical zone should be assessed and if they cannot be 228

eliminated, appropriate controls should be implemented. 229

230

4.11 The transfer of materials, equipment, and components into an aseptic processing area should be 231

carried out via a unidirectional process. Where possible, items should be sterilized and passed into 232

the area through double-ended sterilizers (e.g. through a double-door autoclave or depyrogenation 233

oven/tunnel) sealed into the wall. Where sterilization on transfer of the items is not possible, a 234

procedure which achieves the same objective of not introducing contaminant should be validated and 235

implemented, (e.g. using an effective transfer disinfection, rapid transfer systems for isolators or, for 236

gaseous or liquid materials, a bacteria-retentive filter). 237

238

4.12 Airlocks should be designed and used to provide physical separation and to minimize microbial 239

and particulate contamination of the different areas, and should be present for material and personnel 240

moving between different grades. Wherever possible, airlocks used for personnel movement should 241

be separated from those used for material movement. Where this is not practical, time-based 242

separation of movement (personnel /material) by procedure should be considered. Airlocks should be 243

flushed effectively with filtered air to ensure that the grade of the cleanroom is maintained. The final 244

stage of the airlock should, in the “at rest” state, be of the same cleanliness grade (viable and non-245

viable) as the cleanroom into which it leads. The use of separate changing rooms for entering and 246

leaving Grade B cleanrooms is desirable. Where this is not practical, time-based separation of 247

activities (ingress/egress) by procedure should be considered. Where the CCS indicates that the risk 248

of cross-contamination is high, separate changing rooms for entering and leaving production areas 249

should be considered. Airlocks should be designed as follow: 250

251

i. Personnel airlocks: Areas of increasing cleanliness used for entry of personnel (e.g. from 252

Grade D to Grade C to Grade B). In general hand washing facilities should be provided 253

only in the first stage of the changing room and not be present in changing rooms directly 254

accessing Grade B cleanrooms. 255

256

ii. Material airlocks: used for materials and equipment transfer. 257

258

• Only materials and equipment that have been included on an approved list, developed 259

during validation of the transfer process, should be allowed to be transferred into the 260

Grade A zone or Grade B cleanroom via an airlock or pass-through hatch. Equipment 261

and materials (intended for use in the Grade A zone) should be protected when 262

transiting through the Grade B cleanroom. Any unapproved items that require transfer 263

should be pre-approved as an exception. Appropriate risk assessment and mitigation 264

measures should be applied and recorded as per the manufacturer's CCS and should 265

include a specific disinfection and monitoring programme approved by quality 266

assurance. 267

268

• Pass-through hatches should be designed to protect the higher grade environment, for 269

example by effective flushing with an active filtered air supply. 270

271

• The movement of material or equipment from lower grade or unclassified area to 272

higher grade clean areas should be subject to cleaning and disinfection commensurate 273

with the risk and in line with the CCS. 274

275

4.13 Both sets of doors for pass-throughs and airlocks (for material and personnel) should not be 276

opened simultaneously. For airlocks leading to a Grade A zone and Grade B areas, an interlocking 277

system should be used. For airlocks leading to Grade C and D cleanrooms, a

visual and/or audible 278

warning system should be operated as a minimum. Where required to maintain zone segregation, a 279

time delay between the closing and opening of interlocked doors should be established. 280

281

4.14 Cleanrooms should be supplied with a filtered air supply that maintains a positive pressure 282

and/or an airflow relative to the background environment of a lower grade under all operational 283

conditions and should flush the area effectively. Adjacent rooms of different grades should have 284

pressure differentials of a minimum of 10 pascals (guidance value). Particular attention should be 285

paid to the protection of the critical zone. The recommendations regarding air supplies and pressures 286

may need to be modified where it is necessary to contain certain materials (e.g. pathogenic, highly 287

toxic or radioactive products or live viral or bacterial materials). The modification may include 288

positively or negatively pressurized airlocks that prevent the hazardous material from contaminating 289

surrounding areas. Decontamination of facilities (e.g. the cleanrooms and the heating, ventilation, 290

and air conditioning (HVAC) systems) and the treatment of air leaving a clean area, may be 291

necessary for some operations. Where containment requires air to flow into a critical zone, the 292

source of the air should be from an area of the same grade. 293

294

4.15 Airflow patterns within cleanrooms and zones should be visualised to demonstrate that there is 295

no ingress from lower grade to higher grade areas and that air does not travel from less clean areas 296

(such as the floor) or over operators or equipment that may transfer contaminant to the higher grade 297

areas. Where air movement is shown to be a risk to the clean area or critical zone, corrective actions, 298

such as design improvement, should be implemented. Airflow pattern studies should be performed 299

both at rest and in operation (e.g. simulating operator interventions). Video recordings of the airflow 300

patterns should be retained. The outcome of the air visualisation studies should be considered when 301

establishing the facility's environmental monitoring program. 302

303

4.16 Indicators of pressure differences should be fitted between cleanrooms and/or isolators. Set-304

points and the criticality of pressure differentials should be documented within the CCS. Pressure 305

differentials identified as critical should be continuously monitored and recorded. A warning system 306

should be in place to instantly indicate and warn operators of any failure in the air supply or 307

reduction of pressure differentials (below set limits for those identified as critical). The warning 308

signal should not be overridden without assessment and a procedure should be available to outline 309

the steps to be taken when a warning signal is given. Where alarm delays are set, these should be 310

assessed and justified within the CCS. Other pressure differentials should be monitored and recorded 311

at regular intervals. 312

313

4.17 Facilities should be designed to permit observation of production activities from outside the 314

Grade A zone and Grade B area (e.g. through the provision of windows or remote cameras with a 315

full view of the area and processes to allow observation and supervision without entry). This 316

requirement should be considered when designing new facilities or during refurbishment of existing 317

facilities. 318

319

Barrier Technologies 320

321

4.18 Isolator or RABS technologies, and the associated processes, should be designed to provide 322

protection of the Grade A environment. The entry of materials during processing (and after 323

decontamination) should be minimized and preferably supported by rapid transfer technologies or 324

transfer isolators. 325

326

4.19 The design of the RABS or isolator should take into account all critical factors associated with 327

these technologies including the quality of the air inside and the background environment, the 328

materials and component transfer, the decontamination and/or sterilization processes, the risk factors 329

associated with the manufacturing operations and the operations conducted within the critical zone. 330

331

4.20 The critical zone of the RABS or open isolator used for aseptic processes should meet Grade A 332

requirements with unidirectional airflow. In closed isolator systems where airflow may not be 333

unidirectional, it should provide Grade A conditions and be demonstrated to provide adequate 334

protection for exposed products during processing. The design of the RABS and open isolators should 335

ensure a positive airflow from the critical zones to the supporting background environment; (unless 336

containment is required in which case localized air extraction is required to prevent contamination 337

transfer to the surrounding room). Negative pressure isolators should only be used when containment 338

of the product is considered essential and risk control measures are applied to ensure the critical zone 339

is not compromised. 340

341

4.21 For RABS used for aseptic processing, the background environment should meet at least Grade 342

B. The background environment for open isolators should meet Grade C or D, based on a risk 343

assessment. Airflow studies should be performed to demonstrate the absence of air ingress during 344

interventions, such as door openings. 345

346

4.22 The background environment of a closed isolator should correspond to a minimum of Grade D. 347

The disinfection/decontamination programme should be included as a key consideration when 348

performing the risk assessment for the CCS of an isolator. Where additional process risks are 349

identified, a higher grade of background should be considered. The decision as to the supporting 350

background environment should be documented in the CCS. 351

352

4.23 The materials used for glove systems (for both RABS and isolators), as well as other parts of an 353

isolator, should be demonstrated to have good mechanical and chemical resistance. Integrity testing of 354

the barrier systems, and leak testing of the glove system and the isolator should be performed using a 355

methodology demonstrated to be suitable for the task and criticality. The testing should be performed 356

at defined periods, at a minimum at the beginning and end of each batch, and should include a visual 357

inspection following any intervention that may affect the integrity of the system. For single unit batch 358

sizes, integrity may be verified based on other criteria, such as the beginning and end of each 359

manufacturing session. RABS gloves used in Grade A zone should be sterilized before installation 360

and sterilized (or effectively decontaminated by a validated method which achieves the same 361

objective) prior to each manufacturing campaign. The frequency of glove replacement should be 362

defined within the CCS. 363

364

4.24 For RABS and isolator systems, decontamination methods should be validated and controlled 365

within defined cycle parameters. The cleaning process prior to the disinfection step is essential; any 366

residues that remain may inhibit the effectiveness of the decontamination process: 367

368

i. For isolators, the decontamination process should be automated and should include a 369

sporicidal agent in a suitable form (e.g. gaseous, aerosolized or vaporized form) to ensure 370

thorough microbial decontamination of its interior. Decontamination methods (cleaning and 371

sporicidal disinfection) should render the interior surfaces and critical zone of the isolator free 372

of viable microorganisms. 373

374

ii. For RABS systems, the disinfection should include the routine application of a sporicidal 375

agent using a method that has been validated and demonstrated to robustly disinfect the 376

interior and ensure a suitable environment for aseptic processing. 377

378

Evidence should also be available to demonstrate that the agent used does not have adverse impact on 379

the product produced within the RABS or isolator. The holding time before use of these systems 380

should be validated. 381

382

Cleanroom and clean air equipment qualification 383

384

4.25 Cleanrooms and clean air equipment such as unidirectional airflow units (UDAFs), 385

RABS and isolators, used for the manufacture of sterile products, should be qualified and 386

classified according to the required characteristics of the environment. Each manufacturing 387

operation requires an appropriate environmental cleanliness level in the operational state in 388

order to minimize the risk of particulate or microbial contamination of the product or materials 389

being handled. 390

391

4.26 Cleanrooms and clean air equipment should be qualified using methodology in accordance with 392

the requirements of Annex 15. Cleanroom qualification (including classification) should be clearly 393

differentiated from operational environmental monitoring. 394

395

4.27 Cleanroom Qualification is the overall process of assessing the level of compliance of a 396

classified cleanroom or clean air equipment with its intended use. As part of the qualification 397

requirements of Annex 15, the qualification of cleanrooms and clean air equipment should include 398

(where relevant to the design/operation of the installation): 399

400

i. Installed filter leakage and integrity testing. 401

402

ii. Airflow measurement - Volume and velocity. 403

404

iii. Air pressure difference measurement. 405

406

iv. Airflow direction and visualisation. 407

408

v. Microbial airborne and surface contamination. 409

410

vi. Temperature measurement. 411

412

vii. Relative humidity measurement. 413

414

viii. Recovery testing. 415

416

ix. Containment leak testing. 417

418

4.28 Cleanroom classification is part of a cleanroom qualification and is a method of assessing the 419

level of air cleanliness against a specification for a cleanroom or clean air equipment by measuring 420

the non-viable airborne particulate concentration. Reference for the classification of the cleanrooms 421

and clean air equipment can be found in the ISO 14644 series of standards. 422

423

4.29 For cleanroom classification, the airborne particulates equal to or greater than 0.5 and 5 µm 424

should be measured. For Grade A zone and Grade B at rest, classification should include 425

measurement of particles equal to or greater than 0.5 µm; however, measurement using a second, 426

larger particle size, e.g. 1 µm in accordance with ISO 14644 may be considered. This measurement 427

should be performed both at rest and in operation. The maximum permitted airborne particulate 428

concentration for each grade is given in Table 1. 429

430

431

Table 1: Maximum permitted airborne particulate concentration during classification 432

433

Grade

Maximum limits for particulates

≥ 0.5 µm/m

Maximum limits for particulates

≥ 5 µm/m

at rest

in operation

at rest

in operation

3 520

Not applicable

3 520

352 000

Not applicable

2 900

352 000

3 520 000

2 900

29 000

3 520

000

Not defined

(

)

29 000

Not de

fined

(a)

434

435

(a)

For Grade D, in operation limits are not defined. The company should establish in operation 436

limits based on a risk assessment and historical data where applicable. 437

438

4.30 For classification of the cleanroom, the minimum number of sampling locations and their 439

positioning can be found in ISO 14644 Part 1. In addition, for the aseptic processing room and the 440

background environment (Grade A zone and Grade B area, respectively), sample locations should also 441

consider all critical processing zones such as the point of fill and stopper bowls. Critical processing 442

locations should be based on a documented risk assessment and knowledge of the process and 443

operations to be performed in the area. 444

445

4.31 Clean room classification should be carried out in the “at rest” and “in operation” states. 446

447

i. The definition of “at rest” state is the condition whereby the installation of all the utilities is 448

complete including any functioning HVAC, with the main manufacturing equipment installed 449

as specified and standing by for operation, without personnel in the room. 450

451

ii. The definition of “in operation” state is the condition where the installation of the cleanroom 452

is complete, the HVAC system fully operational, equipment installed and functioning in the 453

manufacturer’s defined operating mode with the maximum number of personnel present 454

performing or simulating routine operational work. In operation classification may be 455

performed during simulated operations or during aseptic process simulations (where worst 456

case simulation is required). 457

458

iii. The particulate limits given in Table 1 above for the “at rest” state should be achieved after 459

a “clean up” period on completion of operations. The "clean up" period should be 460

determined during the classification of the rooms (guidance value of 15 to 20 minutes). 461

462

4.32 The speed of air supplied by unidirectional airflow systems should be clearly justified in the 463

qualification protocol including the location for air speed measurement. Air speed should be designed, 464

measured and maintained to ensure that appropriate unidirectional air movement provides protection 465

of the product and open components at the working height (e.g. where high risk operations and 466

product and/or components are exposed). Unidirectional airflow systems should provide a 467

homogeneous air speed in a range of 0.36 – 0.54 m/s (guidance value) at the working position, unless 468

otherwise scientifically justified in the CCS. Airflow visualization studies should correlate with the air 469

speed measurement. 470

4.33 The microbial concentration of the cleanrooms should be determined as part of the cleanroom 471

qualification. The number of sampling locations should be based on a documented risk assessment, 472

including the results of the classification, air visualization studies and knowledge of the process and 473

operations to be performed in the area. The maximum limits for microbial contamination during 474

qualification for each grade are given in Table 2. Qualification should include both at rest and in 475

operation states. 476

477

Table 2: Limits for microbial contamination during qualification 478

Grade

Air sample

cfu/m

Settle plates

(diameter 90 mm)

cfu/4 hours

(a)

Contact plates

(diameter 55

mm) cfu/plate

(b)

No growth

(b)

B 10 5 5

C 100 50 25

D 200 100 50

(a) Settle plates should be exposed for the duration of operations and changed as required after 4 479

hours. Exposure time should be based on recovery studies and should not allow desiccation of the 480

media used. 481

482

(b) It should be noted that for Grade A, the expected result should be no growth. 483

Note 1: All methods indicated for a specific Grade in the table should be used for qualifying the 484

area of that specific Grade. If one of the methods is not used, or alternative methods are used, the 485

approach taken should be appropriately justified. 486

Note 2: Limits are applied using cfu throughout the document. If different or new technologies 487

are used that present results in a manner different from cfu, the manufacturer should scientifically 488

justify the limits applied and where possible correlate them to cfu. 489

Note 3: For qualification of personnel gowning, the limits given for contact plates and glove prints in 490

Table 7 should apply. 491

Note 4: Sampling methods should not pose a risk of contamination to the manufacturing operations. 492

493

4.34 The requalification of cleanrooms and clean air equipment should be carried out periodically 494

following defined procedures. The requirement for requalification of cleanroom areas is as follows: 495

496

Table 3: Minimum test requirements for the requalification of cleanrooms 497

Grade

Determination

of the

concentration

of airborne

viable and non-

viable particles

Integrity Test

of Terminal

Filters

Airflow

volume

measurement

Verification of

air pressure

difference

between rooms

Air

Velocity

test

A Yes Yes Yes Yes Yes

B Yes Yes Yes Yes *

C Yes Yes Yes Yes *

D Yes Yes Yes Yes *

* performed according to a risk assessment documented as part of the CCS. However, required 498

for filling zones (e.g. when filling terminally sterilised products) and background to Grade A 499

RABS. 500

For Grade A & B areas, the maximum time interval for requalification is 6 months. 501

For Grade C & D areas, the maximum time interval for requalification is 12 months. 502

Appropriate requalification consisting of at least the above tests should also be carried out following 503

completion of remedial action implemented to rectify an out-of-compliance equipment or facility 504

condition or after changes to equipment, facility or processes. The significance of a change should be 505

determined through the change management process. Examples of changes to be considered include 506

but are not limited to the following: 507

508

i. Change in the operational use of the cleanroom, or of the operational setting parameters of 509

the HVAC system. 510

ii. Interruption of air movement which affects the operation of the installation. 511

iii. Special maintenance which affects the operation of the installation (e.g. change of final 512

filters). 513

4.35 Other characteristics, such as temperature and relative humidity, should be controlled within 514

ranges that align with product/processing requirements and support maintenance of defined 515

cleanliness standards (e.g. Grade A or B). 516

517

Disinfection

518

519

4.36 The disinfection of cleanrooms is particularly important. They should be cleaned and disinfected 520

thoroughly in accordance with a written programme. For disinfection to be effective, prior cleaning to 521

remove surface contamination should be performed. More than one type of disinfecting agent should 522

be employed to ensure that where they have different modes of action and their combined usage is 523

effective against all bacteria and fungi. Disinfection should include the periodic use of a sporicidal 524

agent. Monitoring should be undertaken regularly in order to assess the effectiveness of the 525

disinfection program and to detect changes in types of microbial flora (e.g. organisms resistant to the 526

disinfection regime currently in use). Cleaning programs should effectively remove disinfectant 527

residues. 528

529

4.37 The disinfection process should be validated. Validation studies should demonstrate the 530

suitability and effectiveness of disinfectants in the specific manner in which they are used and should 531

support the in-use expiry periods of prepared solutions. 532

533

4.38 Disinfectants and detergents used in Grade A zone and Grade B areas should be sterile

prior to 534

use (disinfectants used in Grade C and D may also be required to be sterile). Where the disinfectants 535

and detergents are made up by the sterile product manufacturer, they should be monitored for 536

microbial contamination. Dilutions

should be kept in previously cleaned containers and should only 537

be stored for defined periods. If the disinfectants and detergents are supplied “ready-made” then results 538

from certificates of analysis or conformance can be accepted subject to successful completion of the 539

appropriate vendor qualification. 540

541

4.39 Fumigation or vapour disinfection (e.g. Vapour-phased Hydrogen Peroxide) of cleanrooms and 542

associated surfaces may be useful for reducing microbial contamination in

inaccessible places. 543

544

5 Equipment 545

546

5.1 A written, detailed description of the equipment design should be available (including process and 547

instrumentation diagrams as appropriate). This should form part of the initial qualification package 548

and be kept up to date as part of the ongoing review of the CCS. 549

550

5.2 Equipment monitoring requirements should be defined in “user requirements specifications” and 551

during early stages of development, and confirmed during qualification. Process and equipment alarm 552

events should be reviewed and approved and evaluated for trends. The frequency at which alarms are 553

assessed should be based on their criticality (with critical alarms reviewed immediately). 554

555

5.3 As far as practicable, equipment, fittings and services should be designed and installed so that 556

operations, maintenance, and repairs can be performed outside the cleanroom. If maintenance has to 557

be performed in the cleanroom, and the required standards of cleanliness and/or asepsis cannot be 558

maintained, then precautions such as restricting access to the work area to specified personnel, 559

generation of clearly defined work protocols and maintenance procedures should be considered. 560

Cleaning, additional disinfection and additional environmental monitoring should also be considered. 561

If sterilization of equipment is required, it should be carried out, wherever possible, after complete 562

reassembly. 563

564

5.4 The cleaning process should be validated to: 565

566

i. Remove any residue or debris that would detrimentally impact the effectiveness of the 567

disinfecting agent used. 568

ii. Minimize chemical, microbial and particulate contamination of the product during the process 569

and prior to disinfection. 570

571

5.5 Direct and indirect contact parts should be sterilized. Direct contact parts are those that the 572

product passes through, such as filling needles or pumps. Indirect product contact parts are 573

equipment parts that come into contact with sterilized critical items and components. 574

575

5.6 All equipment such as sterilizers, air handling systems (including air filtration) and water 576

systems should be subject to qualification, monitoring and planned maintenance. Upon completion 577

of maintenance, their return to use should be approved. 578

579

5.7 Where unplanned maintenance of equipment critical to the sterility of the product is to be carried 580

out, an assessment of the potential impact to the sterility of the product should be performed and 581

recorded. 582

583

5.8 A conveyor belt should not pass through a partition between a Grade A or B area and a

584

processing area of lower air cleanliness, unless the belt itself is continually sterilized (e.g. in a

585

sterilizing tunnel). 586

587

5.9 Particle counters, including sampling tubing, should be qualified. The tubing length should be no 588

greater than 1 meter with a minimum number of bends and bend radius should be greater than 15 cm. 589

Portable particle counters with a short length of sample tubing should be used for classification 590

purpose. Isokinetic sampling heads should be used in unidirectional airflow systems and should be 591

positioned as close as possible to sample air representative of the critical location. 592

593

6 Utilities 594

595

6.1 The nature and extent of controls applied to utility systems should be commensurate with the risk 596

to product quality associated with the utility. The impact should be determined via a risk assessment 597

documented as part of the CCS. 598

599

6.2 In general higher risk utilities are those that: 600

601

i. Directly contact product e.g. water for washing and rinsing, gases and steam for 602

sterilization. 603

604

ii. Contact materials that will ultimately become part of the product. 605

606

iii. Contact surfaces that come into contact with the product. 607

608

iv. Otherwise directly impact the product. 609

610

6.3 Utilities should be designed, installed, operated, maintained and monitored in a manner to ensure 611

that the utility functions as expected. 612

613

6.4 Results for critical parameters and critical quality attributes of high risk utilities should be subject 614

to regular trend analysis to ensure that system capabilities remain appropriate. 615

616

6.5 Records of utility installation should be maintained throughout the system’s life-cycle. Such 617

records should include current drawings and schematic diagrams, construction material lists and 618

specifications. Typically, important information includes attributes such as: 619

620

i. Pipeline flow direction, slopes, diameter and length. 621

622

ii. Tank and vessel details. 623

624

iii. Valves, filters, drains, sampling and user points. 625

626

6.6 Pipes, ducts and other utilities should not be present in cleanrooms. If unavoidable, then they 627

should be installed so that they do not create recesses, unsealed openings and surfaces which are 628

difficult to clean. Installation should allow cleaning and disinfection of outer surface of the pipes. 629

630

Water systems 631

632

6.7 Water treatment plant and distribution systems should be designed, constructed and maintained to 633

minimize the risk of particulates, microbial contamination/proliferation and pyrogens (e.g. sloping of 634

piping to provide complete drainage and the avoidance of dead legs), and prevent the formation of 635

biofilms to ensure a reliable source of water of an appropriate quality. Where filters are included in 636

the system, special attention should be given to the monitoring and maintenance of these filters. Water 637

produced should comply with the current monograph of the relevant Pharmacopeia. 638

639

6.8 Water systems should be qualified to maintain the appropriate levels of physical, chemical and 640

microbial control, taking seasonal variation into account. 641

642

6.9 Water flow should remain turbulent through the pipes to minimize the risk of microbial adhesion, 643

and subsequent biofilm formation. 644

645

6.10 Water for injections (WFI) should be produced from water meeting specifications that have been 646

defined during the qualification process, stored and distributed in a manner which minimizes the risk 647

of microbial growth (for example by constant circulation at a temperature above 70°C). Where the 648

WFI is produced by methods other than distillation, further techniques such as nanofiltration and 649

ultra-filtration as well as electrodeionization (EDI) should be considered in conjunction with reverse 650

osmosis (RO) membranes. 651

652

6.11 Where WFI storage tanks are equipped with hydrophobic bacteria retentive vent filters, the 653

filters should be sterilized and the integrity of the filter tested before installation and after removal 654

following use. 655

656

6.12 To minimize the risk of biofilm formation, sterilization or disinfection or regeneration of water 657

systems should be carried out according to a predetermined schedule and when microbial counts 658

exceed action limits. Disinfection of a water system with chemicals should be followed by a 659

validated rinsing/flushing procedure. Water should be tested after disinfection/regeneration. The 660

results should be approved before the water system is returned to use. 661

662

6.13 Regular ongoing chemical and microbial monitoring of water systems should be performed. 663

Alert levels should be based on the qualification or a review of ongoing monitoring data that will 664

identify an adverse trend in system performance. Sampling programs should reflect the requirements 665

of the CCS and include: 666

667

i. All points of use, at a specified interval, to ensure that representative water samples are 668

obtained for analysis on a regular basis. 669

670

ii. Potential worst case sampling locations. 671

672

iii. A sample from the point at the end of the distribution loop each day that the water is used. 673

674

6.14 Breaches of alert levels should be documented and reviewed, and include investigation of 675

system trends to determine whether the breach is a single (isolated) event or if results are indicative 676

of loss of control or system deterioration. Each breach of action limits should be investigated to 677

determine the root cause of the issue and any impact on the quality of products and manufacturing 678

processes as a result of the potential use of the water. 679

680

6.15 WFI systems should include continuous monitoring systems such as Total Organic Carbon 681

(TOC) and conductivity, (unless justified otherwise) as these may give a better indication of overall 682

system performance than discrete sampling. Sensor locations should be based on risk and the 683

outcome of qualification. 684

685

Steam used as a direct sterilizing agent 686

687

6.16 Feed water to a pure steam (clean steam) generator should be appropriately purified. Pure steam 688

generators should be designed, qualified and operated in a manner to ensure that the quality of steam 689

produced meets defined chemical and endotoxin levels. 690

691

6.17 Steam used as a direct sterilizing agent should be of suitable quality and should not contain 692

additives at a level which could cause contamination of product or equipment. For a pure steam 693

generator supplying pure steam used for the direct sterilization of materials or product-contact 694

surfaces (e.g. porous hard-good autoclave loads), steam condensate should meet the current 695

monograph for WFI of the relevant Pharmacopeia. A suitable sampling schedule should be in place 696

to ensure that representative pure steam samples are obtained for analysis on a regular basis. Other 697

aspects of the quality of pure steam used for sterilization should be assessed periodically against 698

validated parameters. These parameters should include the following: non-condensable gases, 699

dryness value (dryness fraction) and superheat. 700

701

Gases and vacuum systems 702

703

6.18 Gases that come in direct contact with the product/primary container surfaces should be of 704

appropriate chemical, particulate and microbial quality. All relevant parameters, including oil and 705

water content, should be specified, taking into account the use and type of the gas, the design of the 706

gas generation system and, where applicable, comply with the appropriate Pharmacopoeia 707

monographs. 708

709

6.19 Gases used in aseptic processes should be filtered through a sterilizing filter (with a nominal pore 710

size of a maximum of 0.22 µm) at the point of use. Where the filter is used on a batch basis (e.g. for 711

filtration of gas used for overlay of aseptically filled products) or as product vessel vent filter, then the 712

filter should be integrity tested and the results included as part of the batch certification process. Any 713

transfer pipework or tubing that is located after the final sterilizing filter should be sterilized. When 714

gases are used in the process, microbial monitoring of the gas should be performed periodically at the 715

point of use. 716

717

6.20 Where backflow from vacuum or pressure systems poses a potential risk to the product, there 718

should be mechanism(s) to prevent backflow when the vacuum or pressure system is shut off. 719

720

Heating and cooling and hydraulic systems 721

722

6.21 Major items of equipment associated with hydraulic, heating and cooling systems, e.g. such as 723

those associated with Blow-Fill-Seal equipment should, where possible, be located outside the filling 724

room. Where they are located inside the filling room there should be appropriate controls to contain 725

any spillage and/or cross contamination associated with the hydraulic system fluids. Where possible, 726

the system should be at a lower pressure than the processed fluid. 727

728

6.22 Any leaks from these systems that would present a risk to the product should be detectable (i.e. 729

an indication system for leakage). 730

731

6.23 For both vacuum and cooling systems there should be periodic cleaning/disinfection as 732

determined in the CCS. 733

734

7 Personnel 735

7.1 The manufacturer should ensure that there are sufficient appropriate personnel, suitably qualified, 736

trained and experienced in the manufacture and testing of sterile products, and any of the specific 737

manufacturing technologies used in the site’s manufacturing operations, to ensure compliance with 738

GMP applicable to the manufacture and handling of sterile products. 739

740

7.2 Only the minimum number of personnel required should be present in cleanrooms. The 741

maximum number of operators in cleanrooms should be determined, documented and validated 742

during activities such as initial qualification and aseptic process simulations, so as not to 743

compromise sterility assurance. This is particularly important during aseptic processing. 744

745

7.3 Non-essential processes such as product inspection and in process testing should be conducted 746

outside the clean areas wherever possible. 747

748

7.4 All personnel including those performing cleaning, maintenance, monitoring and those that 749

access cleanrooms should receive regular training, gowning qualification and assessment in 750

disciplines relevant to the correct manufacture of sterile products. This training should include the 751

basic elements of microbiology, hygiene, with a specific focus on cleanroom practices, 752

contamination control, aseptic techniques and the protection of sterile products (for those operators 753

entering the Grade B cleanrooms and/or intervening into the Grade A zone) and the potential safety 754

implications to the patient if product is not sterile. The level of training should be based on the 755

criticality of the function and area in which the personnel are working. 756

757

7.5 The personnel working in a Grade A zone and Grade B areas should be trained for aseptic 758

gowning and aseptic practices. Compliance with aseptic gowning procedures should be assessed and 759

confirmed, periodically reassessed at least annually and should involve both visual and microbial 760

assessment (using monitoring locations such as hands, arms, chest and forehead. Refer to paragraph 761

9.30 for the expected limits). The unsupervised access to Grade A zone and Grade B areas where 762

aseptic operations are or will be conducted should be restricted to appropriately qualified personnel, 763

who have passed the gowning assessment and have participated in a successful aseptic process 764

simulation (APS). 765

766

7.6 Unqualified personnel (e.g. building and maintenance contractors and regulatory inspectors) 767

should not enter Grade B cleanrooms or Grade A zones in operation. If needed in exceptional cases, 768

manufacturers should establish written procedures outlining the process by which unqualified 769

personnel are brought into the Grade B and A areas. Access by these persons should be assessed and 770

recorded in accordance with the PQS. An authorized person from the manufacturer should supervise 771

the unqualified personnel during their activities and should assess the impact of these activities on 772

the cleanliness of the area. 773

774

7.7 There should be systems in place for disqualification of personnel from entry into cleanrooms 775

based on aspects including ongoing assessment and/or identification of an adverse trend from the 776

personnel monitoring program and/or after participation in a failed APS. Once disqualified, 777

retraining and requalification should be completed before permitting the operator to have any further 778

involvement in aseptic practices. For operators entering Grade B cleanrooms or performing 779

intervention into Grade A zone, this requalification should include consideration of participation in a 780

successful APS. 781

782

7.8 High standards of personal hygiene and cleanliness are essential to prevent excessive shedding or 783

increased risk of introduction of microbial contamination. Personnel involved in the manufacture of 784

sterile products should be instructed to report any specific health conditions or ailments which may 785

cause the shedding of abnormal numbers or types of contaminants and therefore preclude cleanroom 786

access. Health conditions and actions to be taken with regard to personnel who could be introducing 787

an undue microbial hazard should be provided by the designated competent person and described in 788

procedures. 789

790

7.9 Staff who have been engaged in the processing of human or animal tissue materials or of cultures 791

of micro-organisms, other than those used in the current manufacturing process, or any activities that 792

may have a negative impact to quality (e.g. microbial contamination), should not enter clean areas 793

unless clearly defined and effective decontamination and entry procedures have been followed. 794

795

7.10 Wristwatches, make-up, jewellery, other personal items such as mobile phones and any other 796

non-essential items should not be allowed in clean areas. Electronic devices used in cleanrooms, e.g. 797

mobile phones and tablets, that are supplied by the company solely for use in the cleanrooms, may 798

be acceptable if suitably designed to permit cleaning and disinfection commensurate with the Grade 799

in which they are used. The use and disinfection of such equipment should be included in the CCS. 800

801

7.11 Cleanroom gowning and hand washing should follow a written procedure designed to minimize 802

contamination of cleanroom clothing and/or the transfer of contaminants to the clean areas. 803

804

7.12 The clothing and its quality should be appropriate for the process and the grade of the

805

working area. It should be worn in such a way as to protect the product from contamination. When the 806

type of clothing chosen needs to provide the operator protection from the product, it should not 807

compromise the protection of the product from contamination. Garments should be visually checked 808

for cleanliness and integrity immediately prior to gowning and prior to entry to the cleanroom. Gown 809

integrity should also be checked upon exit. For sterilized or effectively decontaminated garments and 810

eye coverings, particular attention should be taken to ensure they have been processed, are within 811

their specified hold time and that the packaging is visually inspected to ensure it is integral before use. 812

Reusable garments (including eye coverings) should be replaced if damage is identified or at a set 813

frequency that is determined during qualification studies. . Damage to garments may not be identified 814

by visual inspection alone, so the qualification should consider any necessary garment testing 815

requirements. 816

817

7.13 Clothing should be chosen to prevent shedding due to operators moving excessively (when 818

cold) or sweating (when hot). 819

820

7.14 The description of clothing required for each grade is given below: 821

822

i. Grade A / B: Dedicated garments to be worn under a sterilized suit. Sterile headgear should 823

enclose all hair (including facial hair) and where separate from the rest of the gown, it 824

should be tucked into the neck of the sterile suit. A sterile face mask and sterile eye 825

coverings (e.g. goggles) should be worn to cover and enclose all facial skin and prevent the 826

shedding of droplets and particulates. Appropriate sterilized, non-powdered, rubber or 827

plastic gloves and sterilized footwear (such as overboots) should be worn. Trouser-legs 828

should be tucked inside the footwear and garment sleeves into the gloves. The protective 829

clothing should minimize shedding of fibres or particulate matter and retain particulates 830

shed by the body. Garments should be packed and folded in such a way as to allow operators 831

to gown without contacting the outer surface of the garment. 832

833

ii. Grade C: Hair, beards and moustaches should be covered. A single or two-piece trouser suit 834

gathered at the wrists and with high neck and appropriately disinfected shoes or overshoes 835

should be worn. They should minimize the shedding of fibres and particulate

matter. 836

837

iii. Grade D: Hair, beards and moustaches should be covered. A general protective suit

and 838

appropriately disinfected shoes or overshoes should be worn. Appropriate measures should 839

taken to avoid any ingress of contaminants from outside the clean area. 840

841

iv. Gloves should be worn in Grade C and D areas when performing activities considered to be a 842

contamination risk as defined by the CCS. 843

844

7.15 Outdoor clothing (other than personal underwear) should not be brought into changing rooms 845

leading directly to Grade B and C

clean

rooms. Facility suits, covering the full length of the arms 846

and the legs, and socks covering the feet, should be worn before entry to change rooms for Grades B 847

and C. Facility suits and socks should not present a risk of contamination to the gowning area or 848

processes. 849

850

7.16 Every operator entering Grade B or A areas should gown into clean, sterilized protective 851

garments (including eye coverings and masks) of an appropriate size at each entry. The maximum 852

duration of each garment use should be defined as part of the garment qualification. 853

854

7.17 Garments and gloves should be changed immediately if they become damaged and present any 855

risk of product contamination. Gloves should be regularly disinfected during operations. 856

857

7.18 Clean area clothing should be cleaned in a dedicated laundry facility using a qualified process 858

ensuring that the clothing is not damaged and/or contaminated by fibres and particles during the 859

laundry process. Inappropriate handling and use of clothing will damage fibres and may increase the 860

risk of shedding of particles. After washing and before packing, garments should be visually 861

inspected for damage. The garment management processes should be evaluated and determined as 862

part of the garment qualification program. 863

864

7.19 Activities in clean areas that are not critical to the production processes should be kept to a 865

minimum, especially when aseptic operations are in progress. Movement of personnel should be 866

slow, controlled and methodical to avoid

excessive shedding of particulates and organisms due to 867

over-vigorous activity. Operators performing aseptic operations should adhere to aseptic technique 868

at all times to prevent changes in air currents that introduce air of lower quality into the critical zone. 869

Movement adjacent to the critical zone should be restricted and the obstruction of the path of the 870

unidirectional (first air) airflow should be avoided. Airflow visualisation studies should be 871

considered as part of the operator’s training programme. 872

873

874

8 Production and Specific Technologies 875

876

Terminally sterilized products 877

878

8.1 Preparation of components and materials should be performed in at least a Grade D 879

cleanroom in order to limit the risk of microbial, pyrogen and particulate contamination, so that the 880

product is suitable for sterilization. Where the product is at a high or unusual risk of microbial

881

contamination (e.g. the product actively supports microbial growth, the product must

be held for 882

long periods before filling or the product is not processed mostly in closed

vessels), then 883

preparation should be carried out in a Grade C environment. Preparation of ointments, creams, 884

suspensions and emulsions should be carried out in a Grade C environment before

terminal 885

sterilization. 886

887

8.2 Primary packaging containers and components should be cleaned using validated processes to 888

ensure that particulate, pyrogen and bioburden contamination is appropriately controlled. 889

890

8.3 Filling of products for terminal sterilization should be carried out in at least a Grade C

891

environment. 892

893

8.4 Where the product is at an unusual risk of contamination from the environment because, for 894

example, the filling operation is slow, the containers are wide necked or are necessarily exposed for 895

more than a few seconds before closing, then the product should be filled in a Grade A zone with at least 896

a Grade C background. 897

898

8.5 Processing of the bulk solution should include a filtration step with a microorganism retaining 899

filter, where possible, to reduce bioburden levels and particulates prior to filling into the final 900

product containers and there should be a maximum permissible time between preparation and filling. 901

902

8.6 Examples of operations to be carried out in the various grades are given in Table 4.

903

904

Table 4: Examples of operations and grades for terminally sterilized preparation and 905

processing operations 906

A Filling of products, when unusually at risk.

C Preparation of solutions, when unusually at risk. Filling of products.

D Preparation of solutions and components for subsequent filling.

907

Aseptic preparation and processing 908

909

8.7 Aseptic preparation and processing is the handling of sterile product, containers and/or devices in 910

a controlled environment in which the air supply, materials and personnel are regulated to prevent 911

microbial, pyrogenic and particulate contamination. 912

913

8.8 The aseptic process should be clearly defined. The risks associated with the aseptic process, and 914

any associated requirements, should be identified, assessed and appropriately controlled. The site’s 915

CCS should clearly define the acceptance criteria for these controls, requirements for monitoring and 916

the review of their effectiveness. Methods and procedures to control these risks should be described 917

and implemented. Accepted residual risks should be formally documented. 918

919

8.9 Precautions to minimize microbial, pyrogenic and particulate contamination should be taken, 920

as per the site’s CCS, during the preparation of the aseptic environment, during all processing stages

921

(

including the stages before and after bulk product sterilization), and until the product is sealed in its 922

final container. The presence of materials liable to generate particulates and fibres should be minimized 923

in cleanrooms. 924

925

8.10 Where possible, the use of equipment such as RABS, isolators or other systems, should be 926

considered in order to reduce the need for critical interventions into the Grade A zone and to minimize 927

the risk of contamination. Robotics and automation of processes can also be considered to eliminate 928

direct human critical interventions (e.g. dry heat tunnel, automated lyophilizer loading, sterilization in 929

place). 930

931

8.11 Examples of operations to be carried out in the various environmental grades are given in the 932

Table 5. 933

934

Table 5: Examples of operations and grades for aseptic preparation and processing operations 935

936

Grade A

Critical zone for

- Aseptic assembly of filling equipment.

- Connections made under aseptic conditions (where sterilize

d product contact

surfaces are exposed) that are post the final sterilizing filter. T

hese

connections should be sterilized by steam-in-place whenever feasible.

- Aseptic compounding and mixing.

- Replenishment of sterile bulk product, containers and closures.

- Removal and cooling of unprotected (e.g. with no packaging) items

from

sterilizers.

- Staging and conveying of sterile primary packaging components.

- Aseptic filling, sealing of containers such as ampoules, vial closure,

transfer

of open or partially stoppered vials.

- Loading of a lyophilizer.

Grade B

Background support for the Grade A zone (when not in an isolator).

- Transport, while protected from the surrounding environment,

of equipment,

components and ancillary items for introduction into the Grade A zone.

Grade C

- Preparation of solutions to be filtered including weighing.

Grade D

- Cleaning of equipment.

- Handling of components, equipment and accessories after washing.

- Assembly

of cleaned components, equipment and accessories prior to

sterilization.

- Assembly of closed and sterilized SUS using intrinsic aseptic connectors.

937

8.12 For sterile products that cannot be filtered, the following should be considered: 938

939

i. All product and component contact equipment should be sterilized prior to use. 940

941

ii. All raw materials should be sterilized and aseptically added or subsequently sterilized by 942

filtration. 943

944

iii. Bulk solutions should be sterilized by a validated process, e.g. by heat, chemical sterilization 945

or via sterile filtration. 946

947

iv. All materials added to the sterile bulk product should be sterilized prior to addition. 948

949

8.13 The unwrapping, assembly and preparation of sterilized equipment, components and ancillary 950

items and the preparation and filling of the sterile product should be treated as an aseptic process and 951

performed in a Grade A zone with a Grade B background. Where an isolator or RABS is used, the 952

background should be in accordance with paragraphs 4.21 & 4.22. 953

954

8.14 Preparation and filling of sterile products such as ointments, creams, suspensions and 955

emulsions should be

performed in a Grade A zone with a Grade B background when the product and 956

components are exposed to the environment and

the product

is not subsequently filtered (via a 957

sterilizing filter) or terminally sterilized. Where an isolator or RABS is used, the background should 958

be in accordance with paragraphs 4.21 & 4.22. 959

960

8.15 Aseptic connections should be performed in a Grade A zone with a Grade B background unless 961

subsequently sterilized in place or conducted with validated intrinsic sterile connection devices that 962

minimize any potential contamination from the immediate environment. Where an isolator or RABS 963

is used, the background should be in accordance with paragraphs 4.21 & 4.22. Aseptic connections 964

should be appropriately assessed and their effectiveness verified. For requirements regarding intrinsic 965

sterile connection devices refer to paragraph 8.120. 966

967

8.16 Aseptic manipulations (including non-intrinsic aseptic connections) should be minimized 968

through the use of engineering design solutions such as preassembled and sterilized equipment. 969

Whenever feasible, product contact piping and equipment should be pre-assembled, and sterilized in 970

place. 971

972

8.17 There should be an authorized list of allowed interventions, both inherent and corrective, that 973

may occur during production. The procedures listing the types of inherent and corrective 974

interventions, and how to perform them, should be updated, as necessary to ensure consistency with 975

the actual manufacturing activities. In the event that an unauthorized intervention is required, details 976

of the intervention conducted should be recorded and fully assessed under the manufacturer's PQS. 977

978

8.18 The duration of each aspect of aseptic preparation and processing should be limited to a defined 979

and validated maximum time, including: 980

981

i. The holding time between equipment, component, and container cleaning, drying and 982

sterilization. 983

984

ii. The holding time for sterilized equipment, components, and containers before use and 985

during filling/assembly. 986

987

iii. The holding time for a decontaminated environment, such as the RABS and isolator before 988

and during filling /assembly. 989

990

iv. The time between the start of the preparation of a product and its sterilization or filtration 991

through a microorganism-retaining filter (if applicable), through to the end of the aseptic 992

filling process. There should be a maximum permissible time for each product that takes 993

into account its composition and the prescribed method of storage. 994

995

v. The holding time for sterilized product prior to filling. 996

997

vi. The aseptic processing time. 998

999

vii. The filling time. 1000

1001

viii. The maximum exposure time of sterilized containers and closures in the critical processing 1002

zone (including filling) prior to closure. 1003

1004

8.19 Aseptic operations (including APS) should be observed at a regular basis by personnel with 1005

specific expertise in aseptic processing to verify the correct performance of operations and address 1006

inappropriate practices if detected. 1007

1008

Finishing of sterile products 1009

1010

8.20 Open primary packaging containers (including partially stoppered vials or prefilled syringes) 1011

should be maintained under Grade A conditions with Grade B background (e.g. Barrier Technology), 1012

or under Grade A conditions with physical segregation from operators (e.g. UDAF carts) until the 1013

stopper is fully inserted. 1014

1015

8.21 Containers should be closed by appropriately validated methods. Containers closed by fusion, 1016

e.g. Blow-fill-seal (BFS), Form-Fill-Seal (FFS), Small and Large Volume Parenteral 1017

(SVP & LVP) bags, glass or plastic ampoules, should be subject to 100% integrity testing. 1018

Samples of containers closed by other methods should be taken and checked for integrity using 1019

validated methods. The frequency of testing should be based on the knowledge and experience of the 1020

container and closure systems being used. A scientifically valid sampling plan should be utilized. 1021

The sample size should be based on information such as supplier approval, packaging component 1022

specifications and process knowledge. It should be noted that visual inspection alone is not 1023

considered as an acceptable integrity test method. 1024

1025

8.22 Containers sealed under vacuum (where the vacuum is necessary for the product stability) 1026

should be tested for maintenance of vacuum after

an appropriate pre-determined period and during 1027

shelf life. 1028

1029

8.23 The container closure integrity validation should take into consideration any transportation or 1030

shipping requirements that may negatively impact the integrity of the container (e.g. by 1031

decompression or temperature extremes). 1032

1033

8.24 Where the equipment used to crimp vial caps can generate large quantities of non-viable 1034

particulates, measures to prevent particulate contamination such as locating the equipment at a 1035

physically separate station equipped with adequate air extraction should be taken. 1036

1037

8.25 Vial capping can be undertaken as an aseptic process using sterilized caps or as a clean

1038

process outside the aseptic core. Where the latter approach is adopted, vials should be

1039

protected by Grade A conditions up to the point of leaving the aseptic processing area, and 1040

thereafter stoppered vials should be protected with a Grade A air supply until the cap has been 1041

crimped. Where capping is a manual process it should be performed under Grade A conditions either 1042

in an appropriately designed isolator or into Grade A zone with a Grade B background. 1043

1044

8.26 Where capping of aseptically filled sterile product is conducted as a clean process with Grade A 1045

air supply protection, vials with missing or displaced stoppers should be rejected prior to capping. 1046

Appropriately qualified, automated methods for stopper height detection should be in place. 1047

1048

8.27 Where

human intervention is required at the capping station, appropriate technological and 1049

organizational measures should be used to prevent direct contact with the vials and to minimize 1050

microbial contamination. 1051

1052

8.28 RABS and isolators may be beneficial in assuring the required

conditions and minimizing the 1053

microbial contamination associated with direct human interventions into the capping operation. 1054

1055

8.29 All filled containers of parenteral products should be inspected individually for extraneous

1056

contamination or other defects. Defect classification and criticality should be determined during 1057

qualification and based on risk and historical knowledge. Factors to consider include, but are not 1058

limited to, the potential impact of the defect to the patient and the route of administration. Different 1059

defect types should be categorized and batch performance analysed. Batches with unusual levels of 1060

defects, when compared with routine defect numbers for the process (based on historical and trend 1061

data), should lead to an investigation. A defect library should be generated and maintained which 1062

captures all known classes of defects. The defect library should be used for the training of production 1063

and quality assurance personnel. Critical defects should not be identified during any subsequent 1064

sampling and inspection of acceptable containers. Any critical defect identified should trigger an 1065

investigation as it indicates a possible failure of the original inspection process. 1066

1067

8.30 When inspection is done manually, it should be performed under

suitable and controlled 1068

conditions of illumination and background. Inspection rates should be appropriately controlled and 1069

qualified. Operators performing the

inspection should undergo visual inspection qualification (whilst 1070

wearing corrective lenses, if these are normally worn) at least annually. The qualification should be 1071

undertaken using appropriate samples from the manufacturer's defect library sets and taking into 1072

consideration worst case scenarios (e.g. inspection time, line speed where the product is transferred to 1073

the operator by a conveyor system, container size or fatigue at the end of shift) and should include 1074

consideration of eyesight checks. Operator distractions should be minimized and frequent breaks, of 1075

an appropriate duration, from inspection should be taken. 1076

1077

8.31 Where automated methods of inspection are used, the process should be validated to detect 1078

known defects (which may impact the product quality, safety or efficacy) and be equal to, or better 1079

than, manual inspection methods. The performance of the equipment should be challenged using 1080

representative defects prior to start up and at regular intervals. 1081

1082

8.32 Results of the inspection should be recorded and defect types and numbers trended. Reject levels 1083

for the various defect types should also be trended based on statistical principles. Impact to product on 1084

the market should be assessed as part of the investigation when adverse trends are observed. 1085

1086

Sterilization 1087

1088

8.33 Where possible, finished product should be terminally sterilized, using a validated and controlled 1089

sterilization process, as this provides a greater assurance of sterility than a validated and controlled 1090

sterile filtration process and/or aseptic processing. Where it is not possible for a product to undergo 1091

terminal sterilization, consideration should be given to using terminal bioburden reduction steps, such 1092

as heat treatments (e.g. pasteurization), combined with aseptic process to give improved sterility 1093

assurance. 1094

1095

8.34 The selection, design and location of the equipment and cycle/programme used for sterilization 1096

should be based on scientific principles and data which demonstrate repeatability and reliability of the 1097

sterilization process. Critical parameters should be defined, controlled, monitored and recorded. 1098

1099

8.35 All sterilization processes should be validated. Validation studies should take into account the 1100

product composition, storage conditions and maximum time between the start of the preparation of a 1101

product or material to be sterilized and its sterilization. Before any sterilization process is adopted, its 1102

suitability for the product and equipment, and its efficacy in consistently achieving the desired 1103

sterilizing conditions in all parts of each type of load to be processed should be validated notably by 1104

physical measurements and where appropriate by biological indicators (BI). For effective sterilization, 1105

the whole of the product, and surfaces of equipment and components should be subject to the required 1106

treatment and the process should be designed to ensure that this is achieved. 1107

1108

8.36 Particular attention should be given when the adopted sterilization method is not described in the 1109

current edition of the Pharmacopoeia, or when it is used for a product which is not a simple aqueous 1110

solution. Where possible, heat sterilization is the method of choice. 1111

1112

8.37 Validated loading patterns should be established for all sterilization processes and should be 1113

subject to periodic revalidation. Maximum and minimum loads should also be considered as part of 1114

the overall load validation strategy. 1115

1116

8.38 The validity of the sterilizing process should be reviewed and verified at scheduled intervals 1117

based on risk. Heat sterilization cycles should be revalidated with a minimum frequency of at least 1118

annually. 1119

1120

8.39 Routine operating parameters should be established and adhered to for all sterilization 1121

processes, e.g. physical parameters and loading patterns. 1122

1123

8.40 There should be mechanisms in place to detect a sterilization cycle that does not conform to the 1124

validated parameters. Any failed sterilization or sterilization that deviated from the validated process 1125

(e.g. have longer or shorter phases such as heating cycles) should be investigated. 1126

1127

8.41 Suitable BIs placed at appropriate locations may be considered as an additional method to 1128

support the validation of the sterilization process. BIs should be stored and used according to the 1129

manufacturer’s instructions. Where BIs are used to support validation and/or to monitor a 1130

sterilization process (e.g. for ethylene oxide), positive controls should be tested for each sterilization 1131

cycle. If BIs are used, strict precautions should be taken to avoid transferring microbial contamination to 1132

the manufacturing or other testing processes. BI results in isolation do not give assurance of 1133

sterilization and should not be used to override other critical parameters and process design 1134

elements. 1135

1136

8.42 The reliability of BIs is important. Suppliers should be qualified and transportation and storage 1137

conditions should be controlled in order that BI quality is not compromised. Prior to use of a new 1138

batch/lot of BIs, the population and identity of the indicator organism of the batch/lot should be 1139

verified. For other critical parameters, e.g. D-value, Z- value, the batch certificate provided by the 1140

qualified supplier can normally be used. 1141

1142

8.43 There should be a clear means of differentiating products, equipment and components, which 1143

have not been subjected to the sterilization process from those which have. Containers used to carry 1144

products such as baskets or trays, items of equipment and/or components should be clearly labelled 1145

(or electronically tracked) with the material name, product batch number and an indication of 1146

whether or not it has been sterilized. Indicators such as autoclave tape, or irradiation indicators may 1147

be used, where appropriate, to

indicate whether or not a batch (or sub-batch) has passed through a 1148

sterilization process. However, these indicators show only that the sterilization process has occurred, 1149

they do not indicate product sterility or achievement of the required sterility assurance level. 1150

1151

8.44 Sterilization records should be available for each sterilization run. Each cycle should have a 1152

unique identifier. They should be reviewed and approved as part of the batch certification procedure. 1153

1154

8.45 Where possible, materials, equipment and components should be sterilized by validated methods 1155

appropriate to the specific material. Suitable protection after sterilization should be provided to 1156

prevent recontamination. If sterilized items are not used immediately after sterilization, these should 1157

be stored using appropriately sealed packaging. A maximum hold time should also be established. 1158

Where justified, components that have been packaged with multiple sterile packaging layers need not 1159

be stored in a cleanroom if the integrity and configuration of the sterile pack allows the items to be 1160

readily disinfected during transfer by operators into the Grade A zone, (e.g. by the use of multiple 1161

sterile coverings that can be removed at each transfer from lower to higher grade). Where protection is 1162

achieved by containment in sealed packaging, this packaging process should be undertaken prior to 1163

sterilization. 1164

1165

8.46 Where materials, equipment, components and ancillary items are sterilized in sealed packaging 1166

and then transferred into the Grade A zone, this should be done using appropriate, validated methods 1167

(for example, airlocks or pass-through hatches) with accompanying disinfection of the exterior of the 1168

sealed packaging. The use of rapid transfer port technology should also be considered. These methods 1169

should be demonstrated to effectively control the potential risk of contamination of the Grade A zone 1170

and Grade B area and, likewise, the disinfection procedure should be demonstrated to be effective in 1171

reducing any contamination on the packaging to acceptable levels for entry of the item into the Grade 1172

B and Grade A areas. 1173

1174

8.47 Where materials, equipment, components and ancillary items are sterilized in sealed packaging 1175

or containers, the packaging sealing process should be validated. The validation should consider the 1176

integrity of the sterile protective barrier system and the maximum hold time before sterilization and 1177

maximum shelf life assigned to the sterilized items. The integrity of the sterile protective barrier 1178

system for each of the sterilized items should be confirmed prior to use. 1179

1180

8.48 For materials, equipment, components and ancillary items that are necessary for aseptic 1181

processing but cannot be sterilized, an effective and validated disinfection and transfer process should 1182

be in place. These items, once disinfected, should be protected to prevent recontamination. These 1183

items, and others representing potential routes of contamination, should be included in the 1184

environmental monitoring program. 1185

1186

1187

Sterilization by heat 1188

1189

8.49 Each heat sterilization cycle should be recorded either electronically or by hardcopy, on 1190

equipment with suitable accuracy and

precision. Monitoring and recording systems should be 1191

independent of the controlling system (e.g. by the use of duplex/double probes). 1192

1193

8.50 The position of the temperature probes used for controlling and/or recording should

be 1194

determined during the validation which should include heat distribution and penetration studies 1195

and, where applicable, also checked against a

second independent temperature probe located at the 1196

same position. 1197

1198

8.51 Sufficient time should be allowed for the whole of the load to reach the required

temperature 1199

before measurement of the sterilizing time-period starts. For sterilization cycles controlled by 1200

using a reference probe within the load, specific consideration should be given to ensuring the 1201

load probe temperature is controlled within defined temperature range prior to cycle 1202

commencement. 1203

1204

8.52 After completion of the high temperature phase of a heat sterilization cycle, precautions should 1205

be taken

against contamination of a sterilized load during cooling. Any cooling liquid or gas that 1206

comes in contact with the product or sterilized material should be sterilized. 1207

1208

8.53 In those cases where parametric release has been authorized, a robust system should be applied 1209

to the product lifecycle validation and the routine monitoring of the manufacturing process. This 1210

system should be periodically reviewed. Further guidance regarding parametric release is provided in 1211

Annex 17. 1212

1213

Moist heat sterilization 1214

1215

8.54 Moist heat sterilization utilises steam or superheated water, typically at lower temperatures and 1216

shorter duration than dry heat processes, in order to sterilize a product or article. Moist heat 1217

sterilization of hard goods or porous loads is primarily effected by latent heat of condensation of 1218

clean steam and the quality of steam is therefore important to provide consistent results. For aqueous 1219

liquid-filled containers, energy from moist heat is transferred through conduction and/or convection 1220

to the content of the container without direct contact with the autoclave steam. In these cases, time 1221

and temperature are the key parameters and steam quality does not have the same impact to the 1222

process. Moist heat sterilization processes may be utilized to sterilize or control bioburden (for non-1223

sterile applications) of thermally stable materials, articles or products and is the preferred method of 1224

sterilization, where possible. Moist heat sterilization can be achieved using steam, (direct or indirect 1225

contact), but also includes other systems such as superheated water systems. Superheated systems 1226

are typically used for the terminal sterilization of product in flexible containers where the pressure 1227

differentials associated with the steam would cause damage to the primary container. 1228

1229

8.55 For porous cycles (hard goods) time, temperature and pressure should be used to monitor the 1230

process. Each item sterilized should be inspected for damage, packaging material integrity and 1231

moisture on removal from the autoclave. Any item found not to be fit for purpose should be 1232

removed from the manufacturing area and an investigation performed. 1233

1234

8.56 For autoclaves fitted with a drain at the bottom of the chamber, the temperature should be 1235

recorded at this position throughout the sterilization period. For steam in place systems, the 1236

temperature should be recorded at condensate drain locations throughout the sterilization period. 1237

1238

8.57 Validation of porous cycles should include a calculation of equilibration time, exposure time, 1239

correlation of pressure and temperature and maximum temperature range during exposure. 1240

Validation of fluid cycles should include temperature, time and/or F

. These critical processing 1241

parameters should be subject to defined limits (including appropriate tolerances) and be confirmed as 1242

part of the sterilization validation and routine cycle acceptance criteria. 1243

1244

8.58 Leak tests on the sterilizing system should be carried out periodically (normally weekly) when a 1245

vacuum phase is part of the

cycle or the system is returned, post-sterilization, to a pressure lower 1246

than the environment surrounding the sterilized system. 1247

1248

8.59 There should be adequate assurance of air removal prior to and during sterilization when the 1249

sterilization process includes air purging (e.g. porous autoclave loads, lyophilizer chambers). For 1250

autoclaves, this should include an air removal test cycle (normally performed on a daily basis) or an 1251

air detector system. Loads to be sterilized should be designed to support effective air removal and be 1252

free draining to prevent the build-up of condensate. 1253

1254

8.60 The items to be sterilized, other than products in sealed containers, should be dry, wrapped in a

1255

material which allows removal of air and penetration of steam and prevents

recontamination after 1256

sterilization. All loaded items should be dry upon removal from the sterilizer. Load dryness should 1257

be confirmed by visual inspection as a part of the sterilization process acceptance. 1258

1259

8.61 If it is necessary to wet equipment using WFI (e.g. ultrafiltration membrane) prior to the 1260

sterilization process, then a risk-based assessment should be carried out to demonstrate the 1261

acceptable dryness level that will not impact the sterility of the equipment sterilized and the product 1262

sterility assurance level. The hold time between the wetting phase and sterilization should be 1263

justified and validated. 1264

1265

8.62 Distortion and damage of non-rigid containers that are terminally sterilized, such as containers 1266

produced by Blow-Fill-Seal or Form-Fill-Seal technologies, should be prevented by appropriate cycle 1267

design and control (for instance setting correct pressure, heating and cooling rates and loading 1268

patterns). 1269

1270

8.63 Where steam in place systems are used (e.g. for fixed pipework, vessels and lyophilizer 1271

chambers), the system should be appropriately designed and validated to assure all parts of the 1272

system are subjected to the required treatment. The system should be monitored for temperature, 1273

pressure and time at appropriate locations during routine use to ensure all areas are effectively and 1274

reproducibly sterilized. These locations should be demonstrated as being representative of, and 1275

correlated with, the slowest to heat locations during initial and routine validation. Once a system has 1276

been sterilized by steam in place it should remain integral and held under positive pressure prior to 1277

use. 1278

1279

8.64 For systems using superheated water rather than steam, as the sterilizing agent, the heated water 1280

should consistently reach all of the required contact points. Initial qualification studies should 1281

include temperature mapping of the entire load. There should be routine checks on the equipment to 1282

ensure that nozzles (where the water is introduced) are not blocked and drains remain free from 1283

debris. 1284

1285

8.65 For the qualification of superheated systems it should be demonstrated that all parts of the load 1286

meet the minimum required temperature and that routine monitoring probes are located in the worst 1287

case positions identified during the qualification process. 1288

1289

Dry heat sterilization 1290

1291

8.66 Dry heat sterilization is of particular use in the removal of thermally robust contaminants such as 1292

pyrogens and is often used in the preparation of components for aseptic filling. The combination of 1293

time and temperature to which product, components and equipment are exposed should produce an 1294

adequate and reproducible level of lethality and/or pyrogen (endotoxin) inactivation/removal when 1295

operated routinely within the established limits. 1296

1297

8.67 Dry heat sterilization/depyrogenation tunnels should be configured to ensure that airflow protects 1298

the integrity and performance of the Grade A sterilizing zone by maintaining pressure differentials 1299

and airflow through the tunnel from the higher grade area to the lower grade area. Airflow patterns 1300

should be visualised and correlated with temperature studies. The impact of any airflow change 1301

should be assessed to ensure the heating profile is maintained. All air supplied to the tunnel should 1302

pass through at least a HEPA filter and periodic tests should be performed to demonstrate air filter 1303

integrity (at least biannually). Any tunnel parts that come into contact with sterilized components 1304

should be appropriately sterilized or disinfected. Critical process parameters that should be considered 1305

during validation and/or routine processing should include, but may not be limited to: 1306

1307

i. Belt speed or dwell time within the sterilizing zone. 1308

1309

ii. Temperature – minimum and maximum temperatures. 1310

1311

iii. Heat penetration of the material/article. 1312

1313

iv. Heat distribution/uniformity. 1314

1315

v. Airflows – correlated with the heat distribution and penetration studies. 1316

1317

8.68 When a thermal depyrogenation process is used for any component or product contact 1318

equipment, validation studies should be performed to demonstrate that the process provides a suitable 1319

value and results in a minimum 3 log reduction in endotoxins concentration. 1320

1321

8.69 Containers inoculated with endotoxin should be used during validation and should be carefully 1322

managed with a full reconciliation performed. Containers should be representative of the materials 1323

normally processed. Endotoxin quantification and recovery efficiency should also be demonstrated 1324

through biological measurement. 1325

1326

8.70 Dry heat ovens are typically employed to sterilize or depyrogenate primary packaging 1327

components, finished materials or active substances but may be used for other processes. They should 1328

be maintained at a positive pressure relative to lower grade areas throughout the sterilization and post 1329

sterilization hold process. All air entering the oven should pass through a sterilizing filter. Critical 1330

process parameters that should be considered in qualification and/or routine processing should 1331

include, but may not be limited to: 1332

1333

i. Temperature. 1334

1335

ii. Exposure period/time. 1336

1337

iii. Chamber pressure (for maintenance of over pressure). 1338

1339

iv. Air speed. 1340

1341

v. Air quality within the oven. 1342

1343

vi. Heat penetration of material/article (slow to heat spots). 1344

1345

vii. Heat distribution/uniformity. 1346

1347

8.71 For dry heat sterilization of starting materials and intermediates, the same principles should be 1348

applied. Consideration should also be given to factors affecting heat penetration such as the container 1349

type, size and packing matrix. 1350

1351

Sterilization by radiation 1352

1353

8.72 Guidance regarding ionising radiation sterilization can be found within Annex 12. 1354

1355

8.73 Sterilization by radiation is used mainly for the sterilization of heat sensitive materials and

1356

products. Ultraviolet irradiation is not an acceptable method of sterilization. 1357

1358

8.74 Validation procedures should ensure that the effects of variations in density of the product and 1359

packages are considered. 1360

1361

Sterilization with ethylene oxide 1362

1363

8.75 This method should only be used when no other method is practicable. During process 1364

validation, it should be shown that there is no damaging effect on the product and that the

1365

conditions and time allowed for degassing result in the reduction of any residual ethylene oxide 1366

(EO) gas and reaction

products to defined acceptable limits for the given product or material. 1367

1368

8.76 Direct contact between gas and microbial cells is essential, precautions should be taken to avoid 1369

the presence of organisms likely to be enclosed in material such as crystals or dried protein. The 1370

nature, porosity and quantity of packaging materials can significantly affect the process. 1371

1372

8.77 Before exposure to the gas, materials should be brought into equilibrium with the

1373

humidity and temperature required by the process. The time required for this should be

1374

balanced against the opposing need to minimize the time before sterilization. 1375

1376

8.78 Each sterilization cycle should be monitored with suitable BIs, using the

appropriate number of 1377

test units distributed throughout the load at defined locations that have been shown to be worst case 1378

during validation. 1379

1380

8.79 Critical process variables that could be considered as part of the sterilization process validation 1381

and routine monitoring include, but are not limited to: 1382

1383

i. EO gas concentration. 1384

1385

ii. EO gas pressure. 1386

1387

iii. Amount of EO gas used. 1388

1389

iv. Relative humidity. 1390

1391

v. Temperature. 1392

1393

vi. Exposure time. 1394

1395

8.80 After sterilization, the load should be aerated to allow EO gas and/or its reaction products to 1396

desorb from the packaged product to predetermined levels. Aeration can occur within a sterilizer 1397

chamber and/or in a separate aeration chamber or aeration room. The aeration phase should be 1398

validated as part of the overall EO sterilization process validation. 1399

1400

Filter sterilization of products which cannot be sterilized in their final container 1401

1402

8.81 If the product cannot be sterilized in the final container, solutions or liquids should be sterilized 1403

by filtration through a sterile sterilizing grade filter (with a nominal pore size of 0.22 µm (or less) 1404

that has been appropriately validated to obtain a sterile filtrate) and subsequently aseptically filled 1405

into a previously sterilized container. The selection of the filter used should ensure that it is 1406

compatible with the product and as described in the marketing authorization (refer to paragraph 1407

8.125). 1408

1409

8.82 Suitable bioburden reduction prefilters and/or sterilizing grade filters may be used at multiple 1410

points during the manufacturing process to ensure a low and controlled bioburden of the liquid prior 1411

to the primary sterilizing grade filter. Due to the potential additional risks of a sterile filtration 1412

process, as compared with other sterilization processes, a second filtration through a sterile sterilizing 1413

grade filter, immediately prior to filling, should be considered as part of an overall CCS. 1414

1415

8.83 The selection of components for the filtration system and their interconnection and arrangement 1416

within the filtration system, including pre-filters, should be based on the critical quality attributes of 1417

the product, justified and documented. The filtration system should minimize the generation of fibres 1418

and particulates, not cause or contribute to unacceptable levels of impurities, or possess characteristics 1419

that otherwise alter the quality and efficacy of the product. Similarly, the filter characteristics should 1420

be compatible with the fluid and not be adversely affected by the product to be filtered. Adsorption of 1421

product components and extraction/leaching of filter components should be evaluated (refer to 1422

paragraph 8.125). 1423

1424

8.84 The filtration system should be designed to: 1425

1426

i. Allow operation within validated process parameters. 1427

1428

ii. Maintain the sterility of the filtrate. 1429

1430

iii. Minimize the number of aseptic connections required between the sterilizing filter and the 1431

final filling of the product. 1432

1433

iv. Allow cleaning procedures to be conducted as necessary. 1434

1435

v. Allow sterilization procedures, including sterilization in place, to be conducted as necessary. 1436

1437

vi. Permit in-place integrity testing, of the 0.22 µm sterilizing filter, preferably as a closed 1438

system, prior to filtration as necessary. In-place integrity testing methods should be selected 1439

to avoid any adverse impact on the quality of the product. 1440

1441

8.85 Sterile filtration of liquids should be validated in accordance with European (or other relevant) 1442

Pharmacopeia requirements. Validation can be grouped by different strengths or variations of a 1443

product but should be done under worst case conditions. The rationale for grouping should be 1444

justified and documented. 1445

1446

8.86 During filter validation, wherever possible, the product to be filtered should be used for 1447

bacterial retention testing of the sterilizing filter. Where the product to be filtered is not suitable for 1448

use in bacterial retention testing, a suitable surrogate product should be justified for use in the test. 1449

The challenge organism used in the bacterial retention test should be justified. 1450

1451

8.87 Filtration parameters that should be considered and established in validation and monitored in 1452

routine processing should include, but are not limited to: 1453

1454

1455

i. The wetting fluid used for filter integrity testing should be based on the filter manufacturer’s 1456

recommendation or the fluid to be filtered. The appropriate integrity test value specification 1457

should be established. 1458

ii. 1459

If the system is flushed or integrity tested in-situ with a fluid other than the product, 1460

appropriate actions are taken to avoid any deleterious effect on product quality. 1461

1462

iii. Filtration process conditions including: 1463

1464

• Fluid pre-filtration holding time and effect on bioburden. 1465

1466

• Filter conditioning, with fluid if necessary. 1467

1468

• Maximum filtration time/total time filter is in contact with fluid. 1469

1470

• Maximum operating pressure. 1471

1472

• Flow rate. 1473

1474

• Maximum filtration volume. 1475

1476

• Temperature. 1477

1478

• The time taken to filter a known volume of bulk solution and the

pressure difference 1479

to be used across the filter. 1480

1481

Note: Results of these checks should be included in the batch record. Any

significant difference in 1482

parameters from those validated to those observed during routine manufacturing should be noted 1483

and

investigated. 1484

1485

8.88 The integrity of the sterilized filter assembly should be verified by integrity testing before use, to 1486

check for damage and loss of integrity caused by the filter preparation prior to use. A sterilizing grade 1487

filter that is used to sterilize a fluid should be subject to a non-destructive integrity test post-use prior 1488

to removal of the filter from its housing. Test results should correlate to the microbial retention 1489

capability of the filter established during validation. Examples of tests that are used include bubble 1490

point, diffusive flow, water intrusion or pressure hold test. It is recognized that pre-use post 1491

sterilization integrity testing (PUPSIT) may not always be possible after sterilization due to process 1492

constraints (e.g. the filtration of very small volumes of solution). In these cases, an alternative 1493

approach may be taken providing that a thorough risk assessment has been performed and compliance 1494

is achieved by the implementation of appropriate controls to mitigate any risk of non-sterility. Points 1495

to consider in such a risk assessment should include but are not be limited to: 1496

1497

i. In depth knowledge and control of the sterilization process to ensure that the potential for 1498

damage to the filter is minimized. 1499

1500

ii. In depth knowledge and control of the supply chain to include: 1501

• Contract sterilization facilities. 1502

• Defined transport mechanisms. 1503

• Packaging of the sterilized filter, to prevent damage to the filter during transportation 1504

and storage. 1505

iii. In depth process knowledge such as: 1506

• The specific product type, including particulate burden and whether there exists any 1507

risk of impact on filter integrity values, such as the potential to alter integrity testing 1508

values and therefore prevent the detection of a non-integral filter during a post-use 1509

filter integrity test. 1510

• Pre-filtration and processing steps, prior to the sterilizing filter, which would remove 1511

particulate burden and clarify the product prior to the sterile filtration. 1512

1513

8.89 The integrity of critical sterile gas and air vent filters (that are directly linked to the sterility of 1514

the product) should be verified by testing after use, with the filter remaining in the filter assembly. 1515

1516

8.90 The integrity of non-critical air or gas vent filters should be confirmed and recorded at 1517

appropriate intervals. Where gas filters are in place for extended periods such as vent filters, integrity 1518

testing should be carried out pre and post-use. The maximum duration of use should be specified and 1519

monitored based on risk (e.g. considering the maximum number of uses and sterilization cycles 1520

permitted). 1521

1522

8.91 For gas filtration, attention should be paid to avoiding unintended moistening or wetting of the 1523

filter or filter equipment. This can be achieved by the use of hydrophobic filters. 1524

1525

8.92 If the sterilizing filtration process has been validated as a system consisting of multiple filters to 1526

achieve the sterility for a given fluid, the filtration system is considered to be a single sterilizing unit 1527

and all filters within the system should satisfactorily pass integrity testing after use. 1528

1529

8.93 In a redundant filtration system (where a second filter is present as a backup but the sterilizing 1530

process is validated as only requiring one filter), post-use integrity test of the primary sterilizing filter 1531

should be performed and if demonstrated to be integral, then a post-use integrity test of the secondary 1532

filter is not necessary. However, in the event of a failure of the post-use integrity test on the primary 1533

filter, a risk assessment should be carried out to determine the acceptability of performing a post-use 1534

integrity test on the secondary (redundant) filter. 1535

1536

8.94 Bioburden samples should be taken from the bulk product and immediately prior to the final 1537

sterile filtration. Systems for taking samples should be designed so as not to introduce contamination. 1538

1539

8.95 Liquid sterilizing filters should be discarded after the processing of a single lot and the same 1540

filter should not be used for more than one working day unless such use has been validated. 1541

8.96 Where campaign manufacture of a product has been appropriately justified in the CCS and 1542

validated, the filter user should: 1543

i. Assess and document the risks associated with the duration of filter use for the sterile 1544

filtration process for a given fluid. 1545

ii. Conduct and document effective validation and qualification studies to demonstrate that the 1546

duration of filter use for a given sterile filtration process and for a given fluid does not 1547

compromise performance of the sterilizing filter or filtrate quality. 1548

iii. Document the maximum validated duration of use for the filter and implement controls to 1549

ensure that filters are not used beyond the validated maximum duration. Records of these 1550

controls should be maintained. 1551

iv. Implement controls to ensure that filters contaminated with fluid or cleaning agent residues, 1552

or considered defective in any other way, are removed from use. 1553

Form-Fill-Seal 1554

1555

8.97 Form-Fill-Seal (FFS) units include blow moulding from thermoplastic granulate and 1556

thermoforming from thermoplastic film, typically known as Blow-Fill-Seal (BFS) and Vertical-Form-1557

Fill-Seal (VFFS), respectively. VFFS process is an automated filling process, typically for terminally 1558

sterilized products, that may utilize a single or dual web system which constructs the primary 1559

container out of a flat roll of thermoplastic film while simultaneously filling the formed bags with 1560

product and sealing the filled bags in a continuous process. All such containers are considered to be 1561

closed through sealing by fusion and, as such, fall under the requirement to perform 100% integrity 1562

testing (refer to paragraph 8.21). 1563

1564

8.98 Process parameters relating to seal integrity should be qualified and appropriately controlled. 1565

1566

8.99 Critical parameters include, but are not limited to: 1567

1568

i. Seal strength. 1569

1570

ii. Seal uniformity. 1571

1572

iii. Sealing temperatures. 1573

1574

iv. Sealing pressures. 1575

1576

v. Sealing times. 1577

1578

vi. Dwell time for filling. 1579

1580

8.100 Seal strength and uniformity should be monitored routinely. 1581

1582

8.101 The controls identified during qualification should be in alignment with the site’s CCS. Aspects 1583

to be considered include but are not limited to: 1584

1585

i. Determination of the boundaries of the critical zone. 1586

1587

ii. Environmental control and monitoring, both of the machine and the background in which it 1588

is placed. 1589

1590

iii. Integrity testing of the product filling lines. 1591

1592

iv. Integrity testing of the cooling system. 1593

1594

v. Duration of the batch or filling campaign. 1595

1596

vi. Control of polymer starting material (including resin pellets). 1597

1598

vii. Cleaning-in-place and sterilization-in-place of equipment in direct contact to the formulation 1599

(product filling lines); sterilization-in-place of sterile air pathways. 1600

1601

Blow-Fill-Seal 1602

1603

8.102 Blow-Fill-Seal (BFS) units are purpose built machines in which, in one continuous operation, 1604

containers are formed from a thermoplastic granulate, filled and then sealed by one automatic 1605

machine. Air that makes contact with critical surfaces of the container during extrusion, formation or 1606

sealing of the moulded container should undergo appropriate filtration. 1607

1608

8.103 For shuttle type equipment used for aseptic filling, the area between parison cutting and mould 1609

sealing should be covered by a flow of filtered air to provide Grade A conditions at the critical zone. 1610

The equipment should be installed in at least a Grade C environment, provided that Grade A/B 1611

clothing is used. The filling environment should meet Grade A for viable and non-viable limits at rest 1612

and the viable limit only when in operation. 1613

1614

8.104 For rotary-type equipment, used for aseptic filling, the filling environment should be designed 1615

to meet Grade A conditions. Other sterility assurance controls such as monitoring of critical 1616

parameters and alarms during each batch and parison support filter integrity testing should be 1617

considered. 1618

1619

8.105 The environmental control and monitoring program should take into consideration the moving 1620

parts and complex airflow paths generated by the BFS process and the effect of the high heat outputs 1621

of the process, e.g. by performing smoke studies and/or other equivalent studies. Environmental 1622

monitoring should be applied taking into consideration elements such as air-filter configuration, air-1623

filter integrity, cooling systems integrity, equipment design and installation. 1624

1625

8.106 Blow-Fill-Seal equipment used for the manufacture of products which are terminally sterilized 1626

should be installed in at least a Grade D environment. The conditions at the point of fill should 1627

comply with the environmental requirements of paragraphs 8.3 and 8.4. 1628

1629

8.107 External particulate and microbial contamination of the polymer should be prevented by 1630

appropriate design, control, and maintenance of the polymer storage, sampling and distribution 1631

systems. The capability of the extrusion system to provide appropriate sterility assurance for the 1632

moulded container should be fully understood and validated. The sampling frequency, the bioburden 1633

and, where applicable, endotoxins levels of the raw polymer should be defined and controlled within 1634

the CCS. 1635

1636

8.108 Interventions requiring cessation of filling and/or extrusion, moulding and sealing and, where 1637

required, re-sterilization of the filling machine should be clearly defined and well described in the 1638

aseptic filling procedure, and included in the APS (refer to paragraphs 9.36, 9.37 and 9.38). 1639

1640

8.109 The moulds used to form containers are considered critical equipment and any changes or 1641

modification to moulds should result in an assessment of finished product container integrity, and 1642

should be supported by validation. 1643

1644

Lyophilization 1645

1646

8.110 Lyophilization is a critical process step and all activities that can affect the sterility of the 1647

product or material need to be regarded as extensions of the aseptic processing of the sterilized 1648

product. The lyophilization equipment and its processes should be designed to ensure that product or 1649

material sterility is maintained during lyophilization by preventing microbial and particulate 1650

contamination between the filling of products for lyophilization, and completion of lyophilization 1651

process. All control measures in place should be determined by the site’s CCS. 1652

1653

8.111 The sterilization of lyophilizers and associated equipment, (e.g. trays, vial support rings) should 1654

be validated and holding times between sterilization cycles appropriately challenged during aseptic 1655

process simulations. The lyophilizer should be sterilized regularly, based on system design. Re-1656

sterilization should be performed following maintenance or cleaning. Sterilized lyophilizers and 1657

associated equipment should be protected from contamination after sterilization. 1658

1659

8.112 Lyophilizers that are manually loaded or unloaded should normally be sterilized before each 1660

load. For lyophilizers loaded by automated closed systems or located within systems that exclude 1661

operator intervention, the frequency of sterilization should be justified and documented as part of the 1662

CCS. 1663

1664

8.113 The integrity of the lyophilizer system should be maintained following sterilization and during 1665

use. The filter used to maintain lyophilizer integrity should be sterilized before each use of the system 1666

and its integrity testing results should be part of the batch certification. The frequency of vacuum/leak 1667

integrity testing of the chamber should be documented and the maximum permitted leakage of air into 1668

the lyophilizer should be specified and checked at the start of every cycle. 1669

1670

8.114 Lyophilization trays should be checked regularly to ensure that they are not misshapen or 1671

damaged. 1672

1673

8.115 Points to consider for the design of loading (and unloading, where the lyophilised material is 1674

not in a sealed container (e.g. open tray dried materials), include but are not limited to: 1675

1676

i. The loading pattern within the lyophilizer should be specified and documented. 1677

1678

ii. The transfer of partially closed containers to a lyophilizer should be undertaken under Grade 1679

A conditions at all times and handled in a manner designed to minimize direct operator 1680

intervention. Technologies such as conveyor systems, portable transfer systems (e.g. clean air 1681

transfer carts, portable unidirectional airflow workstations) should be used to ensure that the 1682

cleanliness of the system used to transfer the partially closed containers is maintained). 1683

Alternatively, where supported by validation, containers closed in the Grade A zone and not 1684

reopened whilst in the Grade B may be used to protect partially stoppered vials (e.g. sealed 1685

sterilized trays). 1686

1687

iii. Airflow patterns should not be adversely affected by transport devices and venting of the 1688

loading zone. 1689

1690

iv. Unsealed containers (such as partially stoppered vials) should be maintained under Grade A 1691

conditions and should normally be separated from operators by physical barrier technology or 1692

any other appropriate measures. 1693

1694

v. Where seating of the stoppers is not completed prior to opening the lyophilizer chamber, 1695

product removed from the lyophilizer should remain under Grade A conditions during 1696

subsequent handling. 1697

1698

vi. Utensils used during transfer to and loading and unloading of the lyophilizer (such as trays, 1699

bags, placing devices, tweezers, etc.) should be subject to a validated sterilization process. 1700

1701

Closed systems 1702

1703

8.116 Closed systems can be single use systems (i.e. disposable systems) and fixed systems (such as 1704

vessels with fixed pipework). Guidance in this section is equally applicable to both systems. 1705

1706

8.117 The use of closed systems can reduce the risk of extraneous contamination such as microbial, 1707

particulate and chemical from the adjacent environment. Closed systems should always be designed to 1708

reduce the need for, and complexity of manual interventions. 1709

1710

8.118 It is critical to ensure the sterility of all product contact surfaces of closed systems used for 1711

aseptic processing. The design and selection of any closed system used for aseptic processing should 1712

ensure maintenance of sterility. Connection of sterile equipment (e.g. tubing / pipework) to the 1713

sterilized product pathway after the final sterilizing filter should be designed to be connected 1714

aseptically (e.g. by intrinsic aseptic connectors or fusion systems). 1715

1716

8.119 Appropriate measures should be in place to ensure the integrity of components used in aseptic 1717

connections. The means by which this is achieved should be determined and captured in the CCS. 1718

Appropriate system integrity tests should be considered when there is a risk of compromising product 1719

sterility. Supplier assessment should include the collation of data in relation to potential failure modes 1720

that may lead to a loss of system sterility. 1721

1722

8.120 The background in which closed systems are located should be based on their design and the 1723

processes undertaken. For aseptic processing and where there are any risks that system integrity may 1724

be compromised, the system should be located in a Grade A zone. If the system can be shown to 1725

remain integral at every usage (e.g. via pressure testing and/or monitoring) then a lower classified area 1726

may be used. If the closed system is opened (e.g. for maintenance of a bulk manufacturing line) then 1727

this should be performed in a classified area appropriate to the materials (e.g. Grade C for terminally 1728

sterilization processes, or Grade A for aseptic processing) or be subject to further cleaning and 1729

disinfection (and sterilization in case of aseptic processes). 1730

1731

Single use systems (SUS) 1732

1733

8.121 SUS are those technologies used in manufacture of sterile products which are used as an 1734

alternative to reusable equipment. SUS can be individual components or made up of multiple 1735

components such as bags, filters, tubing, connectors, valves, storage bottles and sensors. 1736

1737

8.122 There are some specific risks associated with SUS which should be assessed as part of the CCS. 1738

These risks include but are not limited to: 1739

1740

i. The interaction between the product and product contact surface (such as adsorption, or the 1741

formation of leachables and extractables). 1742

1743

ii. The fragile nature of the system compared to fixed reusable systems. 1744

1745

iii. The increase in the number and complexity of manual operations (including inspection and 1746

handling of the system) and connections made. 1747

1748

iv. The complexity of the assembly. 1749

1750

v. The performance of the pre-use integrity test for sterilizing grade filters (refer to paragraph 1751

8.88). 1752

1753

vi. The risk of holes and leakage. 1754

1755

vii. The potential for compromising the system at the point of opening the outer packaging. 1756

1757

viii. The risk of particulate contamination. 1758

1759

8.123 Sterilization processes for SUS should be validated and shown to have no adverse impact on 1760

system performance. 1761

1762

8.124 Assessment of suppliers of disposable systems including sterilization is critical to the selection 1763

and use of these systems. For sterile SUS, verification of sterility should be performed as part of the 1764

supplier qualification and on receipt and use of each unit. 1765

1766

8.125 The adsorption and reactivity of the product with product contact surfaces should be evaluated 1767

under process conditions. 1768

1769

8.126 The extractable and leachable profile of the SUS and any impact on the quality of the product 1770

especially where the system is made from polymer-based materials should be evaluated. An 1771

assessment should be carried out for each component to evaluate the applicability of the extractable 1772

profile data. For components considered to be at high risk from leachables, including those that may 1773

absorb processed materials or those with extended material contact times, an assessment of leachable 1774

profile studies, including safety concerns, should be taken into consideration. If applying simulated 1775

processing conditions, these should accurately reflect the actual processing conditions and be based 1776

on a scientific rationale. 1777

1778

8.127 SUS should be designed to maintain integrity throughout processing under the intended 1779

operational conditions. Attention to the structural integrity of the single use components is necessary 1780

where these may be exposed to more extreme conditions (e.g. freezing and thawing processes) either 1781

during routine processing or transportation. This should include verification that intrinsic aseptic 1782

connections (both heat sealed and mechanically sealed) remain integral under these conditions. 1783

1784

8.128 Acceptance criteria should be established and implemented for SUS corresponding to the risks 1785

or criticality of the products and its processes. On receipt, each piece of SUS should be checked to 1786

ensure that they have been manufactured, supplied and delivered in accordance with the approved 1787

specification. A visual inspection of the outer packaging (e.g. appearance of exterior carton, product 1788

pouches), label printing, and review of attached documents (e.g. certificate of conformance and proof 1789

of sterilization) should be carried out and documented prior to use. 1790

1791

8.129 Critical manual handling operations of SUS such as assembly and connections should be 1792

subject to appropriate controls and verified during the APS. 1793

1794

9 Viable and non-viable environmental & process monitoring 1795

1796

General 1797

1798

9.1 The site’s environmental and process monitoring program forms part of the overall CCS and is 1799

used to monitor the controls designed to minimize the risk of microbial and particulate contamination. 1800

It should be noted that the reliability of each of the elements of the monitoring system (viable, non-1801

viable and APS) when taken in isolation is limited and should not be considered individually to be an 1802

indicator of asepsis. When considered together, their reliability is dependent on the design, validation 1803

and operation of the system that they are monitoring. 1804

1805

9.2 This program is typically comprised of the following elements: 1806

i. Environnemental monitoring – non-viable particles. 1807

1808

ii. Environmental and personnel monitoring – viable particles. 1809

1810

iii. Aseptic process simulation (aseptically manufactured product only). 1811

1812

9.3 The information from these systems should be used for routine batch certification and for periodic 1813

assessment during process review or investigation. This applies for both terminal sterilization and 1814

aseptic processes, however, the criticality of the impact may differ depending upon the product and 1815

process type. 1816

1817

Environmental monitoring 1818

1819

9.4 Risk assessments should be performed in order to establish a comprehensive environmental 1820

monitoring program, i.e. sampling locations, frequency of monitoring, monitoring method used and 1821

incubation conditions (e.g. time, temperature(s), aerobic and/or anaerobic conditions). These risk 1822

assessments should be conducted based on detailed knowledge of; the process inputs and final 1823

product, the facility, equipment, specific processes, the operations involved, historical monitoring 1824

data, monitoring data obtained during qualification and knowledge of typical microbial flora isolated 1825

from the environment. Consideration of other information such as air visualization studies should 1826

also be included. These risk assessments should be reviewed regularly in order to confirm the 1827

effectiveness of the site’s environmental monitoring program. The monitoring program should be 1828

considered in the overall context of the trend analysis and the CCS for the site. 1829

1830

9.5 Routine monitoring of cleanrooms, clean air equipment and personnel should be performed in 1831

operation throughout all critical stages, including equipment set-up. 1832

1833

9.6 The monitoring of Grade A zones should demonstrate the maintenance of aseptic processing 1834

conditions during critical operations. Monitoring should be performed at locations posing the highest 1835

risk of contamination to the sterile equipment surfaces, container, closures and product. The selection 1836

of monitoring locations and the orientation and positioning of sampling devices should be justified 1837

and appropriate to obtain reliable data from the critical zones. 1838

1839

9.7 Sampling methods should not pose a risk of contamination to the manufacturing operations. 1840

1841

9.8 Appropriate alert levels and action limits should be set for the results of viable and non-viable 1842

particle monitoring. Alert levels should be established based on results of cleanroom qualification 1843

tests or trend data and should be subject to periodic review. 1844

1845

9.9 Alert levels for Grade A (non-viable particles only) Grade B, Grade C and Grade D should be set 1846

such that adverse trends (e.g. a numbers of events or individual events that indicate a deterioration of 1847

cleanliness) are detected and addressed. 1848

1849

9.10 Monitoring procedures should define the approach to trending. Trends can include, but are not 1850

limited to: 1851

1852

i. Increasing numbers of action limit or alert level breaches. 1853

1854

ii. Consecutive breaches of alert levels. 1855

1856

iii. Regular but isolated breaches of action limits that may have a common cause, for example 1857

single excursions that always follow planned preventative maintenance. 1858

1859

iv. Changes in microbial flora type and numbers and predominance of specific organisms. 1860

Particular attention should be given to objectionable organisms or those that can be difficult 1861

to control such as spore-forming microorganisms. 1862

1863

9.11 The monitoring of Grade C and D cleanrooms in operation should be performed based on data 1864

collected during qualification and historical data to allow effective trend analysis. The requirements 1865

of alert levels and action limits will depend on the nature of the operations carried out. Action limits 1866

may be more stringent than those listed in Table 6 and Table 7. 1867

1868

9.12 If action limits are exceeded, operating procedures should prescribe a root cause investigation, 1869

an assessment of the potential impact to product and requirements for corrective and preventive 1870

actions. If alert levels are exceeded, operating procedures should prescribe assessment and follow-1871

up, which should include consideration of an investigation and/or corrective actions to avoid any 1872

further deterioration of the environment. 1873

1874

9.13 Results from environmental monitoring should be considered when reviewing batch 1875

documentation for finished product batch certification. 1876

1877

Environmental monitoring- non-viable particles 1878

1879

9.14 Non-viable particulate monitoring systems should be established to obtain data for assessing 1880

potential contamination risks and to ensure the maintenance of the environment for sterile operations 1881

in a qualified state. 1882

1883

9.15 The limits for environmental monitoring of airborne particulate concentrations for each graded 1884

area are given in Table 6. 1885

1886

Table 6: Limits for airborne particulate concentration for the monitoring of non-viable 1887

contamination. 1888

1889

Grade

aximum limits for

rticulate

≥ 0.5 μm/m

aximum limits for

particulate

≥ 5 μm/m

at rest

in operation

at rest

in operation

3 520

352 000

2 900

352 000

3 520 000

2 900

29 000

3 520

000

Not defined

(

)

29 000

Not defined

(

)

1890

1891

1892

(a)

For Grade D, in operation limits are not defined. The company should establish in operation 1893

limits based on a risk assessment and on historical data, where applicable. 1894

1895

Note 1: The particulate limits given in the table for the “at rest” state should be achieved after 1896

a short “clean up” period (defined during qualification with a guidance value of 15 to 20 1897

minutes) in an unmanned state, after the completion of operations (refer to paragraph 4.30 and 1898

4.31). 1899

1900

Note 2: With regards to the monitoring of airborne particulates ≥5 μm particulate 1901

concentration, the limit of 29 (Grade A) is selected due to the limitations of monitoring 1902

equipment. Alert levels should be set based on historical data, such that frequent sustained 1903

counts below the action limit which may be indicative of system contamination or 1904

deterioration should trigger an investigation. For the Grade A zone and Grade B area the 1905

importance of monitoring the ≥5 μm particulates is to identify negative trends as defined in the 1906

manufacturer's CCS. 1907

1908

9.16 For the Grade A zone, particulate monitoring should be undertaken for the full duration of 1909

critical

processing, including equipment assembly. 1910

1911

9.17 The Grade A zone should be monitored continuously (for particulates ≥0.5 and ≥5 µm) and 1912

with a suitable sample flow rate (at least 28 litres (1ft

) per minute) so that all interventions, transient 1913

events and any system deterioration is captured. The system should frequently correlate each 1914

individual sample result with the limits in Table 6 at such a frequency that any potential excursion 1915

can be identified and responded to in a timely manner. Alarms should be triggered if alert levels are 1916

exceeded. Procedures should define the actions to be taken in response to alarms including the 1917

consideration of additional microbial monitoring. 1918

1919

9.18 It is recommended that a similar system be used for Grade B area although the sample frequency 1920

may be decreased. The Grade B zone should be monitored at such a frequency and with suitable 1921

sample size that the programme captures any increase in levels of contamination and system 1922

deterioration. If alert or action levels are exceeded, alarms should be triggered. 1923

1924

9.19 The selection of the monitoring system should take i n t o account any risk presented by 1925

the

materials used in the manufacturing operation (for example, those involving live organisms, 1926

powdery products or radiopharmaceuticals) that may give rise to biological or chemical hazards. 1927

1928

9.20 In the case where contaminants are present due to the processes involved and would potentially 1929

damage the particle counter or present a hazard (e.g. live organisms, powdery products and radiation 1930

hazards), the frequency and strategy employed should be such as to assure the environmental 1931

classification both prior to and post exposure to the risk. An increase in viable particle monitoring 1932

should be considered to ensure comprehensive monitoring of the process. Additionally, monitoring 1933

should be performed during simulated operations. Such operations should be performed at appropriate 1934

intervals. The approach should be defined in the CCS. 1935

1936

9.21 The size of monitoring samples taken using automated systems will usually be a

function of the 1937

sampling rate of the system used. It is not necessary for the sample volume to

be the same as that 1938

used for formal classification of cleanrooms and clean air equipment. Monitoring sample volumes 1939

should be justified. 1940

1941

9.22 The occasional indication of macro particulate counts, especially ≥ 5 µm, may be considered to 1942

be false counts due to electronic noise, stray light, coincidence, etc. However, consecutive or regular 1943

counting of low levels may be indicative of a possible contamination event and should be 1944

investigated. Such events may indicate early failure of the room air supply filtration system, filling 1945

equipment failure, or may also be diagnostic of poor practices during machine set-up and routine 1946

operation. 1947

1948

9.23 Monitoring conditions such as frequency, sampling volume or duration, alert levels and action 1949

limits and corrective actions (including an investigation) should be established in each 1950

manufacturing area based on data generated during the initial qualification process, ongoing routine 1951

monitoring and periodic review of data. 1952

1953

Environmental and personnel monitoring-viable particles 1954

1955

9.24 Where aseptic operations are performed, microbial monitoring should be frequent using a 1956

combination of methods such as settle plates, volumetric air sampling, glove, gown and surface 1957

sampling (e.g. swabs and contact plates). The method of sampling used should be justified within 1958

the CCS and should be demonstrated not to have a detrimental impact on Grade A and B airflow 1959

patterns. 1960

1961

9.25 Monitoring should include sampling of personnel at periodic intervals during the process. 1962

Sampling of personnel should be performed in such a way that it will not compromise the process. 1963

Particular consideration should be given to monitoring personnel following involvement in critical 1964

interventions and on each exit from the Grade B cleanroom. 1965

1966

9.26 Viable particle monitoring should also be performed within the cleanrooms when normal 1967

manufacturing operations are not occurring (e.g. post disinfection, prior to start of manufacturing, 1968

on completion of the batch and after a shutdown period), and in associated rooms that have not 1969

been used, in order to detect potential incidents of contamination which may affect the controls 1970

within the cleanrooms. In case of an incident, additional sample locations may be used as a 1971

verification of the effectiveness of a corrective action (i.e. cleaning and disinfection). 1972

1973

9.27 Continuous viable air monitoring in the Grade A zone (e.g. air sampling or settle plates) should 1974

be undertaken for the full duration of critical processing, including equipment (aseptic set-up) 1975

assembly and filling operations. A similar approach should be considered for Grade B cleanrooms 1976

based on the risk of impact on the aseptic processing. The monitoring should be performed in such a 1977

way that all interventions, transient events and any system deterioration would be captured and any 1978

risk caused by interventions of the monitoring operations is avoided. 1979

1980

9.28 The adoption of suitable rapid or automated monitoring systems should be considered by 1981

manufacturers in order to expedite the detection of microbiological contamination issues and to 1982

reduce the risk to product. These rapid and automated microbial monitoring methods may be adopted 1983

after validation has demonstrated their equivalency or superiority to the established methodology. 1984

1985

9.29 Sampling methods and equipment used should be fully understood and procedures should be in 1986

place for the correct operation and interpretation of results obtained. The recovery efficiency of the 1987

sampling methods chosen should be qualified. 1988

1989

9.30 Action limits for viable particle contamination are shown in Table 7 1990

1991

Table 7: Maximum action limits for viable particle contamination 1992

1993

Grade

Air sample

cfu/m

Settle plates

(diam. 90 mm)

cfu/4 hours

(a)

Contact plates

(diam. 55mm),

cfu/ plate

(c)

Glove print,

Including 5 fingers on

both hands

cfu/ glove

No growth

(b)

B 10 5 5 5

C 100 50 25 -

D 200 100 50 -

1994

(a)

Settle plates should be exposed for the duration of operations and changed as required after 1995

4 hours (exposure time should be based on validation including recovery studies and it should 1996

not have any negative effect on the suitability of the media used). Individual settle plates may 1997

be exposed for less than 4 hours. 1998

(b)

It should be noted that for Grade A, any growth should result in an investigation. 1999

2000

(c)

Contact plate limits apply to equipment room and gown surfaces within the Grade A zone 2001

and Grade B area. Routine gown monitoring is not normally required for Grade C and D areas, 2002

depending on their function. 2003

2004

Note 1: It should be noted that the types of monitoring methods listed in the table above are 2005

examples and other methods can be used provided they meet the intent of providing 2006

information across the whole of the critical process where product may be contaminated (e.g. 2007

aseptic line set-up, filling and lyophilizer loading). 2008

2009

Note 2: Limits are applied using cfu throughout the document. If different or new technologies 2010

are used that present results in a manner different from cfu, the manufacturer should 2011

scientifically justify the limits applied and where possible correlate them to cfu. 2012

2013

9.31 Microorganisms detected in Grade A zone and Grade B area should be identified to species level 2014

and the potential impact of such microorganisms on product quality (for each batch implicated) and 2015

overall state of control should be evaluated. Consideration should also be given to the identification of 2016

microorganisms detected in Grade C and D areas (for example where action limits or alert levels are 2017

exceeded or where atypical or potentially objectionable microorganisms are recovered). The approach 2018

to organism identification and investigation should be documented. 2019

2020

9.32 Personnel gloves (and any part of the gown that may potentially have direct impact on the 2021

product sterility (e.g. the sleeves if these enter a critical zone) should be monitored for viable 2022

contamination after critical operations and on exit from the cleanroom. Other surfaces should be 2023

monitored at the end of an operation. 2024

2025

9.33 Microbial monitoring of personnel in the Grade A zone and Grade B area should be performed to 2026

assess their aseptic behaviour. Where filling operations are manual in nature e.g. hand filling, the 2027

process in its entirety may be considered as one critical intervention. In these cases, the frequency of 2028

microbial monitoring of gowning should be based on scientific principles and justified as part of the 2029

CCS. Where monitoring is routinely performed by manufacturing personnel, consideration should be 2030

given to periodic monitoring under the supervision of the quality unit. 2031

2032

Aseptic process simulation (APS) (also known as media fill) 2033

2034

9.34 Periodic verification of the effectiveness of the controls in place for aseptic processing should 2035

include a process simulation test using a sterile nutrient media and/or surrogate in place of the 2036

product. Selection of an appropriate nutrient media should be made based on the ability of the media 2037

and/or surrogate to imitate product characteristics at all processing stages. Where processing stages 2038

may indirectly impact the viability of any introduced microbial contamination, (e.g. sterile aseptically 2039

produced semi-solids, powders, solid materials, microspheres, liposomes and other formulations 2040

where product is cooled or heated or lyophilized), alternative procedures that represent the operations 2041

as closely as possible can be developed and justified. Where surrogate materials, such as buffers, are 2042

used in parts of the process simulation, the surrogate material should not inhibit the growth of any 2043

potential contamination. 2044

2045

9.35 The process simulation test should imitate as closely as possible the routine aseptic 2046

manufacturing process and include all the critical manufacturing steps, specifically: 2047

2048

i. Process simulation tests should assess all aseptic operations performed subsequent to the 2049

sterilization and decontamination cycles of materials utilised in the process to the point 2050

where the container is sealed. 2051

2052

ii. For non-filterable formulations, any additional aseptic steps should be assessed. 2053

2054

iii. Where aseptic manufacturing is performed under an inert atmosphere, the inert gas should 2055

be substituted with air in the process simulation unless anaerobic simulation is intended. 2056

2057

iv. Processes requiring the addition of sterile powders should use an acceptable surrogate 2058

material in containers identical to those used in the process under evaluation. 2059

2060

v. Separate simulations of individual unit operations (e.g. processes involving drying, 2061

blending, milling and subdivision of a sterile powder) should generally be avoided. Any use 2062

of individual simulations should be supported by a documented justification and ensure that 2063

the sum total of the individual simulations continues to fully cover the whole process. 2064

2065

vi. The process simulation procedure for lyophilized products should represent the entire 2066

aseptic processing chain including filling, transport, loading, chamber dwell, unloading and 2067

sealing under specified, documented and justified conditions representing worst case 2068

operating parameters. 2069

2070

vii. The lyophilization process simulation should duplicate all aspects of the process, except 2071

those that may affect the viability or recovery of contaminants. For instance, boiling-over or 2072

actual freezing of the solution should be avoided. Factors to consider in determining APS 2073

design include, where applicable: 2074

2075

• The use of air to break vacuum instead of nitrogen. 2076

2077

• Replicating the maximum interval between sterilization of the lyophilizer and its 2078

use. 2079

2080

• Replicating the maximum period of time between sterilization and lyophilization. 2081

2082

• Quantitative aspects of worst case situations, e.g. loading the largest number of 2083

trays, replicating the longest duration of loading where the chamber is open to the 2084

environment. 2085

2086

9.36 The process simulation testing should take into account various aseptic manipulations and 2087

interventions known to occur during normal production as well as worst case situations, including: 2088

2089

i. Inherent interventions representative of the routine process at the maximum accepted 2090

frequency per number of filled units (e.g. loading of vials into a lyophilizer). 2091

2092

ii. Corrective interventions, that occur frequently during routine production, in a representative 2093

number and with the highest degree of acceptable intrusion (e.g. correcting jammed 2094

stoppers). 2095

2096

9.37 Interventions should not be designed or selected to justify poor process or facility design or to 2097

assess unacceptable interventions that rarely occur and which should lead to a thorough investigation 2098

and product assessment when they do occur. 2099

2100

9.38 In developing the process simulation test plan, consideration should be given to the following: 2101

2102

i. Identification of worst case conditions covering the relevant variables, such as container size 2103

and line speed, and their impact on the process. The outcome of the assessment should 2104

justify the variables selected. 2105

2106

ii. Determining the representative sizes of container/closure combinations to be used for 2107

validation. Bracketing or matrix approach may be considered for validation of the same 2108

container/closure configuration for different products where process equivalence is 2109

scientifically justified. 2110

2111

iii. The volume filled per container, which should be sufficient to ensure that the media contacts 2112

all equipment and component surfaces that may directly contaminate the sterile product. The 2113

volume used should provide sufficient headspace to support potential microbial growth and 2114

ensure that turbidity can be detected during inspection. 2115

2116

iv. Maximum permitted holding times for sterile product and associated sterile components and 2117

equipment exposed during the aseptic process. 2118

2119

v. The method of detection of microbial contamination should be scientifically justified to 2120

ensure that any contamination is detectable. 2121

2122

vi. The selected nutrient media should be capable of growing a designated group of reference 2123

microorganisms as described by the relevant pharmacopeia and suitably representative local 2124

isolates and supporting recovery of low numbers of these microorganisms. 2125

2126

vii. The requirement for substitution of any inert gas used in the routine aseptic

manufacturing 2127

process by air unless anaerobic simulation is intended. In these situations, inclusion of 2128

occasional anaerobic simulations as part of the overall validation strategy should be 2129

considered (refer to paragraph 9.35 point iii). 2130

2131

viii. The process simulation should be of sufficient duration to challenge the process, the 2132

operators that perform interventions, shift changes and the capability of the processing 2133

environment to provide appropriate conditions for the manufacture of a sterile product. 2134

2135

ix. Where the manufacturer operates different shifts then the APS should be designed to capture 2136

specific factors (e.g. for those manufacturing during a night or extended shift, fatigue should 2137

be considered). 2138

2139

x. Simulating normal aseptic manufacturing interruptions where the process is idle (e.g. shift 2140

changeovers, recharging dispensing vessels, introduction of additional equipment, etc.). 2141

2142

xi. Ensuring that environmental monitoring is conducted as required for routine production, and 2143

throughout the entire duration of the process simulation. 2144

2145

xii. Where campaign manufacturing occurs, such as in the use of Barrier Technologies or 2146

manufacture of sterile active substances, consideration should be given to designing and 2147

performing the process simulation so that it simulates the risks associated with both the 2148

beginning and the end of the campaign and demonstrating that the campaign duration does 2149

not pose any risk. The performance of "end of production or campaign APS" may be used as 2150

additional assurance or investigative purposes; however, their use should be justified in the 2151

CCS and should not replace routine APS. If used, it should be demonstrated that any 2152

residual product does not negatively impact the recovery of any potential microbial 2153

contamination. 2154

2155

9.39 For sterile active substances, batch size should be large enough to represent routine operation, 2156

simulate intervention operation at the worst case, and cover potential contact surfaces. In addition, all 2157

the simulated materials (surrogates or growth medium) should be subjected to microbial evaluation. 2158

The simulation materials should be sufficient to satisfy the evaluation of the process being simulated 2159

and should not compromise the recovery of micro-organisms. 2160

2161

9.40 Process simulation tests should be performed as part of the initial validation, with at least three 2162

consecutive satisfactory simulation tests that cover all working shifts that the aseptic process may 2163

occur in, and after any significant modification to operational practices, facilities, services or 2164

equipment (e.g. modification to the HVAC system, equipment, major facility shut down, changes to 2165

process, number of shifts and numbers of personnel etc.). Normally, process simulation tests (periodic 2166

revalidation) should be repeated twice a year (approximately every six months) for each aseptic 2167

process, each filling line and each shift. Each operator should participate in at least one successful 2168

APS annually. Consideration should be given to performing an APS after the last batch prior to shut 2169

down, before long periods of inactivity or before decommissioning or relocation of a line. 2170

2171

9.41 Where manual operation (e.g. aseptic compounding or filling) occurs, each type of container, 2172

container closure and equipment train should be initially validated with each operator participating in 2173

at least 3 consecutive successful APS and revalidated with one APS approximately every 6 months for 2174

each shift. The APS batch size should mimic that used in the routine aseptic manufacturing process. 2175

2176

9.42 The number of units processed (filled) for APS tests should be sufficient to effectively simulate 2177

all activities that are representative of the aseptic manufacturing process. Justification for the number 2178

of units to be filled should be clearly captured in the PQS. Typically, a minimum of 5000 to 10000 2179

units are filled. For small batches (e.g. those under 5000 units), the number of containers for media fill 2180

should at least equal the size of the production batch. 2181

2182

9.43 Filled APS units should be agitated, swirled or inverted before incubation to ensure contact of 2183

the media with all interior surfaces in the container. Units with cosmetic defects or those who have 2184

gone through non-destructive in process control checks should be identified and incubated. Units 2185

discarded during the process simulation and not incubated should be comparable with units discarded 2186

during a routine fill. Examples may include those normally discarded after the set-up process or due to 2187

an intervention or where the integrity of the unit is compromised as would be identified by the routine 2188

inspection process for the product. 2189

2190

9.44 Where processes have materials that contact the product contact surfaces but are then discarded, 2191

the discarded material should be simulated with nutrient media and be incubated as part of the APS, 2192

unless it can be clearly demonstrated that this waste process would not impact the sterility of the 2193

product. 2194

2195

9.45 Filled APS units should be incubated in a clear container to ensure visual detection of microbial 2196

growth. Where the product container is not clear (i.e. amber glass, opaque plastic), clear containers of 2197

identical configuration may be substituted to aid in the detection of contamination. When a clear 2198

container of identical configuration cannot be substituted, a suitable method for the detection of 2199

microbial growth should be developed and validated. Microorganisms isolated from contaminated 2200

units should be identified to at least genus, and to the species level when practical, to assist in the 2201

determination of the likely source of the contaminant. The selection of the incubation conditions and 2202

duration should be scientifically justified and validated to provide an appropriate level of sensitivity 2203

of detection of microbial contamination. 2204

2205

9.46 Filled APS units should be incubated without unnecessary delay to achieve the best possible 2206

recovery of potential contamination. 2207

2208

9.47 On completion of incubation: 2209

2210

i. Filled APS units should be inspected by staff, who have been trained and qualified in the 2211

visual inspection procedures, under conditions similar to those for visual inspection, that 2212

facilitate the identification of any microbial contamination. 2213

2214

ii. Samples of these units should undergo positive control by inoculation with a suitable range of 2215

reference organisms and local isolates. 2216

2217

9.48 The target should be zero growth. Any contaminated unit should result in a failed process 2218

simulation and the following actions should occur: 2219

2220

i. An investigation to determine the most probable root causes. 2221

2222

ii. Determination and implementation of appropriate corrective measures. 2223

2224

iii. A sufficient number of successful, consecutive repeat media fills (normally a minimum of 3) 2225

should be conducted in order to demonstrate that the process has been returned to a state of 2226

control. 2227

2228

iv. A prompt review of all appropriate records relating to aseptic production since the last 2229

successful APS. The outcome of the review should include a risk assessment of potential 2230

sterile breaches in batches manufactured since the last successful process simulation. All other 2231

batches not released to the market should be included in the scope of the investigation. Any 2232

decision regarding their release status should consider the investigation outcome. 2233

2234

v. All products that have been manufactured on a line subsequent to a process simulation failure 2235

should be quarantined until a successful resolution of the process simulation failure has 2236

occurred. 2237

2238

vi. Production should resume only after completion of successful revalidation. 2239

2240

9.49 APS should be carefully observed by personnel with specific expertise in aseptic processing to 2241

assess the correct performance of operations and address inappropriate practices if detected. 2242

9.50 Where results indicate that an operator may have failed qualification, actions to limit the 2243

operator’s activities, until retrained and requalified, should be taken. 2244

2245

9.51 An aseptic process or filling should be subject to a repeat of the initial validation when: 2246

2247

i. The specific aseptic process has not been in operation for an extended period of time. 2248

2249

ii. There is a change to the process, equipment, procedures or environment that has the 2250

potential to affect the aseptic process or an addition of new product containers or container-2251

closure combinations. 2252

2253

9.52 All process simulation runs should be fully documented and include a reconciliation of units 2254

processed (e.g. units filled, incubated, not incubated, and rejected). All interventions performed 2255

during the process simulations should be recorded, including the start and end of each intervention. 2256

All microbial monitoring data as well as other testing data should be recorded in the APS batch 2257

record. 2258

2259

10 Quality Control (QC) 2260

2261

10.1 It is important that there are personnel with appropriate training and experience in microbiology 2262

and knowledge of the process to support the design of the manufacturing process, environmental 2263

monitoring regime and any investigation assessing the impact of microbiologically linked events to 2264

the safety of the sterile product. 2265

2266

10.2 Specifications for raw materials, components and products should include requirements for 2267

microbial quality when the need for this has been indicated by monitoring and/or by the CCS. 2268

2269

10.3 The bioburden assay should be performed on each batch for both aseptically filled product and 2270

terminally sterilized products and the results considered as part of the final batch review. There should 2271

be defined limits for bioburden immediately before the sterilizing filter or the terminal sterilization 2272

process, which are related to the efficiency of the method to be used. Samples should be taken to be 2273

representative of the worst case scenario (e.g. at the end of hold time). Where overkill sterilization 2274

parameters are set for terminally sterilized products, bioburden should be monitored at suitable 2275

scheduled intervals. 2276

2277

10.4 A pre-sterilization bioburden monitoring program for the product and components should be 2278

developed to support parametric release. The bioburden should be performed for each batch. The 2279

sampling locations of filled units before sterilization should be based on a worst case scenario and be 2280

representative of the batch. Any organisms found during bioburden testing should be identified and 2281

their impact on the effectiveness of the sterilizing process determined. Where appropriate, the level of 2282

pyrogen (endotoxins) should be monitored. 2283

2284

10.5 The sterility test applied to the finished product should only be regarded as the last in a series of 2285

control measures by which sterility is assured. It cannot be used to assure sterility of a product that 2286

does not meet its design, procedural or qualification parameters. The test should be validated for the 2287

product concerned. 2288

2289

10.6 The sterility test should be performed under aseptic conditions. Samples taken for sterility 2290

testing should be representative of the whole of the batch but should in particular include samples 2291

taken from parts of the batch considered to be most at risk of contamination, for example: 2292

2293

i. For products which have been filled aseptically, samples should include containers filled at

2294

the beginning, middle and end of the batch and after any significant intervention (e.g. 2295

interventions where the integrity of a barrier is breached (open door)) or an operator 2296

intervention into critical zones. 2297

2298

ii. For products which have been heat sterilized in their final containers, samples taken 2299

should

representative of the worst case locations (e.g. the potentially coolest or slowest to 2300

heat part of each load). 2301

2302

iii. For products that are lyophilized, samples taken from different lyophilization loads. 2303

2304

Note: Where the manufacturing process results in sub-batches (e.g. for terminally sterilized 2305

products) then sterility samples from each sub-batch should be taken and a sterility test for each sub-2306

batch performed. Consideration should also be given to performing separate testing for other 2307

finished product tests. 2308

2309

10.7 For some products it may not be possible to perform a sterility test prior to release because the 2310

shelf life of the product is too short to allow completion of a sterility test. In these cases, the CCS 2311

should clearly capture the identified risks, the additional considerations of design of the process and 2312

additional monitoring required to mitigate the identified risks. 2313

2314

10.8 Any process (e.g. Vaporized Hydrogen Peroxide or VH202, Ultra Violet) used to decontaminate 2315

the external surfaces of sterility samples prior to testing should not negatively impact the sensitivity of 2316

the test method. 2317

2318

10.9 Media used for environmental monitoring and APS should be tested for its growth promotion 2319

capability, in accordance with a formal written program. 2320

2321

10.10 Environmental monitoring data and trend data generated for classified areas should be reviewed 2322

as part of product batch certification. A written plan should be available that describes the actions to 2323

be taken when data from environmental monitoring are found out of trend or exceeding the 2324

established limits. For products with short shelf life, the environmental data for the time of 2325

manufacture may not be available; in these cases, the certification should include a review of the most 2326

recent available data. Manufacturers of these products should consider the use of rapid monitoring 2327

systems. 2328

2329

10.11 Where rapid and automated microbial methods are used for general manufacturing purposes, 2330

these methods should be validated for the product(s) or processes concerned. 2331

2332

Glossary 2333

2334

Airlock – An enclosed space with interlocked doors, constructed to maintain air pressure control 2335

between adjoining rooms (generally with different air cleanliness standards). The intent of an airlock 2336

is to preclude ingress of particulate matter and microorganism contamination from a lesser controlled 2337

area. 2338

2339

Action limit – An established relevant measure (e.g. microbial, or airborne particulate limits) that, 2340

when exceeded, should trigger appropriate investigation and corrective action based on the 2341

investigation. 2342

2343

Alert level – An established relevant measure (e.g. microbial, or airborne particulate levels) giving 2344

early warning of potential drift from normal operating conditions and validated state, which does not 2345

necessarily give grounds for corrective action but triggers appropriate scrutiny and follow-up to 2346

address the potential problem. Alert levels are established based on historical and qualification trend 2347

data and periodically reviewed. The alert level can be based on a number of parameters including 2348

adverse trends, individual excursions above a set limit and repeat events. 2349

2350

Aseptic processing room – A room in which one or more aseptic activities or processes are performed. 2351

2352

Aseptic Process Simulation (APS) –A simulation of the entire aseptic formulation and filling process 2353

in order to determine the capability of the process to assure product sterility. 2354

2355

Asepsis – A state of control attained by using an aseptic work area and performing activities in a 2356

manner that precludes microbial contamination of the exposed sterile product. 2357

2358

Bacterial retention testing – This test is performed to validate that a filter can remove bacteria from a 2359

gas or liquid. The test is usually performed using a standard organism, such as Brevundimonas 2360

diminuta at a minimum concentration of 10

Colony Forming Units/cm

. 2361

2362

Barrier – A physical partition that affords aseptic processing area (usually Grade A) protection by 2363

separating it from the background environment. Such systems frequently use in part or totally the 2364

Barrier Technologies known as RABS or isolators. 2365

2366

Bioburden – The total number of microorganisms associated with a specific item such as personnel, 2367

manufacturing environments (air and surfaces), equipment, product packaging, raw materials 2368

(including water), in-process materials, or finished products. 2369

2370

Biological Indicator (BI) – A population of microorganisms inoculated onto a suitable medium (e.g. 2371

solution, container or closure) and placed within a sterilizer or load or room locations to determine the 2372

sterilization or disinfection cycle efficacy of a physical or chemical process. The challenge 2373

microorganism is selected and validated based upon its resistance to the given process. Incoming lot 2374

D value, microbiological count and purity define the quality of the BI. 2375

2376

Blow-Fill-Seal (BFS) – A technology in which containers are formed from a thermoplastic granulate, 2377

filled with product, and then sealed in a continuous, integrated, automatic operation. The two most 2378

common types of BFS machines are the Shuttle type (with Parison cut) and the Rotary type (Closed 2379

Parison) types. 2380

2381

Classified area – An area that contains a number of cleanrooms (see cleanroom definition). 2382

2383

Cleaning – A process for removing contamination e.g. product residues and disinfectant residues. 2384

2385

Clean area – An area with defined particulate and microbiological cleanliness standards usually 2386

containing a number of joined cleanrooms. 2387

2388

Cleanroom – A room designed, maintained, and controlled to prevent particulate and microbial 2389

contamination of drug products. Such a room is assigned and reproducibly meets an appropriate air 2390

cleanliness level. Grade A will be referred to as Grade A zone. 2391

2392

Cleanroom classification – A method of assessing the level of air cleanliness against a specification 2393

for a cleanroom or clean air equipment by measuring the non-viable airborne particulate 2394

concentration. 2395

2396

Cleanroom qualification – A method of assessing the level of compliance of a classified cleanroom or 2397

clean air equipment with its intended use. 2398

2399

Closed system – A system in which the sterile product is not exposed to the surrounding environment. 2400

For example, this can be achieved by the use of bulk products holders (such as tanks or bags) that are 2401

connected to each other by pipes or tubes as a system, with the system being sterilized after the 2402

connections are made. Examples of these can be (but are not limited to) large scale reusable systems, 2403

such as those seen in active substance manufacturing, or disposable bag and manifold systems, such 2404

as those seen in the manufacture of biological products. Closed systems, when used in this document, 2405

does not refer to systems such as RABS or isolator systems which are referred to as Barrier 2406

Technologies. 2407

2408

Colony Forming Unit (CFU) – A microbiological term that describes a single detectable colony that 2409

originates from one or more microorganisms. Colony forming units are typically expressed as cfu per 2410

ml for liquid samples, and cfu per cm

for samples captured on solid medium such as settle or contact 2411

plates. 2412

2413

Contamination – The undesired introduction of impurities of a microbiological nature (quantity and 2414

type of microorganisms, pyrogens), or of foreign particulate matter, into or onto a raw material, 2415

intermediate, active substance or drug product during production, sampling, packaging or 2416

repackaging, storage or transport with the potential to adversely impact product quality. 2417

2418

Contamination Control Strategy (CCS) – A planned set of controls for microorganisms, pyrogens and 2419

particulates, derived from current product and process understanding that assures process performance 2420

and product quality. The controls can include parameters and attributes related to active substance, 2421

excipient and drug product materials and components, facility and equipment operating conditions, in-2422

process controls, finished product specifications, and the associated methods and frequency of 2423

monitoring and control. 2424

2425

Corrective intervention – An intervention that is performed to correct or adjust an aseptic process 2426

during its execution. These may not occur with the same frequency in the routine aseptic process. 2427

Examples include such as clearing component jams, stopping leaks, adjusting sensors, and replacing 2428

equipment components. Corrective measures should be taken to reduce their extent and frequency. 2429

2430

Critical surfaces – Surfaces that may come directly into contact with, or directly affect, a sterile 2431

product or its containers or closures. Critical surfaces are rendered sterile prior to the start of the 2432

manufacturing operation, and sterility is maintained throughout processing. 2433

2434

Critical zone – A location within the aseptic processing area in which product and critical surfaces are 2435

exposed to the environment. 2436

2437

Critical intervention – An intervention (corrective or inherent) into the critical zone. 2438

2439

D value – The value of a parameter of sterilization (duration or absorbed dose) required to reduce the 2440

number of viable organisms to 10 per cent of the original number. 2441

2442

Dead leg – Length of non-circulating pipe (where fluid may remain static) that is greater than 3 2443

internal pipe diameters. 2444

2445

Decommission – When a process, equipment or cleanroom are closed where they will not be used 2446

again. 2447

2448

Decontamination – The overall process of removal or reduction of any contaminants (chemical, waste, 2449

residue or microorganisms) from an area, object, or person. The method of decontamination used (e.g. 2450

cleaning, disinfection, sterilization) should be chosen and validated to achieve a level of cleanliness 2451

appropriate to the intended use of the item decontaminated. 2452

2453

Depyrogenation – A process designed to remove or inactivate pyrogenic material (e.g. endotoxins) to 2454

a specified minimum quantity. 2455

2456

Disinfection – The process by which the reduction of the number of microorganisms is achieved by 2457

the irreversible action of a product on their structure or metabolism, to a level judged to be 2458

appropriate for a defined purpose. 2459

2460

Endotoxin – A pyrogenic product (e.g. lipopolysaccharide) present in the bacterial cell wall. 2461

Endotoxin can lead to reactions in patients receiving injections ranging from fever to death. 2462

2463

Extractables - Chemical entities that migrate from the surface of the process equipment, exposed to an 2464

appropriate solvent at extreme conditions, into the product or material being processed. 2465

2466

First Air – Refers to filtered air that has not been interrupted by items (such as operators) with the 2467

potential to add contamination to the air prior to reaching the critical zone. 2468

2469

Form-Fill-Seal (FFS) – Similar to Blow fill Seal, this involves the formation of a large tube formed 2470

from a flexible packaging material, in the filling machine, and generally the tube is filled to form the 2471

bags. 2472

2473

Gowning qualification – A program that establishes, both initially and on a periodic basis, the 2474

capability of an individual to don the complete sterile gown in an aseptic manner. 2475

2476

Grade A air supply – Air which is passed through a filter qualified as capable of producing Grade A 2477

non-viable quality air, but where there is no requirement to perform continuous non-viable monitoring 2478

or meet Grade A viable monitoring limits and the area itself is not classified. Specifically used for the 2479

protection of fully stoppered vials where the cap has not been crimped and the equipment and 2480

engineering systems that have a direct impact on product quality. 2481

2482

HEPA filter - High efficiency particulate air filter with 0.3 µm particulate retaining efficiency of no 2483

less than 99.95 percent according to the relevant norms (e.g. EN 1822).. 2484

2485

Inherent interventions – An intervention that is an integral part of the aseptic process and is required 2486

for either set-up, routine operation and/or monitoring (e.g. aseptic assembly, container replenishment, 2487

environmental sampling, etc.). Inherent interventions are required by procedure or work instruction 2488

for the execution of the aseptic process. 2489

2490

Integrity test - A test to confirm that a filter (product, gas or HVAC filter) retain their retentive 2491

properties and have not been damaged during handling, installation or processing. 2492

2493

Intrinsic Sterile Connection device – A device that reduces the risk of contamination during the 2494

connection process; these can be mechanical or fusion sealing. 2495

2496

Isokinetic sampling head – A sampling head designed to disturb the air as little as possible so that the 2497

same particulates go into the nozzle as would have passed the area if the nozzle had it not been there 2498

i.e. the sampling condition in which the mean velocity of the air entering the sample probe inlet is 2499

nearly the same (± 20 percent) as the mean velocity of the airflow at that location. 2500

2501

Isolator – A decontaminated unit, with an internal work zone meeting Grade A conditions that 2502

provides uncompromised, continuous isolation of its interior from the external environment (e.g. 2503

surrounding cleanroom air and personnel). There are two major types of isolators 2504

2505

i. Closed isolator systems exclude external contamination of the isolator’s interior by 2506

accomplishing material transfer via aseptic connection to auxiliary equipment, rather than 2507

use of openings to the surrounding environment. Closed systems remain sealed throughout 2508

operations. 2509

ii. Open isolator systems are designed to allow for the continuous or semi-continuous ingress 2510

and/or egress of materials during operations through one or more openings. Openings are 2511

engineered (e.g. using continuous overpressure) to exclude the entry of external contaminant 2512

into the isolator. 2513

2514

Leachables – Chemical entities that migrate into products from the product contact surface of the 2515

process equipment or containers under normal condition of use and/or storage. 2516

2517

Local Isolates – Suitably representative microorganisms of the site that are frequently recovered 2518

through environmental monitoring within the classified zone/areas especially Grade A zone and 2519

Grade B area, personnel monitoring or positive sterility test results. 2520

2521

Lyophilization – A physical-chemical drying process designed to remove solvents, by way of 2522

sublimation, from both aqueous and non-aqueous systems, primarily to achieve product or material 2523

stability. Lyophilization is synonymous to the term freeze-drying. 2524

2525

Manual Filling – A filling process where operator intervention is required to complete the filling of 2526

each container (e.g. as occurs during aseptic compounding operations). 2527

2528

Operator - Any individual participating in the processing operation, including line set-up, filler, 2529

maintenance, or other personnel associated with manufacturing activities. 2530

2531

Overkill sterilization – A process that is sufficient to provide at least a 12 log reduction of 2532

microorganisms having a minimum D value of 1 minute. 2533

2534

Pass-through hatch – Synonymous with airlock (refer to airlock definition) but typically smaller in 2535

size. 2536

2537

Pyrogen – A substance that induces a febrile reaction in a patient. 2538

2539

Rapid transfer system (RTP) – A System used for the transfer of items into RABS and isolators that 2540

minimize the risk to the critical zone. An example would be a rapid transfer container with an 2541

alpha/beta port. 2542

2543

Raw material – Any ingredient intended for use in the manufacture of a sterile product, including 2544

those that may not appear in the final drug product. 2545

2546

Restricted Access Barrier System (RABS) – System that provides an enclosed, but not sealed, 2547

environment meeting defined cleanroom conditions (for aseptic processing Grade A, (but where used 2548

for non-sterile applications can be lesser grade) and using a rigid-wall enclosure and air overspill to 2549

separate its interior from the surrounding environment. The inner surfaces of the RABS are 2550

disinfected and decontaminated with a sporicidal agent. Operators use gloves, half suits, rapid transfer 2551

systems (RTPs) and other integrated transfer ports to perform manipulations or convey materials to 2552

the interior of the RABS. Depending on the design, doors are rarely or never opened: 2553

2554

i. Active RABS: integral HEPA-filtered air supply. 2555

2556

ii. Passive RABS: air supply by ceiling mounted HEPA-filters. 2557

2558

iii. Closed RABS: where the air is vented in return ducts within the cabinet. 2559

2560

iv. Open RABS: Where there are vents in the barrier that allow air to move from the Grade A 2561

zone to the Grade B area. 2562

2563

Single Use Systems (SUS) – Systems in which product contact components are used only once (i.e. 2564

single use components) to replace reusable equipment such as stainless steel transfer lines or bulk 2565

containers. SUS covered in this document are those that are used in manufacturing processes of sterile 2566

products (e.g. sterile active substance, sterile bio bulk, sterile finished dosage), and are typically made 2567

up of disposable components such as bags, filters, tubing, connectors, storage bottles and sensors. 2568

2569

Sporicidal agent – An agent that destroys bacterial and fungal spores when used in sufficient 2570

concentration for specified contact time. It is expected to kill all vegetative microorganisms. 2571

2572

Sterile Product – For purpose of this guidance, sterile product refers to one or more of the sterilized 2573

elements exposed to aseptic conditions and ultimately making up the sterile active substance or 2574

finished sterile product. These elements include the containers, closures, and components of the 2575

finished drug product. Or, a product that is rendered sterile by a terminal sterilization process. 2576

2577

Sterilizing grade filter – A filter that, when appropriately validated, will remove a defined microbial 2578

challenge from a fluid or gas producing a sterile effluent. Usually such filters have a pore size equal or 2579

less than 0.22 µm (for the purposes of this document 0.2 µm and 0.22 µm are used interchangeably 2580

and deemed equivalent). 2581

2582

Terminal Sterilization – The application of a lethal sterilizing agent or conditions to a product within a 2583

sealed container to achieve a predetermined sterility assurance level (SAL) of 10⁻⁶ or better (i.e. the 2584

theoretical probability of there being a single viable microorganism present on or in a sterilized unit is 2585

equal to or less than 1 x 10

-6

(one in a million)). 2586

2587

Turbulent airflow – Air that is not unidirectional. Turbulent air in cleanrooms should flush the 2588

cleanroom via mixed flow dilution and ensure maintenance of acceptable air quality. 2589

2590

Unidirectional airflow – An airflow moving in a single direction, in a robust and uniform manner, and 2591

at sufficient speed, to reproducibly sweep particulates away from the critical processing or testing 2592

area. 2593

2594

Unidirectional Airflow Unit (UDAF) – A cabinet supplied with filtered unidirectional airflow 2595

(previously referred to as a Laminar Airflow Unit or LAF). 2596

2597

Vertical-Form-Fill-Seal (VFFS) – An automated filling process, typically for terminally sterilized 2598

products, that may utilize a single or dual web system which constructs the primary container out of a 2599

flat roll of thermoplastic film while simultaneously filling the formed bags with product and sealing 2600

the filled bags in a continuous process. 2601

2602

Worst case – A set of conditions encompassing processing limits and circumstances, including those 2603

within standard operating procedures, that pose the greatest chance of process or product failure 2604

(when compared with ideal conditions). Such conditions have the highest potential to, but do not 2605

necessarily always induce, product or process failure. 2606

2607

Water system – A system for producing, storing and distributing water, usually compliant to a specific 2608

pharmacopeia grade e.g. purified and water for injection (WFI). 2609

2610