SBNet 2010 Proper autoclaving of solid wastes. Bernard Jenni

Transcription

1 SBNet 2010 Proper autoclaving of solid wastes Bernard Jenni

2 Autoclaves: Hazards are often underestimated USERS (the main hazards!) Incorrect transport of wastes Incorrect handling Incorrect processing of materials Incorrect operation Inadequate maintenance Hazards High temperatures High pressure Steam Hot liquids Biohazard 2 SBNet 2010 B.Jenni solid wastes

3 Explosion hazard Explosion in an autoclave caused by cellulose nitrate tubes Nature 199, 102 (July 1963); doi: /199102a0, I. H. SILVER Explosion in an autoclave and subsequent fire. Most likely cause: Incorrect disposal of chemical waste in the biowaste bin (2006, USMW). Do not autoclave bleach (Javel) Do not autoclave volatile chemicals Do not autoclave radioactive materials. 3 SBNet 2010 B.Jenni solid wastes

4 Transport >> << CONFLICT >> << Autoclaving Containers for BL2-biowastes must be closed during the transport - with a visible biohazard sign They must be open during the autoclaving to allow the penetration of steam Biohazard sign should not be visible for final discard 4 SBNet 2010 B.Jenni solid wastes

5 The basic principle of autoclaving Saturated STEAM UNDER PRESSURE is the sterilizing agent Hot steam = very high heat content = high killing power Dry, saturated steam condenses on colder surfaces Condensation releases high amount of heat heat transfer killing effect Condensation decrease of volume lower pressure draws more steam Potential issue: condensed water wets the surfaces (objets) 5 SBNet 2010 B.Jenni solid wastes

6 Why is an autoclave such an effective sterilizer? The world of autoclaves An autoclave is a large pressure cooker; it operates by using steam under pressure as the sterilizing agent. High pressures enable steam to reach high temperatures, thus increasing its heat content and killing power. Most of the heating power of steam comes from its latent heat of vaporization. This is the amount of heat required to convert boiling water to steam. This amount of heat is large compared to that required to make water hot. For example, it takes 80 calories to make 1 liter of water boil, but 540 calories to convert that boiling water to steam. Therefore, steam at 100º C has almost seven times more heat than boiling water. Steam is able to penetrate objects with cooler temperatures because once the steam contacts a cooler surface it immediately condenses to water, producing a concomitant 1,870 fold decrease in steam volume. This creates negative pressure at the point of condensation and draws more steam to the area. Condensations continues so long as the temperature of the condensing surface is less than that of steam. These properties ensure rapid heating of surfaces, good penetration of dense materials, and coagulation of proteins. 6 SBNet 2010 B.Jenni solid wastes

7 Liquid water,...,water vapour, steam, & air Common belief indicates the contrary, but water vapour & steam are invisible! (only condensed water droplets are visible, e.g., clouds)... AIR IS HEAVIER THAN STEAM! MW = molecular weight Gas MW H 2 O 18 N 2 28 O 2 32 CO 2 44 Air density (masse volumique) in kg/m 3 under 1013 mbar RH= relative humidity Temp. 100% RH 1% RH 0 C C C SBNet 2010 B.Jenni solid wastes

8 Autoclaving of solid wastes Air is the enemy since the heat power is the steam Air does not mix readily with steam and sinks to the bottom of the autoclave chamber Entrapped air pockets prevent steam from touching surfaces, impairing killing effect. Air must be removed 8 SBNet 2010 B.Jenni solid wastes

9 Air removal by autoclaves without vacuum Gravity displacement: limited efficiency to remove entrapped air pockets Source: Steam sterilization. Rolf Dittli (Systec Schweiz GmbH) 9 SBNet 2010 B.Jenni solid wastes

10 Air removal by vacuum assisted autoclaves Air removal by pulsed, fractionated pre-vacuum Usually multiple pre-vacuum pulse(s) Vacuum steps below atmospheric pressure - For solid loads! - Perfect air removal by hollow objects, tubings, pipette tips, etc... - Liquids will boil, splash and spoil the autoclave chamber! 10 SBNet 2010 B.Jenni solid wastes

11 Air removal by pressure pulsation steps Air removal by several pressure releases (not vacuum) Pulses steps above atmospheric pressure - Less efficient than fractionated, pre-vacuum steps - A better choice for not-so-solid wastes, agar plates, etc SBNet 2010 B.Jenni solid wastes

12 Sterilization of agar plates Pressure released steps, 124ºC, 90 min Pressure Temp chamber Temperature inside load 12 SBNet 2010 B.Jenni solid wastes QA-run performed by Philippe Wirth

13 Sterilization of agar plates Pressure released steps, 124ºC, 90 min Agar melts and melting is an endothermic reaction Temperature decreases and much time is needed to reach the set temperature again. 13 SBNet 2010 B.Jenni solid wastes QA-run performed by Philippe Wirth

14 The three phases of a cycle Conditioning phase - Air is removed and the load is heated to a set temperature. Sterilizing phase - Time, which is maintained at the set temperature. Exhaust / cooling / post-vacuum drying - Steam is replaced with air (ev. positive pressure to avoid boiling) - Solid wastes are usually not actively cooled - Return to atmospheric pressure - Post-vacuum to minimize odors. 14 SBNet 2010 B.Jenni solid wastes

15 Program parameters, printout and graphs A sterilizer without such data is a blackbox! A sterilizer without somebody (BSO?) able to interpret these data is a blackbox as well! 15 SBNet 2010 B.Jenni solid wastes

16 Parameters for semi-solid biowastes Parameters for the different types of use. Solids Semi-solid biowastes Liquids (glassware, pipette tips) (plasticware containing liquids, agar plates, etc...) (media, cell cultures) Conditioning phase Pre-vacuum pulses Steam release pulses Pre-vacuum pulses (okay for 90% of cases) Gravitation Single vacuum pulse Sterilizing phase Cooling No active cooling No active cooling Active cooling Positive pressure Drying Post-vacuum No vacuum No vacuum 16 SBNet 2010 B.Jenni solid wastes

17 Process indicators Autoclave tapes: only not treated / treated Chemical indicators irreversible reaction with heat, steam & time: CrCl3 (pink) + 6H2O + heat CrCl3 6H2O (green) Biological indicators (spores of Geobacillus stearothermophilus) Vials for liquids 3M Attest & paper strips: only for solids 17 SBNet 2010 B.Jenni solid wastes

18 Proper autoclaving of wastes Questions: What is the GOAL of the process? 1) GLP/GMP sterilisation? 2) Make-safe inactivation for final discard? Is validation with G. stearothermophilus necessary for the inactivation of (BL1-BL2) biowastes (which do not contain thermoresistant microorganisms)? Would it not be sufficient to use a microorganism representative of the load, e.g., a mesophilic bacterium like Bacillus subtilis or B. atrophaeus? 18 SBNet 2010 B.Jenni solid wastes

19 Message to take home Autoclaving is a very complex process, which requires a lot of diverse knowledge. Proper waste autoclaving needs a knowledgeable BSO, able to be an efficient platform between all involved parties (management, autoclaving service, lab users, autoclave suppliers, further external parties). The BSO should know, understand and able to interpret the physical parameters and printouts of the autoclave cycles. 19 SBNet 2010 B.Jenni solid wastes

20 BACKUP SLIDE Dampfdesinfektion versus Dampfsterilisation Din Deutsche Norm Desinfektion - DampfDesinfektionsapparate Teil 3: Prüfung auf Wirksamkeit 1. Anwendungsbereich Diese Norm gilt für die Prüfung auf Wirksamkeit von Dampf-Desinfektionsapparaten nach DIN einschliefllich des Desinfektionsprozesses. Sie gilt auch für die Dampfdesinfektion von Abfällen, an die aus infektionspräventiver Sicht besondere Anforderungen zu stellen sind, d. h. für Abfälle, die aufgrund von 17 Infektionsschutzgesetz behandelt oder gemäss Europäischem Abfallkatalog dem Schlüssel entsprechen. ABFALLVERZEICHNIS : = Infektiöse Abfälle 6.2 Biologische Indikatoren Die Wahl des biologischen Indikators ist abhängig vom Desinfektionsverfahren. Für das 75 C-Verfahren mit einer Haltezeit von 20 min ist der biologische Indikator DIN Bi-AB (Enterococcus faecium nach DIN ) zu verwenden. Für das 105 C-Verfahren mit einer Haltezeit von 5 min ist der biologische Indikator DIN Bi-ABC (Bacillus subtilis nach DIN ) zu verwenden. 20 SBNet 2010 B.Jenni solid wastes

21 BACKUP SLIDE Dampfdesinfektion mit B. subtilis als Bioindikator D-Werte von B. subtilis Quelle: 2. Vorlesung Steriltechnik - Galenische Pharmazie Boiling water for 10 minutes can reduce B. anthracis spore counts by at least 6log10. Inactivation of Bacillus anthracis spores. Ellen A. Spotts Whitney et al. Emerging Infectious Diseases, 9, No. 6, , SBNet 2010 B.Jenni solid wastes

22 BACKUP SLIDE Dampfdesinfektion mit B. subtilis als Bioindikator Bioburden Quelle: 2. Vorlesung Steriltechnik - Galenische Pharmazie 22 SBNet 2010 B.Jenni solid wastes