A Basic concepts:
Pyrogen refers to a substance that causes fever reactions in mammals in clinical practice, and is a common problem faced by manufacturers of injectable raw materials and preparations. Pollution may come from raw materials, water, reagents, workshop environment, equipment, etc. Medications containing pyrogens entering the human body can cause adverse reactions such as chills, shivering, elevated body temperature, body pain, sweating, nausea, and vomiting. Sometimes the body temperature can rise to 40 ° C, and in severe cases, coma, collapse, and even life-threatening situations may occur.
laws and regulations:
Chinese GMP Aseptic Drug Appendix [1]:
Article 3: The production of sterile drugs must meet the requirements of their quality and intended use, and should minimize contamination by microorganisms, various particles, and pyrogens. The skills, training, and work attitude of production personnel are key factors in achieving the above goals. The production of sterile drugs must strictly follow carefully designed and validated methods and procedures. The sterility or other quality characteristics of products must not rely solely on any form of final processing or finished product inspection (including sterility testing).
FDA CFR 211 Subpart E - Control of Components, Drug Containers, and Bottle Caps [2]:
Section 211.94: Drug containers and bottle caps.
(c) Drug containers and caps must be cleaned and sterilized and processed to remove their pyrogenic properties, as indicated by the drug properties, to ensure they are suitable for their intended use. This de heating process should be validated.
(d) Drug containers and caps should be written and follow standards or specifications, testing methods, and, where specified, cleaning, sterilization, and handling methods to remove pyrogenic properties.
FDA CFR 211 Subpart H - Holding and Distribution [3]:
Section 211.165 testing and release for distribution.
(a) For each batch of drugs, prior to release, an appropriate laboratory should determine whether they satisfactorily meet the final specifications of the drug, including the characteristics and strength of each active ingredient. If specific batches of short-lived radioactive drugs are subjected to sterile and/or pyrogen testing, as long as such testing is completed as soon as possible, these batches can be released before the sterile and/or pyrogen testing is completed.
Section 211.167 Special Testing Requirements.
(a) For each batch of drugs claiming to be sterile and/or pyrogen free, appropriate laboratory testing should be conducted to determine compliance with these requirements. The examination procedure should be in written form and should be followed.
Section 211.170: Retain Samples.
(a) Adequate reserve samples should be retained, which should represent each batch and each active ingredient. Reserve samples are at least twice the quantity required for all tests to determine whether the active ingredient meets its established specifications, except for sterile and pyrogen testing. The storage time is as follows:
(b) Properly determined reserve samples representing each batch or batch of drugs should be stored and preserved under conditions consistent with the product label. Reserve samples should be stored in the same immediately sealed container system used for drug sales, or in systems with essentially identical characteristics. Reserve samples at least twice the quantity required for all necessary tests, except for sterile and pyrogen samples. Except for the drugs described in paragraph (b) (2) of this section, a visual inspection of representative sample batches or retained samples from batches selected according to acceptable statistical procedures shall be conducted at least once a year to detect evidence of spoilage, unless visual inspection would affect the integrity of the retained samples. Any evidence of deterioration of reserve samples should be investigated in accordance with 211.192. The inspection results should be recorded and saved together with other stability data on the drug. No need to retain reserve samples of compressed medical gases.
EU GMP Appendix 1 Manufacturing of Sterile Drugs [4]：
principle
The manufacturing of sterile products is subject to special requirements to minimize the risk of microbial contamination as well as particulate matter and pyrogen contamination. This largely depends on the skills, training, and attitude of the personnel involved. Quality assurance is particularly important, and this type of manufacturing must strictly follow carefully established and validated preparation methods and procedures. Do not place the sole reliance on sterility or other quality aspects on any terminal process or finished product testing.
The process used should include air circulation within the chamber and maintaining positive pressure to prevent non sterile air from entering. Any incoming air should pass through a HEPA filter. If the process also aims to remove pyrogens, a challenge test using endotoxins should be used as part of the validation.
Implementation guidance
Pyrogen pollution pathways：
There are four main pathways of pyrogen pollution: injection water; Bring in from raw materials; Bringing in from containers, utensils, pipelines, and devices; Pollution during the preparation process. Refer to GMP guidelines for sterile preparations 3.3.1 endotoxin control [6].
Remove Pyrogen process:
The Chinese Pharmacopoeia requires that the removal of heat sources must meet 250 ° C for 45 minutes. According to the FH calculation formula: FH=10 (T2-T1)/Z=10 (250-170)/54=1365
Each manufacturer of dry heat sterilizers can make theoretical calculations based on the recommended conditions in the pharmacopoeia, but should confirm the equipment's heat removal performance and set the sterilization cycle time for daily use based on actual validation data. Refer to GMP guidelines for sterile preparations 10.3 Dry heat sterilization [7].
Quality defects of Pyrogen contamination in traditional Chinese medicine injections:
Due to incomplete sterilization, inadequate packaging, or poor quality of medicinal materials, pyrogen is often introduced. Therefore, strictly control the quality of raw materials and pay attention to preventing microbial contamination in the prepared intermediates. If necessary, perform a hot spot check before feeding. The method of destroying the heat source of glassware can be done by washing and cleaning it, and then treating it at 250 ° C for 30 minutes or 300 ° C for more than 5 minutes to destroy the heat source; Soak glass, plastic, and other containers in strong acids and alkalis to remove heat sources; When preparing injections, add 0.1% to 0.5% (g/ml) activated carbon adsorption, boil and stir for more than 15 minutes, but the effect of activated carbon adsorption on active ingredients should be investigated. Reference to the GMP Guidelines for Sterile Preparations Appendix 4-2 Implementation Guidelines for GMP of Traditional Chinese Medicine Injections [8].
Dry heat removal Pyrogen and removal Pyrogen validation of glass containers:
Container dry heat removal pyrogen operation can inactivate organisms and degrade endotoxins. Set specific sterilization/remove pyrogen temperatures and times based on container size, material, quality, and loading structure. In large-scale production, the usual method is for containers to be automatically circulated through conveyor machinery, using integrated cleaning equipment and tunnel ovens to clean and remove pyrogen the containers. The removal pyrogen of the plunger can be achieved by repeatedly rinsing with hot injection water. For containers made of high temperature resistant materials such as stainless steel or glass, dry heat sterilization can be used to remove the pyrogen. Alternatively, the removal pyrogen can be cleaned with hot injection water and then subjected to wet heat sterilization. The validation of dry heat sterilization and removal of pyrogen should include appropriate thermal distribution and penetration studies, as well as the use of worst-case operating procedures, container characteristics (such as large capacity containers), and special loading methods to simulate production state testing under worst-case conditions. If necessary, rinsing with water multiple times can effectively remove the pyrogen from these containers. Refer to GMP guidelines for sterile preparations 5.2 Glass containers [9] and GMP guidelines for sterile preparations 5.4 Tools [10].
Pyrogen pollution:
Pyrogen is a substance that can cause mammals to generate heat. Pyrogens are usually endotoxins, organic compounds (lipopolysaccharides) shed during bacterial cell growth, or dead cell residues. They are chemically and physically stable, and may not necessarily be eradicated under conditions that can kill bacteria. Their molecular weight may vary, typically ranging from 12000 to 320000. The level of pyrogen is usually quantified in units of endotoxin per milliliter (EU). The pharmaceutical industry is very concerned about pyrogen, as high concentrations of pyrogen can cause adverse reactions in the human body, leading to fever, fainting, or even death. Refer to the GMP guidelines for water systems 6.1 Basic knowledge of water chemistry and microbiology [11].
【 Reference 】
1.Appendix of Chinese GMP Aseptic Drugs
2. FDA CFR 211 Subpart E - Control of Components, Drug Containers, and Bottle Caps
3. FDA CFR 211 Subpart H - Holding and Distribution
4. EU GMP Annex 1 Aseptic Drug Production
5.Annex 2 of EU GMP 2012 Manufacturing of Bioactive Substances and Pharmaceutical Products
6. GMP Guidelines for Sterile Preparations 3.3.1 Control of endotoxins
7. Drug GMP Guidelines for Sterile Preparations 10.3 Dry Heat Sterilization
8. GMP Guidelines for Sterile Preparations Appendix 4 2 Implementation Guidelines for GMP of Traditional Chinese Medicine Injections
9. GMP Guidelines for Sterile Preparations 5.2 Glass Containers
10. GMP Guidelines for Sterile Preparations 5.4 Tools and Instruments
11. Drug GMP Guidelines for Water Systems 6.1 Basic Knowledge of Water Chemistry and Microbiology
A In order to minimize the risks of contamination and cross contamination during drug production, it is necessary to develop a cleaning and validation plan for co production equipment that can produce multiple varieties in parallel, in accordance with GMP requirements.
Traditionally, HPLC has been the most commonly used quantitative analysis method for cleaning validation of residual active pharmaceutical ingredients (APIs) in pharmaceutical production equipment. In recent years, the preferred clean validation analysis method for pharmaceutical manufacturers, regulatory agencies, and industries has shifted from HPLC to Total Organic Carbon (TOC) analysis. The reasons for this change include but are not limited to better expressing the clean process of the equipment lifecycle, reducing costs, improving productivity, and thus increasing profits.
For equipment cleaning verification using wiping method, corresponding residue inspection is carried out. Cotton swab wiping method is used to collect inspection samples, and total organic carbon TOC is measured by dissolving and diluting with water for TOC inspection; If the equipment cleaning is validated using the rinsing method, the corresponding residue inspection is carried out, and the rinsing solution is collected as the inspection sample to determine the total organic carbon TOC.
So, how should the TOC method be defined for clean validation in the pharmaceutical and biopharmaceutical industries?
● Simplify the verification method based on the cleaning process rather than the product;
● Quickly evaluate the cleaning ability of the cleaning process;
● Optimize key cleaning parameters (TACT, i.e. temperature, action, concentration, and time) through wiping and rinsing sampling methods;
● Estimate the worst-case scenario and complex compounds, such as cleaning agents;
● Accelerate product replacement speed and better produce according to plan.
Why is TOC used in cleaning validation and routine cleaning monitoring?
● Non-exclusive methods can theoretically detect all residual molecules containing organic carbon, which is easy to verify.
● Universal sample analysis methods: cleaning validation and confirmation, raw materials and factory water
● Able to analyze a wide range of dissolved, "insoluble," and most common compounds in the pharmaceutical industry.
● TOC can detect organic acrobatics and residues that have not been completely cleaned accidentally in the equipment.
● High and excellent cost savings. Simple verification method, easy to train new users.
● Improve equipment turnover time.
The unique advantages of TOC method
For the TOC method, when organic matter is designated as an exclusive substance, regardless of whether its source is products, cleaning agents, chemicals, solvents, by-products, or microbial contamination, it is necessary to determine the presence of any residual substances to effectively accomplish this task. As long as its molecular structure contains carbon, any active pharmaceutical ingredient or residual substance for cleaning can be detected. And the test results must be below the limit of the target compound, which improves the strictness of cleaning verification.
The TOC method has gradually been accepted by the pharmaceutical industry, and the strictness of cleanliness verification in the entire industry has been significantly improved, which is increasingly in line with the overall requirements of the industry. The universality of TOC method saves time and labor costs for pharmaceutical companies. The scientific, reasonable, and feasible cleaning verification process ensures the cleanliness required for drug safety production.
A Water system
6.7 Water treatment facilities and their distribution systems should be designed, constructed, installed, debugged, confirmed, monitored, and maintained to prevent microbial contamination and ensure a reliable source of high-quality water. Measures should be taken to minimize the risks of particulate matter, microbial contamination/proliferation, and endotoxins/pyrogens to the greatest extent possible (such as tilting pipes for complete drainage and avoiding dead corners). When filters are included in the system, special attention should be paid to their monitoring and maintenance. The produced water should comply with the current monographs of the relevant pharmacopoeia.
6.8 The water system should be confirmed and validated to maintain appropriate levels of physical, chemical, and microbiological control, taking into account the impact of seasonal changes.
6.9 The pipeline water flow in the water distribution system should maintain end flow to minimize the risk of microbial adhesion and subsequent biofilm formation. The flow rate should be determined during the confirmation process and monitored on a daily basis.
6.10 Water for injection (WFI) should be produced from water that meets quality standards (defined during the confirmation process) and stored and distributed in a manner that minimizes the risk of microbial growth (such as cycling at temperatures above 70 ° C). WFI should be produced through steam distillation or purification processes equivalent to steam distillation. This may include combining reverse osmosis with other appropriate technologies such as electrodeionization (EDI), ultrafiltration, or nanofiltration.
6.11 If the WFI storage tank is equipped with a hydrophobic sterilization filter, the filter should not become a source of contamination and should undergo integrity testing before installation and after use. Control should be implemented to prevent condensation on the filter (such as through heating).
6.12 To minimize the risk of biofilm formation, the water system should be sterilized, disinfected, or regenerated according to the predetermined schedule and as a corrective measure after exceeding or exceeding the limit. After disinfecting the water system with chemicals, a validated rinsing/flushing procedure should be carried out. Water should be tested after disinfection/regeneration. The chemical test results should be approved before the water system is restored to use, and the microbiological/endotoxin results should be verified within the quality standard range and approved before considering batch certification/release for water production using the system.
6.13 Regular chemical and microbiological monitoring should be conducted on the water system to ensure that the water continues to meet pharmacopoeia requirements. The alert limit should be based on initial confirmed data, and then periodically reassessed based on data obtained from subsequent reconfirmation, daily monitoring, and investigation. Review continuous monitoring data to identify any adverse trends in system performance. The sampling plan should reflect the requirements of CCS and include all outlets and water points, with designated time intervals to ensure regular acquisition of representative water samples for analysis. The sampling plan should be based on confirmed data, consider potential worst-case sampling points, and ensure that at least representative samples of water used in the production process are included daily.
6.14 Deviation from alert limits should be recorded and reviewed, including investigation to determine whether the deviation is a single (independent) event, or whether the results indicate adverse trends or system deterioration. Conduct an investigation into each deviation from the action limit to determine possible root causes and any potential impacts on product quality and production processes due to water usage.
6.15 The WFI system should include a continuous monitoring system, such as total organic carbon (TOC) and conductivity, as these parameters better indicate overall systematicity than discrete sampling.
Steam for direct sterilization
6.16 The raw water of the pure steam generator should be appropriately purified. The design, confirmation, and operation of the pure steam generator should ensure that the quality of the steam produced meets the specified chemical and endotoxin levels.
6.17 Steam used for direct sterilization should have appropriate quality and the amount of additives should not contaminate the product or equipment. For generators that provide pure steam for direct sterilization of materials or product contact surfaces (such as porous hard material high-pressure steam sterilization kettles), the steam condensate should comply with the current WFI monograph of the relevant pharmacopoeia (microbiological testing of steam condensate is not mandatory). There should be a suitable sampling plan to ensure that representative pure steam is obtained regularly for analysis. Other quality aspects of pure steam used for sterilization should be evaluated regularly according to validated parameters. These parameters should include the following (unless otherwise specified): non condensable gases, steam dryness value (dryness), and steam superheat.
