在国际无菌制药行业，有2部关键法规和指南被尊为圣经，其中一部是FDA发布的无菌工艺指南，另外一部就是EU GMP附录1。经过漫长的修订期，在2022年8月，EU GMP附录1修改工作终于结束，定稿版本发布。这份新的GMP指南影响很大，因此为了让企业有适应期，设定生效日期是2023年8月。

       在近期，PIC/S也根据已经发布的EU GMP附录1发布了自己的GMP指南的附录1。而中国已经在2021年提出加入PIC/S的申请，可以预见的是，EU GMP附录1对中国制药行业的影响也会不断加强。

       为了帮助国内无菌企业更快的熟悉EU GMP附录1的具体要求，并加强拟进入国际市场的无菌制药企业的质量提升工作，特梳理和分析EU GMP附录1中的强制性条款，帮助企业技术人员更快理解和熟悉这份关键法规的要求。

       说明：

       1- 下面为了行文流畅，从EU GMP附录1中摘引的原文，只是引用强制性和要求非常具体的段落；无关内容没有被引用。

       2- 下面内容只是引用了部分EU GMP附录1的内容，不是全部内容。

       第一部分：厂房设施部分的强制性条款

       4.4-The maintenance of unidirectional airflow should be demonstrated and qualified across the whole of the grade A area.

       解析：应该在整个A级区域内部证明和确认对单向流的维护。通过这句话可以看出，A级区域的内部验证和测试不能只测试某个位置，应该基于A级区域面积和A级区域内部的关键工位的评估，来选择足够测试点，来证明A级区域内部的单向流。

       4.4-Air pressure differences should be continuously monitored.

       解析：应该对压差进行持续监控。从这句可以看出，应该对关键位置压差，进行持续监控。这些关键位置的压差指的是A级-B级区域之间的压差，以及B级-C级区域之前的压差。

       4.6-Sliding doors may be undesirable for this reason.

       解析：在无菌车间不能使用推拉门，也不能使用悬吊门。因为这样类型的门，存在更多不易清洁的死角。

       4.9-Sinks and drains should be prohibited in the grade A and grade B areas.

       解析：在A级和B级区域不能出现水池和地漏。这个问题，不需要解释，这是GMP基本原则和无菌制药特征决定的。

       4.10-The transfer of materials, equipment, and components into the grade A or B areas should be carried out via a unidirectional process. The removal of items from the grade A and B areas (e.g. materials, waste, environmental samples) should be carried out via a separate unidirectional process.

       解析：将物料、设备和部件传递进入A级区域或者B级区域，应该采用单向流程。将物料、废弃物、环境监控样品从A级区域和B级区域转移出来时，应该从一个单独的单向流通道转移出来。从这句要求可以看出，进入A/B级区域的通道应该单独设置，进入通道和生产废弃物退出通道，应该分开。

       4.11-The final stage of the airlock should, in the "at rest" state, be of the same cleanliness grade (viable and total particle) as the cleanroom into which it leads.

       解析：人流物流通道最后一个阶段的房间，应该和拟进入区域在静态下保持相同的标准（微生物项目和悬浮粒子项目）。

       4.12-Personnel airlocks: Areas of increasing cleanliness used for entry of personnel (e.g. from the grade D area to the grade C area to the grade B area). In general hand washing facilities should be provided only in the first stage of the changing room and not be present in changing rooms directly accessing the grade B area.

       解析：人流通道气锁房间：这些气锁房间是用于人员进入的房间，采用逐级提升洁净度的方式来实现人员进入（例如从D级进入C级，然后再进入B级）。通常，洗手设施应该仅仅在更衣室通道的第一个房间设置，而不应该出现在紧靠B级区域的房间设置洗手装置。

       从上面这段要求，相信制药企业明白无菌车间更衣通道设置的一般性要求。虽然措辞没有使用必须，但是欧盟GMP附录1采用举例的方式告诉制药行业，这是最常见和最推荐的方式之一。

       4.12-Only materials and equipment that have been included on an approved list and assessed during validation of the transfer process should be transferred into the grade A or grade B areas via an airlock or pass-through hatches.

       解析：只有纳入批准清单的物料和设备才可以通过传递窗或者缓冲间传递进入A级或者B级区域。从这句可以看出，凡是进入A级或者B级区域的物料和设备，必须纳入一个特定的批准目录中。

       4.13-For pass-through hatches and airlocks (for material and personnel), the entry and exit doors should not be opened simultaneously.

       解析：对于人员和物料进出的传递窗和缓冲间，进出的门不能同时打开。这是为了控制污染提出的基本要求，不需要过多解释。

       4.14-Adjacent rooms of different grades should have an air pressure difference of a minimum of 10 Pascals (guidance value).

       解析：洁净区内部不同级别相邻房间之间的压差应该保持最低10Pa（指导值）。从这条可以看出，具体保持多大的压差数据，需要结合更具体的QRM来判断，而不是一律保持10Pa就算可以。

       4.15 Airflow patterns within cleanrooms and zones should be visualised to demonstrate that there is no ingress from lower grade to higher grade areas and that air does not travel from less clean areas (such as the floor) or over operators or equipment that may transfer contamination to the higher grade areas.

       解析：应该对洁净区和洁净室的气流模式进行可视化研究，以证明没有空气从低级别区域向高级别区域侵入，以及空气不会从较低洁净区域（例如地面）携带污染物越过人员或者设备向较高等级区域传递。

       从这句要求看，洁净区内部送风、回风、排风设置要非常精细的设计才可以实现这样的工艺目的。而且验证要求也提到很高程度，需要证明这些风险是不存在的。

       4.19-Negative pressure isolators should only be used when containment of the product is considered essential (e.g. radiopharmaceutical products) and specialized risk control measures should be applied to ensure the critical zone is not compromised.

       解析：当产品的污染风险（例如放 射 性药品）被认为是基本问题时，才允许使用负压隔离器，采用采用特定的风险控制措施，以确保关键区域不被污染。这句话描述了负压隔离器使用的特定情况。

       4.20-The background environment for open isolators should generally correspond to a minimum of grade C. The background for closed isolators should correspond to a minimum of grade D.

       解析：O-隔离器的背景环境通常最低是C级。而C-隔离器的背景环境通常最低是D级。这句描述了在为隔离器设置背景环境是的特定要求。

       4.21-Isolators-Generally glove integrity testing should be performed at a minimum frequency of the beginning and end of each batch or campaign.

       解析：对于隔离器上面的手套完整性测试最低频率是每个批次或者每个生产周期的开始和结束，各测试一次。

       RABS-Gloves should be visually examined with each use, and integrity testing should be performed at periodic intervals.

       解析：对于RABS，应该每次使用是目视检查手套的完整性，并且应该在周期性间隔测试手套完整性。

       4.30-Unidirectional airflow systems should provide a homogeneous air speed in a range of 0.36 - 0.54 m/s (guidance value) at the working position, unless otherwise scientifically justified in the CCS. Airflow visualization studies should correlate with the air speed measurement.

       解析：在工作位置，单向流系统应该提供均一的空气风速，范围是0.36 - 0.54 m/s，除非在CCS中对于风速另有论证。气流可视化研究应该和风速测量相互关联。

       通过上面这段描述，我们知道，具体单向流风速要求，应该和工艺和具体位置相关，而不是必须采用0.36 - 0.54 m/s。

       4.32-The maximum time interval for requalification of grade A & B areas, is 6 months. The maximum time interval for requalification of grade C & D areas, is 12 months.

       解析：A/B级区域的再确认周期最短是6个月。C/D级区域再确认周期最短是12个月。上面描述无菌制药企业对于洁净区再确认的最低周期要求。

       4.35 Disinfectants and detergents used in grade A and grade B areas should be sterile prior to use.

       解析：用于A级和B级区域的消毒剂和清洁剂应该在使用前灭菌或者除菌。从这条可以看出，A/B级是无菌区域，必须严格控制微生物污染。

       第二部分：设备部分的强制性条款

       5.2-The frequency at which alarms are assessed should be based on their criticality (with critical alarms reviewed immediately).

       解析：对于报警信息的审核应该基于它们的关键程度（关键报警信息必须立即审核）。从这段可以看出，如果制药公司对于某些报警信息列为关键信息，应该安排人员24小时值班来处理。

       5.8-A conveyor belt should not pass through a partition between a grade A or B area and a processing area of lower air cleanliness, unless the belt itself is continually sterilised (e.g. in a sterilising tunnel).

       解析：传送带不能在A级区域之外运行，或者不能在B级和较低级别区域之间运行，除非传送带自身可以持续灭菌，例如隧道烘箱。

       这句告诉我们，在设置传送带的时候，必须遵守这些基本原则。

       5.9-Tube length should typically be no longer than 1m unless justified and the number of bends should be minimized.

       解析：悬浮粒子计数器取样管长度通常不能超过1米，除非另有说明。而且，取样管上面的弯曲数量应该最小化。这个要求应该是行业共识了，请牢记山东省某企业被欧盟警告的案例。

       第三部分：公用系统部分的强制性条款

       6.10 Water for injections (WFI) should be produced from water meeting specifications that have been defined during the qualification process, stored and distributed in a manner which minimizes the risk of microbial growth (e.g. by constant circulation at a temperature above 70°C).

       解析：注射用水 (WFI) 应使用符合在确认过程中定义的标准的水生产，以将微生物生长风险降至最低的方式储存和分配（例如在高于 70°C 的温度下持续循环）。从这条可以看出，WFI循环温度是硬性要求。

       6.12-Chemical testing results should be approved before the water system is returned to use and microbiological/endotoxin results verified to be within specification and approved before batches manufactured using water from the system are considered for certification/release.

       解析：在水系统恢复使用之前，应该对于化学测试结果进行审核批准；使用相关工艺用水生产的药品在放行之前，微生物项目和内毒素项目应该被确认符合标准并得到批准。从这条要求看，应该对于WFI进行批次划分，在生产记录上面记录生产使用的WFI批号，并按照规定完成放行。

       6.13-Sample plans should be based on the qualification data, should consider the potential worst case sampling locations and should ensure that at least one representative sample is included every day of the water that is used for manufacturing processes.

       解析：对于工艺用水的取样计划应该基于确认数据，应该考虑潜在的最差取样位置，应该确保每天至少取样一个代表性的取样点，来评估用于生产的工艺用水的质量。根据上面这段要求，对于水系统取样计划，应该采用常见的3+N或者4+N方法。

       6.15 WFI systems should include continuous monitoring systems such as Total Organic Carbon (TOC) and conductivity, as these may give a better indication of overall system performance than discrete sampling.

       解析：应该对于WFI系统采用持续监控，测试TOC和电导率项目，因为这样可以及时反馈整个系统的性能，比离线测试要更好。从这句看，对于WFI系统，在线监控TOC和电导率是必须项目。

       第四部分：人员管理部分的强制性条款

       7.5 Unqualified personnel should not enter grade B cleanrooms or grade A in operation. If needed in exceptional cases, manufacturers should establish written procedures outlining the process by which unqualified personnel are brought into the grade B and A areas.

       解析：未经过无菌更衣确认的人员，不能进入A级或者B级区域。如果需要设置额外情况，制造商应该建立书面规程，来描述未经过更衣确认人员如何进入A/B级区域，如何被管控。

       第五部分：生产和特定技术部分的强制性条款

       8.3 Filling of products for terminal sterilisation should be carried out in at least a grade C environment.

       解析：最终灭菌产品的灌装区域应该至少设置在C级区域。

       8.22- Glass ampoules, BFS units and small volume containers (≤100 ml) closed by fusion should be subject to 100% integrity testing using validated methods.

       解析：采用融封工艺生产的玻璃安瓿、BFS工艺生产的单元包装、小容量容器（≤100ml）应该采用验证的方法进行100%的完整性检测。

       8.67- All air supplied to the tunnel should pass through at least a HEPA filter and periodic tests (at least biannually) should be performed to demonstrate air filter integrity.

       解析：针对干热灭菌，供应到隧道的所有空气都应至少通过HEPA，并且应进行定期测试（至少每两年一次）以证明空气过滤器的完整性。

       从这条可以看出，用于干热隧道烘箱的过滤器必须是HEPA，不能采用非HEPA。而且完整性测试频率也进行详细规定。

       8.68 When a thermal process is used as part of the depyrogenation process for any component or product contact equipment/material, validation studies should be performed to demonstrate that the process provides a suitable Fh value and results in a minimum 3 log10 reduction in endotoxin concentration.

       解析：当干热工艺被用作任何部件或产品接触设备/材料的去热原过程时，应进行验证研究以证明该工艺提供了合适的Fh值并导致内毒素浓度至少降低3个log10。

       8.79 If the product cannot be sterilised in its final container, solutions or liquids should be sterilised by filtration through a sterile sterilising grade filter (with a nominal pore size of a maximum of 0.22 μm that has been appropriately validated to obtain a sterile filtrate) and subsequently aseptically filled into a previously sterilised container.

       解析：如果产品不能在其最终容器中灭菌，溶液或液体应通过无菌级过滤器（标称孔径最大为0.22 μm，经适当验证以获得无菌滤液）过滤来除菌，随后填充到已经预先灭菌的容器中。

       第六部分：环境和工艺监控部分的强制性条款

       9.16 For grade A, particle monitoring should be undertaken for the full duration of critical processing, including equipment assembly.

       解析：对于A级区域，应在关键加工的整个过程中进行悬浮粒子监测，包括设备组装操作。

       9.17 The grade A area should be monitored continuously (for particles ≥0.5 and ≥5 μm) and with a suitable sample flow rate (at least 28 litres (1ft3) per minute) so that all interventions, transient events and any system deterioration is captured.

       解析：应连续监测A级区域（对于 ≥0.5um和 ≥5 μm的颗粒）并以合适的采用速度（至少28升（1ft3）每分钟的流速），以便捕获所有干预、瞬态事件和任何系统恶化信息。从这句可以看出，A级区域悬浮粒子监控的采样流速，需要符合上面的硬性要求。

       第七部分：质量控制部分的强制性条款

       10.3 The bioburden assay should be performed on each batch for both aseptically filled product and terminally sterilised products and the results considered as part of the final batch review.

       解析：对无菌灌装产品和最终灭菌产品的每批产品都应进行生物负荷量测定，并将结果视为最终批次审核的一部分。根据这句，应该在每批次无菌产品生产时，在合适位置取样测试生物负荷量。

       总结

       根据上面汇总和梳理的信息可以看出，刚发布的EU GMP附录1对于无菌药品提出了明显的严苛要求。因此说，想进军欧盟市场的中国企业，要提前准备，才可以取得预期的成绩。

       说明：本文不构成价值判断和投资建议。

       作者简介：zhulikou431，高级工程师、PDA会员、ISPE会员、ECA会员、PQRI会员、资深无菌GMP专家，在无菌工艺开发和验证、药品研发和注册、CTD文件撰写和审核、法规审计、国际认证、国际注册、质量体系建设与维护领域，以及无菌检验、环境监控等领域皆具有较深造诣。近几年开始着力关注制药宏观领域趋势分析和制药企业并购项目的风险管理工作。