Preservation Efficacy Testing Scott Sutton, Ph.D. scott.sutton@microbiol.org 38 52 Material in this presentation is copyright 2011 Microbiology Network Inc. All rights reserved Overview of Presentation What is the Antimicrobial Effectiveness Test? What it can do What it can t do AET (PET) Topics Packaging Effects Formulation Effects Phoenix Phenomenon Linear Regression Unique Microorganisms FDA s Role in this Use of contract labs for generation of PET data 2 1
Antimicrobial Efficacy Test USP <51>/ISO11930 Designed to demonstrate the ability of a multidose product to withstand microbial challenge. High-level challenge, frequent sampling of the challenge suspension for survivors 3 USP Antimicrobial Efficacy Test Finished product test in US, development test in Europe Required for multi-dose presentations, anhydrous ointments that contain preservative Method basically harmonized, criteria not Different criteria for different categories of products 4 2
Consensus Antimicrobial Effectiveness Test Effort of the European Pharmacopoeia, Japanese Pharmacopeia, and United States Pharmacopeia to come to a harmonized assay. Sutton, SVW and D Porter. 2002. Development of the Antimicrobial Effectiveness Test As USP Chapter <51>. PDA J Pharm Sci Tech. 56(6):300-311 Matthews, BR. 2003. Preservation and Preservative Efficacy Testing: European Perspectives. Eur J Parent Pharm Sci. 8(4):99-107. 5 Proposed Cosmetics AET ISO 11930 Cosmetics Microbiology Evaluation of the antimicrobial protection of a cosmetic product (DRAFT) Critical Features Requires Method Suitability Test Describes standard test Provides two levels of acceptance criteria based on risk analysis. 6 3
CTFA Guidelines 13. Determination of Preservative Adequacy in Cosmetic Formulations 14. Preservation Testing of Eye Area Cosmetics 17. Determination of Preservation Efficacy in Nonwoven Substrate Products Product Type Methodology o 20. M-3 A Method for Preservation Testing of Water-Miscible Personal Care Products o 21. M-4 Method for Preservation Testing of Eye Area Cosmetics o 22. M-5 Methods for Preservation Testing of Nonwoven Substrate Personal Care Products o 23. M-6 A Method for Preservation Testing of Atypical Personal o Care Products o 24. M-7 A Rapid Method for Preservation Testing of Water-Miscible Personal Care Products 7 AET - Validation Comparisons Test Group Peptone Control Group Viability Group Neutralizer Efficacy Neutralizer Toxicity 8 4
Method Suitability Acceptance Criteria ISO 11930 Performed in duplicate must be within 50% USP <1227> 5-6 plates in replicate must exceed 70% between populations USP <51> Pharm Eur 5.1.6 9 Standard Antimicrobial Efficacy Test ISO 11930 & USP <51> 6 10 CFU/mL Each of five challenge microorganisms - 3 bacteria, 2 fungi 10-fold serial dilutions Incubate microbial suspension Plate in growth agar to count survivors Sample at 7 days, 14 days, *21 days, 28 days Determine Log Reduction (log value of inoculum) - (log value of survivors at the time point) 10 5
Challenge Organisms Challenge Organism ISO 11930 USP Pharm Eur Pseudomonas aeruginosa ATCC 9027 ATCC 9027 ATCC 9027; NCIMB 8626; CIP 82.118 Staphylococcus aureus ATCC 6538 ATCC 6538 ATCC 6538; NCTC 10788; MCIMB 9518; CIP 4.83 Escherichia coli ATCC 8739 ATCC 8739 N/A Candida albicans ATCC 10231 ATCC 10231 ATCC 10231; NCPF 3179; IP 48.72 Aspergillus brasiliensis ATCC 16404 ATCC 16404 ATCC 16404; IMI 149007; IP 1431.83 11 CTFA PET Challenge Organisms Water- Miscible Non- Woven Eye Area* Atypical Gram (+) Cocci Staphylococcus aureus (6538) NLT One NLT One NLT One NLT One** Gram (+) Cocci S. epidermidis (12228) NLT One NLT One NLT One NLT One Fermentative Gram(-) Bacilli Klebsiella pneumoniae (10031) NLT One NLT One NLT One NLT One Fermentative Gram(-) Bacilli Enterobacter cloacae (13047)) NLT One NLT One NLT One NLT One Fermentative Gram(-) Bacilli Escherichia coli (8739) NLT One NLT One NLT One NLT One Fermentative Gram(-) Bacilli Enterobacter gergoviae (33028) NLT One NLT One NLT One NLT One Fermentative Gram(-) Bacilli Proteus spp N/A NLT One N/A N/A Non-fermentative Gram(-) Bacilli Pseudomonas aeruginosa (9027) NLT One NLT One NLT One NLT One Non-fermentative Gram(-) Bacilli Burkholderia cepacia (25416) NLT One NLT One NLT One NLT One Non-fermentative Gram(-) Bacilli Pseudomonas fluorescens (13525) NLT One NLT One NLT One NLT One Non-fermentative Gram(-) Bacilli Pseudomonas putida (31483) NLT One NLT One NLT One NLT One Non-fermentative Gram(-) Bacilli Flavobacterium spp N/A NLT One N/A N/A Non-fermentative Gram(-) Bacilli Acinetobacter spp N/A NLT One N/A N/A Yeast Candida albicans (10231) Recommended NLT One NLT One Recommended Yeast Candida parapsilosis (22019) N/A NLT One NLT One N/A Mold Aspergillus niger (16404) NLT One NLT One NLT One NLT One Mold Penicillium species NLT One N/A NLT One NLT One Mold Chaetomium globosum (6205) N/A N/A NLT One N/A Mold Trichoderma reesei (13631) N/A N/A NLT One N/A Mold Cladosporium oxysporum (76499) N/A N/A NLT One N/A Mold Penicillium luteum N/A NLT One N/A N/A Spore-Forming Bacilli Bacillus subtilis (6051) Optional Optional Optional Optional Other organisms. Optional NLT One Optional Optional *No ATCC strain designations listed for this method ** Read as "Not Less Than One" or "Select at least one" 12 6
CTFA Preparation of Inocula All CTFA methods except eyecosmetics which does not allow Malt Extract Agar for growth of molds 13 Criteria for Passage USP/Pharm Eur Log 10 Reduction 6 Hr 24 Hr 7 Day 14 Day 21 Day 28 Day USP: Bacteria -- -- 1.0 3.0 -- NI EP-A: Bacteria 2 3 -- -- -- NR EP-B: Bacteria -- 1 3 -- -- NI USP: Fungi -- -- NI NI -- NI EP-A: Fungi -- -- 2 -- -- NI EP-B: Fungi -- -- -- 1 -- NI 7
USP Categories Category Product Description 1 Injections and other parenterals, otic, sterile nasal products, and opthalmics 2 Topical Products 3 Oral Products 4 Liquid Antacids 15 Proposed ISO 11930 Acceptance Criteria 16 8
Proposed ISO 11930 Risk Analysis 17 CTFA Criteria for Passage Bacteria Fungi Cosmetics 99.9% in 7 days 90% in 7 days Eye-Area Cosmetics 99.9% in 7 days 90% in 7 days Nonwovens Determined by risk assessment Determined by risk assessment Note that the challenges can be individual or pooled inocula, and that the test may, or may not, incorporate a rechallenge. 18 9
Scope of AET It can Provide relative estimates of the biological activity of a preservative system in a particular formulation at a particular time. It cannot Predict the preservative efficacy of the multidose finished product in all patients hands under all conditions. 19 AET General Reviews Machtiger, N., et al. 2001. Determination of the Efficacy of Preservation of Non-Eye Area Water-Miscible Cosmetic and Toiletry Formulations: Collaborative Study. J AOAC Intl. 84(1):101 110. Farrington, J.K, et al. 1994. Ability of Lab Methods to Predict In-Use Efficacy Antimicrobial Preservatives In an Experimental Cosmetic. Appl Envir Microbiol. 60(12):4553-4558. Geis, P.A. 1988. Preservation of Cosmetics and Consumer Products: Rationale and Application. J Ind Microbiol. 29(Suppl 3):305-315. 20 10
Overview of Presentation What is the Antimicrobial Effectiveness Test? What it can do What it can t do AET (PET) Topics Packaging Effects covered in the excellent presentation of Dr. Machtiger Formulation Effects Phoenix Phenomenon Linear Regression Unique Microorganisms FDA s Role in this Use of Contract Labs for generation of data 21 Formulation Effects Cyclodextrins Simpson, W.J. 1992. Neutralization of the Antibacterial Action of Quaternary Ammonium Compounds with Cyclodextrins. Fed Eur Microbiol Soc. pp.197-200. Colorants Sakamoto, T.,et al. 1987. Effects of Some Cosmetic Pigments On the Bactericidal Activities of Preservatives. J Soc Cosmet Chem. 38:83-98. Emulsions Rieger, M.M. 1981. Current Aspects of Cosmetic Science:The Inactivation of Phenolic Preservatives In Emulsions. Cosmet Toilet 96:39-43. Myburgh, J.A. et al. 1980. The Influence of Suspending Agents On Preservative Activity In Aqueous Solid/Liquid Dispersions. Pharm Weekblad Sci Ed 2:1411-1416. Myburgh, J.A. et al. 1980. Inactivation of Preservatives In the Presence of Particulate Solids Pharm Weekblad Sci Ed. 2:137-142. 22 11
Formulation Effects Excellent Review Geis, P. 2007. Fundamental Elements of Cosmetic and OTC Drug Microbiological Quality. Amer Pharm Rev. 10(4):86-95. Also: Geis, P. and T Rook. 2011. Microbiological Quality Of Consumer Products HAPPI May, 2011. 23 Formulation Effects - Physical Anhydrous Bases No need for preservation Low Water Activity May not require preservation, technically challenging to test Buffer Effects Example: Phosphate buffer base vs Borate ph Halotolerant/halophilic growth in alkaline shampoos 24 12
Phoenix Phenomenon The apparent regrowth (rebound) of microorganisms after initial testing revealing an aerobic plate count (APC) of <10 cfu/g, is known as the Phoenix Phenomenon (Orth, D.S. 1999. Putting the Phoenix Phenomenon Into Perspective. Cosmet Toilet 114(Apr):61-66) Requires product to have sufficient nutrients for growth 25 Fungal Growth Apparent growth of fungal spores in a preservative efficacy test Most commonly seen with products containing surfactants Dissociation of spore aggregates? 26 13
Linear Regression Model Linear Regression Model Orth, D.S. 1979. Linear Regression Method for Rapid Determination of Cosmetic Preservative Efficacy. J Soc Cosmet Chem. 30:321-332. Orth, D.S. et al. 1980. Establishing Cosmetic Preservative Efficacy by Use of D-Values. J Soc Cosmet Chem. 31:165-172. Orth, DS and LR Brueggen. 1982. Preservative Efficacy Testing of Cosmetic Products: Rechallenge Testing and Reliability of the Linear Regression Method. Cosmet Toilet. 97(May):61-65. 27 Linear Regression Good Screen Many (most?) preservative systems are not linear except in approximate terms Sutton, SVW. et al. 1991. D-Value Determinations Are an Inappropriate Measure of Disinfecting Activity of Common Contact Lens Disinfecting Solutions. Appl Envir Microbiol. 57(7):2021-2026. All chemical preservative systems are consumed in the reaction 28 14
Linear Regression Good Screen Bou-Chacra, NA et al. 2003. Evaluation of Preservative Systems in a Sunscreen Formula by the Linear Regression Method. J Cosmet Sci. 54:1-7. 29 Unique Microorganisms FDA regulatory focus in recent years Bacillus cereus Gram-positive, facultatively aerobic sporeformer. -hemolytic and may be an emerging pathogen. Burkholderia cepacia Pseudomonad, clearly pathogenic to cystic fibrosis patients who can develop pneumonia. THE CONCERN IS THAT CF PATIENTS ARE EXPOSED TO PRODUCTS THAT MAY LEAD TO PNEUMONIA. 30 15
Overview of Presentation What is the Antimicrobial Effectiveness Test? What it can do What it can t do AET (PET) Topics Packaging Effects Formulation Effects Phoenix Phenomenon Linear Regression Unique Microorganisms FDA s Role in this Use of Contract Labs for generation of data 31 FDA Role Auditable GMP GMP Topic Good Manufacturing Practice (GMP) Guidelines/Inspection Checklist ISO 22716 Buildings and Facilities X X Equipment X X Personnel X X Raw Materials X X Production X X Records X X Labeling X X Complaints X X QC Lab Operation X Subcontractor QA X 32 16
ISO 22716 Topics 1. Scope 2. Terms and definitions 3. Personnel 4. Premises 5. Equipment 6. Raw materials and packaging materials 7. Production 8. Finished products 9. Quality control laboratory 10.Treatment of product that is out of specification 11. Wastes 12. Subcontracting 13. Deviations 14. Complaints and recalls 15. Change control 16. Internal audit 17. Documentation 33 FDA Actions Recalls Warning Letters Halting Transport 34 17
Overview of Presentation What is the Antimicrobial Effectiveness Test? What it can do What it can t do AET (PET) Topics Packaging Effects Formulation Effects Phoenix Phenomenon Linear Regression Unique Microorganisms FDA s Role in this Use of Contract Labs for generation of data 35 Contract Lab Relationship Must meet your quality standards Sutton, S. 2011. Successful Use of a Contract Microbiology Laboratory. J GXP Compliance. 15(1):54-64. You are responsible for data generated on your products Diebel Labs Warning Letter Micro issues: No Method Suitability Testing Poor investigations Inadequate control of microbiological growth media Poor control of water Poor documentation practices http://www.fda.gov/iceci/enforcementactions/warningletters/ucm271728.htm 36 18
Summary What is the Antimicrobial Effectiveness Test? What it can do What it can t do AET (PET) Topics Packaging Effects Formulation Effects Phoenix Phenomenon Linear Regression Unique Microorganisms FDA s Role in this Use of Contract Labs for generation of data 37 Thank you for your attention Scott Sutton, Ph.D. scott.sutton@microbiol.org +1 585-298-0767 http://www.linkedin.com/in/scottvwsutton Twitter - @microbiologynet Material in this presentation is copyright 2011 Microbiology Network Inc. All rights reserved 19