Determination of TTR. Acceptance Specification, Recovery ≥85% of the Visual Count for the Maximal Incubation Time
| Siteand Media | System Incubation Temperature and time | Time to Results | Comments |
|---|---|---|---|
| Site 1 TSA R2A SDA TSA + LP80 | EM, water and product: 30°C–35°C for 72 to 120 hours | Product: 36 hoursRinse Water 44 hours EM: 60 hours | Many species recovered >85% much earlier for EM but minimum times will be driven by worst case recovery organism(s) |
| Site 2 TSA + LP80 | EM: 22.5°C–27.5°C for 120 hours | EM: 36 hours | 100% recovery at 72 hours Microorganisms included standard molds and bacteria and in-house molds and bacteria |
| Site 3 TSA + LP80 | EM: 30°C–35°C for 120 hours | EM 44 hours | TTR is defined by ATCC strains, site-isolates, slow-growers, and heat-stressed organisms. Verified with real samples in routine |
| Site 4R2ATSA + LP80TSA | Water: 30°C–35°C for 144 hoursEM: 28°C–32°C for 120 hoursProduct: 30°C–35°C for 120 hours | Water: 116 hours EM: 76 hours Product: 52 hours | Representative purified water samples from site water loop Real air and surface samples from C/D/CNC areas pH stressed ATCC strains on phosphate buffer (without product matrix) |
| Site 5 R2ATSA + LP80HT | EM:25°C–30°C(surface and air monitoring)30°C–35°C(personnel monitoring)for 96 hoursWater:30°C–35°C for 164 hours | EM: 56 hours Water: 116 hours | Microorganisms for EM TTR: environmental samples from surface, air and personnel monitoring Microorganisms for water TTR: Stenotrophomonas maltophilia (in-house strain) Methylobacterium extorquens (in-house strain) Water samples of different quality |
| Site 6 TSA + LP80HT | EM: 25°C–30°C for 120 hours | EM: 72 hours | Site-isolates from air and surface monitoring |
| Site 7TSA + LP80HTR2ATSA | EM30°C–35°C for 168 hours Water 30°C–35°C for 120 hours Product 30°C–35°C for 120 hours | EM: 52 hours Water: 64 and 108 hours Product: 36 hours | Water TTR varies by facility flora.Bioburden used pH 4.0 stress method |
| Site 8TSA + LP80R2A | EM30°C–35°C for 72 hours Water 30°C–35°C for 100 hours | EM: 68 hours Water: 100 hours | Microorganisms for WFI water TTR were each “stressed” per JP pharmacopeia: Pseudomonas protegens (ATCC 17386)Stenotrophomonas maltophilia (in-house strain) Methylobacterium extorquens (ATCC BAA-2500) Endogenous bacteria were used for purified water (PUW) and pretreatment water systems, using historically highest count sites based on annual trend reports. EM used typical 5 USP organisms plus 3 in-house organisms. |