培養(yǎng)基無(wú)菌模擬灌裝試驗(yàn)指南染菌測(cè)試
發(fā)布時(shí)間：
2022-11-16
作者：
微生物技術(shù)的品牌
培養(yǎng)基無(wú)菌模擬灌裝試驗(yàn)指南染菌測(cè)試
我做凍干粉注射劑生產(chǎn)工作有段時(shí)間了，培養(yǎng)基模擬罐裝實(shí)驗(yàn)多次進(jìn)行，模擬罐裝染菌也會(huì)發(fā)生，今天挑一個(gè)案例來(lái)談?wù)剬?duì)于預(yù)防污染的看法，希望與同行有很好的交流。
這家公司連續(xù)做了三批培養(yǎng)基無(wú)菌模擬灌裝，每批量為10000支，第一批培養(yǎng)培養(yǎng)不到兩天就長(zhǎng)菌了，第二、三批到培養(yǎng)結(jié)束都沒(méi)有檢出陽(yáng)性污染品。而且第一批有一半是長(zhǎng)菌的，問(wèn)我是那里出現(xiàn)問(wèn)題。
先說(shuō)一下下車(chē)間的生產(chǎn)設(shè)備。小容量注射液生產(chǎn)車(chē)間主要生產(chǎn)設(shè)備有：洗烘灌聯(lián)動(dòng)生產(chǎn)線、全自動(dòng)配液系統(tǒng)和濕熱滅菌柜都是國(guó)內(nèi)品牌設(shè)備，設(shè)備性能和自動(dòng)化程度高。
安瓿清洗滅菌流程：安瓿瓶--純化水超聲波清洗--注射用水沖洗--壓縮空氣吹掃--注射用水沖洗--壓縮空氣吹掃--進(jìn)入300℃滅菌隧道烘箱--B級(jí)灌封機(jī)進(jìn)瓶轉(zhuǎn)盤(pán)。
全自動(dòng)配液流程：P1罐--鈦棒脫炭--P2罐--0.45 um和 0.22um串聯(lián)除菌--P3罐（無(wú)菌貯備）--兩個(gè)0.22um串聯(lián)除菌--灌封緩沖罐--分液器--陶瓷泵--針頭。
全自動(dòng)配液系統(tǒng)具有在線CIP和SIP功能，CIP具有在線檢測(cè)最終清洗水電導(dǎo)率，SIP滅菌完成后具有正壓保護(hù)功能。滅菌吹掃后正壓保護(hù)在24小時(shí)后壓力≥0.08Mpa，從這可以看出設(shè)備滅菌后被污染可能性小。
生產(chǎn)前：公司小容量注射液車(chē)間（非終端滅菌）生產(chǎn)不是很飽和，在上個(gè)星期生產(chǎn)了一批培養(yǎng)基無(wú)菌模似灌封，為何要重新再做一次？原因是操作工在模似灌封后充氮充了氮?dú)?，方案是不能充氮?dú)獾?，所以需重做。在此段時(shí)間沒(méi)有生產(chǎn)過(guò)任何產(chǎn)品，時(shí)間間隔了一個(gè)星期多。
之后詳細(xì)了解了生產(chǎn)情況：
1，在上一次生產(chǎn)完培養(yǎng)基后，有沒(méi)有立即徹底清場(chǎng)？回答是： B級(jí)灌封間當(dāng)天有徹底清場(chǎng)和消毒，全自動(dòng)配液系統(tǒng)當(dāng)天只在線清洗了，到第二天才滅菌。這一點(diǎn)似乎好好像沒(méi)有問(wèn)題。其實(shí)我認(rèn)為，問(wèn)題的所在就在此，全自動(dòng)配液系統(tǒng)在CIP后沒(méi)有馬上滅菌，存在一定風(fēng)險(xiǎn)。
,2，本次模似灌裝生產(chǎn)前準(zhǔn)備：全自動(dòng)配液系統(tǒng)按規(guī)程進(jìn)行了清洗和滅菌，潔凈室經(jīng)過(guò)清潔和消毒，B級(jí)潔凈區(qū)還進(jìn)行了過(guò)氧化氫濕霧法消毒，進(jìn)入B級(jí)所用到的工器具、潔凈服、維修工具等都經(jīng)過(guò)濕熱滅菌。除菌過(guò)濾器都經(jīng)過(guò)完整性測(cè)試后安裝設(shè)備在位滅菌。
,3，在無(wú)菌模似操作過(guò)程：人員嚴(yán)格按照無(wú)菌更衣程序進(jìn)入灌裝間，整個(gè)操作過(guò)程項(xiàng)目按方案執(zhí)行，生產(chǎn)過(guò)程中在線塵埃粒子未見(jiàn)異常報(bào)警，B級(jí)動(dòng)態(tài)塵埃粒子監(jiān)測(cè)達(dá)到動(dòng)態(tài)標(biāo)準(zhǔn)。
,4，本批生產(chǎn)結(jié)束相關(guān)了解：生產(chǎn)結(jié)束后除菌過(guò)濾器完整性測(cè)試符合要求，灌裝設(shè)備表面微生物符合要求，操作人員手指和潔凈服取樣微生物限度符合要求，生產(chǎn)過(guò)程監(jiān)控浮游菌、沉降菌也未出現(xiàn)異常情況。用于灌裝的培養(yǎng)基在除菌過(guò)濾前測(cè)微生物負(fù)荷≤10Cfu/100mL。
個(gè)人分析：在全自動(dòng)配液系統(tǒng)方面，不管做什么產(chǎn)品，必須在生產(chǎn)結(jié)束后盡快清洗后滅菌。我認(rèn)為在配液系統(tǒng)污菌可能性比較大，主要在除菌過(guò)濾器后管道和灌封緩沖罐清洗不徹底，灌封緩沖罐是連著配液系統(tǒng)一起清洗和滅菌的，唯有一點(diǎn)缺陷是灌封緩沖罐不帶清洗噴淋球，罐體中還有兩個(gè)上下液位電極感應(yīng)器，同時(shí)灌封緩沖罐呼吸過(guò)濾器沒(méi)用在位滅菌功能，現(xiàn)在系統(tǒng)清洗滅菌時(shí)必須關(guān)掉呼吸過(guò)濾器閥門(mén)，系統(tǒng)在清洗過(guò)程中對(duì)灌封緩沖罐清洗不徹底，培養(yǎng)基繁殖大量的微生物。
建議灌封緩沖罐離線清洗和滅菌，用軟連接管代替緩沖罐清洗和滅菌。之后他們拆下灌封緩沖罐，發(fā)現(xiàn)呼吸過(guò)濾器閥門(mén)有少量異物，最終查找到原因。從而也說(shuō)明了設(shè)備URS方案起草時(shí)沒(méi)有考慮多方面因素，做FAT人員不專(zhuān)業(yè)。（注：在職時(shí)，未參與該車(chē)間的管理，也沒(méi)有參與URS編寫(xiě)）
總結(jié)：
第一，企業(yè)關(guān)鍵人員和專(zhuān)業(yè)人員在無(wú)菌生產(chǎn)一線經(jīng)驗(yàn)很重要，人員穩(wěn)不穩(wěn)定直接影響到產(chǎn)品質(zhì)量，企業(yè)應(yīng)該重視在B級(jí)無(wú)菌操作生產(chǎn)員工以及生產(chǎn)管理多年的管理人員。一支穩(wěn)定的團(tuán)隊(duì)重要。在人員這方面，我認(rèn)為員工的能力經(jīng)驗(yàn)和責(zé)任心很重要，學(xué)歷是次要的。高學(xué)歷不一定有能力經(jīng)驗(yàn)，有很多企業(yè)在招聘無(wú)菌生產(chǎn)管理人員時(shí)，學(xué)歷要求很高，非得全日制本科以上學(xué)歷，無(wú)菌操作人員非得大專(zhuān)學(xué)歷。還有好多企業(yè)存在著高級(jí)管理人員看不起低學(xué)歷的員工，大致上認(rèn)為低學(xué)歷低能力。這樣大大抹殺了很多中專(zhuān)學(xué)歷有多年無(wú)菌生產(chǎn)經(jīng)驗(yàn)并有責(zé)任心的員工。好好體會(huì)下產(chǎn)品質(zhì)量是生產(chǎn)出來(lái)的，而不是檢驗(yàn)出來(lái)的這句話。
第二，連續(xù)三批無(wú)菌模擬灌裝，第一批是驗(yàn)證前清場(chǎng)和無(wú)菌操作保證水平，第二、三批是驗(yàn)證生產(chǎn)后清場(chǎng)和無(wú)菌操作保證水平，也是說(shuō)驗(yàn)證清潔操作規(guī)程清潔方法。有些企業(yè)第一批、第二批生產(chǎn)清場(chǎng)結(jié)束后潔凈室內(nèi)用了空氣消毒，用VHP或臭氧，以保證三批培養(yǎng)基無(wú)菌模擬灌裝挑戰(zhàn)成功。我認(rèn)為不是很妥，這樣不能驗(yàn)證清潔操作規(guī)程清潔方法可行性。建議在第三批生產(chǎn)結(jié)束后，必須徹底對(duì)設(shè)備、工器具和環(huán)境清潔消毒。
第三，在停產(chǎn)后恢復(fù)生產(chǎn)時(shí)，還要注意下B級(jí)更衣室的手消器應(yīng)清潔徹底，之前我就碰到過(guò)手消器清潔不徹底，以前殘留酒精含有大量細(xì)菌，更換了除菌過(guò)濾消毒液，操作人員手消毒被污染，操作后造成培養(yǎng)基長(zhǎng)菌，是通過(guò)操作人員手套和潔凈服手袖表面微生物取樣檢測(cè)出有大量菌，基本上每個(gè)進(jìn)入的人員都有，在手消器取樣測(cè)微生物證實(shí)有大量的菌。建議在B級(jí)不要用自動(dòng)手消器，用可滅菌的手壓噴壺每批滅菌后使用，消毒液現(xiàn)用現(xiàn)配除菌過(guò)濾。
第四，培養(yǎng)基無(wú)菌模擬灌裝的整個(gè)過(guò)程盡可能模擬設(shè)備和環(huán)境最條件下進(jìn)行，比如：灌裝設(shè)備頻繁出現(xiàn)故障，進(jìn)入人員超限等。特別是維修人員有無(wú)參與。
作者：楓子 來(lái)源：蒲公英 日水培養(yǎng)基qdrishui.cn
I have been doing the production of freeze-dried powder for some time. The simulation canning experiment is carried out many times. The simulated canned bacteria will happen. Today, I choose a case to talk about the prevention of pollution. I hope to have a good communication with the colleagues.
The company has continuously made three batches of culture based aseptic simulation filling, each batch of 10000, the first batch of cultivation and culture less than two days long bacteria, second, third batch to the end of the culture did not detect positive pollution. And half of the first group was long bacteria, and asked me where there was a problem.
First, say, the production equipment between the car. The main production equipment in the production workshop of small capacity injection is: the linkage production line, the full automatic liquid distribution system and the damp heat sterilization cabinet are all domestic brand equipment, and the equipment performance and automation are high.
Ampoule cleaning and sterilization process: ampoule bottle - purify water ultrasonic cleaning - injection water flushing - compressed air blowing - injection water flushing - compressed air blowing - into the 300 centigrade sterilizing tunnel oven --B grade sealing machine into the bottle turntable.
Automatic liquid mixing process: P1 tank -- titanium rod --P2 tank --0.45 um and 0.22um series --P3 tank (sterile reserve) - two 0.22um series degenerate bacteria - sealing buffer tank - liquid separator - ceramic pump - needle.
The automatic liquid distribution system has the functions of on-line CIP and SIP. CIP has the on-line detection of the conductivity of the final cleaning water and has positive pressure protection function after the SIP sterilization is completed. After positive pressure protection after sterilization, the pressure is greater than 0.08Mpa after 24 hours. From this we can see that the possibility of contamination after sterilization is small.
Simulated filling workshop map of diurnal water biological medium
Before production: the production of the company's small capacity injection workshop (non terminal sterilization) is not very saturated. In the last week, a batch of culture - based aseptic mold was produced. Why should we do it again? The reason is that the operators fill nitrogen with nitrogen after the mold is sealed, and the scheme is not filled with nitrogen, so it needs to be redone. No product has been produced during this period, and the interval is more than a week.
After that, the production situation was understood in detail.
1, was there a thorough clearance after the last production of the medium? The answer is: the B level potting room was completely cleared and sterilized on that day. The automatic liquid distribution system was cleaned online on the same day, and it was sterilized on the second day. It seems as if there is no problem. In fact, I think the problem lies here. The automatic dispensing system has not been sterilized immediately after CIP, and there is a certain risk.
2, the preparation of this mold before filling: fully automatic liquid distribution system cleaning and sterilizing according to the regulations, cleaning and disinfection of the clean room, the B clean area also carried out the hydrogen peroxide wet fog method disinfection, into the B level of equipment, clean clothes, maintenance tools, etc. have been sterilized by wet heat. The sterilized filter is sterilized after being tested for completeness.
3, in the aseptic mode like operation process: the personnel strictly follow the aseptic dressing procedure into the filling room, the whole operation process is carried out according to the plan, the dust particles in the production process have no abnormal alarm, and the dynamic dust particle monitoring of the B level reaches the dynamic standard.
4, this batch of production end related understanding: after the end of production, the integrity test of the bacteria removal filter meets the requirements, the surface microbes of the filling equipment meet the requirements, the operator fingers and clean clothing sampling microbial limit meet the requirements, and the production process monitoring zooplankton and sedimentation bacteria have not been abnormal. The culture medium used for filling is measuring the microbial load less than 10Cfu/100mL before sterilization.
Personal analysis: in the automatic dispensing system, no matter what product is made, it must be cleaned and sterilized as soon as possible after the production is finished. I think the possibility of bacteria contamination in the dispensing system is relatively large, mainly in the pipe and the sealing buffer tank after the bacteria removal filter, and the sealing buffer tank is cleaned and sterilized together with the dispensing system. The only defect is that the seal buffer tank does not have a cleaning spray ball, and there are two upper and lower liquid level electrode sensors in the tank. During the system cleaning and sterilization, the breathing filter valve must be turned off when the system is cleaned and sterilized. The system does not clean the sealing tank thoroughly during the cleaning process and propagate a large number of microbes in the culture medium.
It is suggested that the potting buffer tank be cleaned and sterilized off-line, and the soft connecting tube should be used instead of the buffer tank to clean and sterilize. Then they removed the potting buffer tank and found that there were few foreign bodies in the breathing filter valve, and finally found the cause. It also shows that there are not many factors to consider when drafting the URS plan, and FAT staff is not professional. (Note: no involvement in the management of the workshop and no involvement in URS).
Finally, the conclusion is as follows:
First, the experience of the key personnel and professionals in the production of aseptic production is very important. The stable and unstable personnel directly affect the quality of the product. The enterprise should pay attention to the production staff in the B level aseptic operation and the management staff for years of production management. A stable team is important. In terms of personnel, I think the ability, experience and responsibility of employees are very important, and education is secondary. A high degree of education does not necessarily have the ability to experience, many enterprises in the recruitment of aseptic production managers, a high degree of education, a full-time bachelor's degree or more, aseptic operators have a college degree. There are still a lot of enterprises that are despised by senior managers for low academic qualifications. Generally speaking, they have low academic qualifications and low ability. This greatly obliterates many secondary vocational education with many years of aseptic production experience and responsible staff. A good understanding of the quality of the product is produced rather than tested.
Second, the three consecutive batch of aseptic simulation filling, the first batch is to verify the pre clearance and aseptic operation assurance level, the second, third batch is to verify the post production clearance and aseptic operation assurance level, but also to verify the clean operation procedures cleaning method. In some enterprises, the first and second batch of production clearing houses are sterilized in the clean room, using VHP or ozone to ensure the success of the three batch of culture based aseptic simulation filling challenges. I don't think it's proper enough to verify the feasibility of cleaning procedures. It is suggested that after the third batch of production, the equipment, equipment and environment must be thoroughly cleaned and sterilized.
Third, in the recovery of production after the shutdown, we should also pay attention to the B level dressing room hand elimination device should be clean thoroughly, before I met the hand elimination device cleaning not thoroughly, before the residual alcohol contains a large number of bacteria, the replacement of bactericidal filtrate disinfectant, operator hand disinfection was polluted, after operation, the cultivation of long bacteria, through operation, is through operation A large number of bacteria were detected on the surface of the gloves and the surface of the hand sleeve of the clean clothes. It is suggested that the automatic hand disinfectant should not be used in the B class. The sterilizing hand pressure sprinkling can be used after every batch of sterilization, and the disinfectant is now filtered with bacteria.
Fourth, the whole process of the culture based aseptic simulation filling is as much as possible under the conditions of the simulation equipment and the environment, such as the frequent failure of the filling equipment and the exceeding the limit of the personnel. In particular, the maintenance personnel have no participation.