bio-decontamination with h2o2 – vapor presentation …€¦ · bio-decontamination with h2o2 –...

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Bio-decontamination with H2O2 - Vapor

PA-PH/EPP | 10/09/2007 | © Robert Bosch GmbH 2007. All rights reserved, also regarding any disposal, exploitation, reproduction, editing, distribution, as well as in the event of applications for industrial property rights.

Packaging Technology

Bio-Decontamination with H2O2 – Vapor in Pharmaceutical Industry

Dr. Johannes Rauschnabel, Robert Bosch GmbH

SBM Expert’s Congress 13th-14th of September, 2007

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StructureIntroduction

“VHP” for Isolators

Results

More Applications

Summary

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Chemistry / Biocidal Effect

Challenges

Regulatory


Results

More Applications

Summary

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IntroductionHydrogen Peroxide is an old sterilant – well established in food industry, where liquid H2O2 is applied for disinfection of packaging material and plastic bottles.

H2O2 has a broad antimicrobial activity spectrum and shows low toxicity.

Residues of hydrogen peroxide decompose into neutral species, which underlines the environmental friendliness of this agent:

2 H2O2 2 H2O + O2

But: hydrogen peroxide can decompose with fast kinetics – under some circumstances it can explode at high concentrations.

For complex geometries disinfection with liquid peroxide has limits. For this case H2O2 in the vapor state is widely used.

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Chemistry / Biocidal EffectThe hydrogen peroxide molecule has a unique geometry with a high energy angle,which makes splitting of O-O-bond easy:

H2O2 2 HO●2 HO● H-O-H + ½O2

The decomposition can be catalyzed by metallic surfaces (Cu, Pb, Mg…) – whichcan be used to decompose after use.On the other side this makes the need fornoble surfaces in the equipment to be disinfected.

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Chemistry / Biocidal EffectThe biocidal effect of hydrogen peroxidecomes from the oxygen atom/radical, which is released under influence of theenzyme catalase at the microbiotic surface.The oxidative power of oxygen radicalscause irreversible damage to enzymicsystems and DNA.

A minimum of humditiy at the surfaceto be treated helps to promote this meta-bolismism – especially bacterial sporesneed a basic humidity to show any bio-chemical activity.

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Challenges

Regulatory


Results

More Applications

Summary

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ChallengesVaporized Hydrogen Peroxide is usedto disinfect bottles and pharmaceutical equipment interior, such as isolators (mini-cleanrooms on top of machinery).

Pioneer in that technique was AMSCO,later bought by Steris Co., with their VHP® technology back in the 80 ies.

Disadvantage of vaporizing H2O2 are thetwo physical states present in the system,when using diluted H2O2 for evaporation.This results in different concentrations andthereby in different inactivation efficacy.

T [°C]

Conc. H2O2 [%]

25

352 79

Gas state

liquid state

Gas + liquid

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ChallengesSlow evaporation of e.g. 35%H2O2 separates the both components(”destillation”) during transition intovapor. Condensation on the surfacesmakes the next separation (“concentration change”).

Sterilisation means absence of viablemicrobiotics. In complex geometriesthis is a challenge, because homogene-ous inactivation is required. This can bequestioned with vapor/liquid mixtures.

H202 vapor is known as bio-decontaminating, not sterilizing surfaces –though 6 log inactivation can be achieved/proven.

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Challenges

Regulatory


Results

More Applications

Summary

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RegulatoryIn “Guidance for Industry: Sterile DrugProducts Produced by Aseptic Processing — Current Good Manu-facturing Practice” published by FDAin September 2004, within Appendix 1:

Aseptic Processing Isolators the needfor decontaminated surfaces is explained:

“1. Surface Exposure Decontamination procedures should ensure full exposure of all isolator surfaces to the chemical agent. The capability of a decontaminant to penetrate obstructed or covered surfaces is limited….”

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RegulatoryContinuation “Aseptic Guidance”:

“2. EfficacyThe decontamination method should render the inner surfaces of the isolator free of viable microorganisms. Multiple available vaporized agents are suitable for achieving decontamination…. Cycles should be developed with an appropriate margin of extra kill to provide confidence in robustness of the decontamination processes. Normally, a four- to six-log reduction can be justified depending on the application.”

“E. Filling Line Sterilization…Where decontamination methods are used to render certain product contact surfaces free of viable organisms, a minimum of a six-log reduction should be demonstrated using a suitable biological indicator.”

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RegulatoryContinuation “Aseptic Guidance”:“E. Filling Line Sterilization…Where decontamination methods are used to render certain product contact surfaces free of viable organisms, a minimum of a six-log reduction should be demonstrated using a suitable biological indicator.”

In EC GUIDE to Good Manufacturing Practice, Annex 1 a list of sterilization techniques (heat, moist heat, radiation, ethylene oxide…) does not include hydrogen peroxide. The description of sterilization/decontamination of isolators does not specifically discussissues coming from bio-decontamination instead sterilization.

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RegulatoryIn EC GUIDE to Good ManufacturingPractice, Annex 1 a list of sterilization techniques (heat, moist heat, radiation,ethylene oxide…) does not include

hydrogen peroxide. The description ofsterilization/decontamination of isolatorsdoes not specifically discuss issues coming from bio-decontamination instead “sterilization”.

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RegulatoryPIC/S Recommendation “Isolators used for Aseptic Processing and Sterility Testing”; Chapter “An expansion of the detailed points to be considered for the implementation of the principles to isolators subjected to a sporicidal process” collects 14 points for the sporicidal process. Point 9.4.1 mentions H2O2 bio-decontamination:

“…The agent selected for gas generation should be sporicidal. The agent used for gas generation or other means of applicationshould be capable of rapidly killing bacterial

endospores, fungal spores and vegetativemicroorganisms.

… Peracetic acid, hydrogen peroxide andformaldehyde are used. …”

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Results

More Applications

Summary

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“VHP” for IsolatorsVaporized hydrogen peroxide is widely used to decontaminate theinside of pharmaceutical isolators.The good old pioneering days applied a mixture of steam and H2O2 to be on the safe side,but validation was a nightmare.

Nowadays several H2O2 gas gen-erators are on the market –different philosophy, but more orless reliable technology.

Cycle development and validation has become a routine job.

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“VHP” for IsolatorsIsolator design principle

Air Management System (HVAC)

Unidirectional Air Flow Unit

Aseptic Processing Chamber

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“VHP” for IsolatorsExample (isolated vial/bottle fill line in Austria)

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“VHP” for IsolatorsClosed loop system:

Liquid H2O2 is dropped on a heatedplate inside the filter level of an iso-lator. Recirculation and continousinjection of peroxide goes to saturationof atmosphere including strong visiblecondensation

Open loop system:

Liquid hydrogen peroxide is evaporatedoutside of the isolator in a gas generator;which carries the vapor with air into theisolator, where it is recirculated. Some o-l-systems perform “flash evaporation”.

H2O2

H2O2

Heat plateFilter

Isolator

Dry air

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“VHP” for IsolatorsClosed loop system:

Simple air management

Vapor passes filters first

Incomplete evaporation

Limited injection rate

Saturation/condensation soon

Open loop system:

Complete (“flash”) evaporation

Controlled atmosphere

Capability for high gas conc.

Air management more complex

More sophisticated generator

H2O2

H2O2

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“VHP” for IsolatorsPreparation (Phase I)

- Expanding gloves, load monitoring goods- Preparation of isolator atmosphere- Preheating of peroxide duct work

Conditioning (Phase II)- Injection of H2O2 with a high rate to

achieve a high gas concentration

Biodecontamination (Phase III)- Injection with a lower rate to replace

inactivated peroxide and keep level

Aeration (Phase IV)- Removal of H2O2 from injection system- Removal of peroxide from all isolator surfaces including filters.

H2O

2-co

nc.

Process time

I+ II

Idealized process cycle

IVIII

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SafeVAP = Safe Vacuum Assisted Peroxide Evaporation

Bosch SafeVAP (open loop system)Features:

Venturi transport

3-step evaporation

Inj.rates up to 80g / min

Controlled atmosphere

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“VHP” for IsolatorsDescription

Filter

Venturi

Evaporator

Particle filterBlower

Peroxidesource(35%)

HEPA H14

Peroxide duct

Compressed air HVAC

Aseptic Processing ChamberScale

Exhaust air

IsolatorAir to isolator

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“VHP” for IsolatorsDescription

-100

0

100

200

300

400

500

600

700

800

15:05:0

6

15:11:0

4

15:17:0

2

15:23:0

1

15:28:5

9

15:34:5

8

15:40:5

6

15:46:5

4

15:52:5

3Time

H2O

2 con

cent

ratio

n [p

pm]

-5000

0

5000

10000

15000

20000

25000

H2O

con

cent

ratio

n [p

pm]

Isolator vol. 8 m³Cycle time 22 minAeration 1,5hoursPeroxide: 310 gInj. rate I: 25 g/minInj. rate II: 9 g/min

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“VHP” for IsolatorsCycle Development for H2O2 bio-decontamination include

OQ-studies

tests with chemical indicators (CI’s)

Tests with bio indicators (BI’s), which are stainless steel couponsinoculated with 6 log of the most resistant viable (spores of Geobacillus stearothermophilus).

To ease handling (and to put an extrachallenge on the biological testsystem), BI’s wrapped in Tyvek becameindustrial standard.

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Worst location CIsfor H2O2

concentration

Worst location with triplicate BIs

Air flow

study

Air velocity mapping

Critical design area

Temperature mapping

Physical System Evaluation

Chemical System Evaluation

Biological System Characterization and Optimization

Safety Evaluation

Biological Verification

D-value determination

Worst location BIs 3 times

Evaluate aeration

time

TestingComplete

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Results

Recent Studies

Results

More Applications

Summary

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Recent StudiesMany studies were performed atBosch during the last 3 years (PhD thesis). Basis for these studies was a reference isolatorequipped with the SafeVAP systemand placed in the Bosch PharmaLab.

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Recent StudiesMaterial of Construction Study:Goal of this study was to learnthe influence of different surfaces(material, finish) on inactivationefficacy of bioburden.

Humidity StudyInfluence of relative humidityat different H2O2 gas con-centration levels was studiedand compared with dew pointanalysis.

300

500

700

900

1100

1300

1500

1700

1900

2100

00:00 00:02 00:05 00:08 00:11 00:14 00:17

Duration of bio-decontamination phase [hh:mm]

Hyd

roge

n pe

roxi

de c

once

ntra

tion

[ppm

]

0

5000

10000

15000

20000

Wat

er c

once

ntra

tion

[ppm

]

Water concentration (humidity level 4)Water concentration (humidity level 3)Water concentration (humidity level 2)Water concentration (humidity level 1)Hydrogen peroxide concentration

D-Values of tested Materials in Pre-, Main- and Poststudy

00,5

11,5

22,5

33,5

44,5

5

avera

ge s

tanda

rd co

ntrols

(ES-1)

(ES-F

G)

(ES-E

P)

(ES-G

)

(ES-44

35)

(ES-43

01)

(Alum

inium

)

(POM)

(EPDM)

(PVDF)

(FPM)

(HYPA)

(PA-12

GF)

(PVC-T

R)

(CG-T

ISS)

(PVC-S

CH)(P

C)

(PEEK-N

at)

(PETP)

(PVC-U

)

(PEEK-P

VX)

(PE-U

HMW)

(VMQ)

(TRFX)

Material

D-V

alue

in m

in

Prestudy Mainstudy Poststudy

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Results

Recent Studies

Results

More Applications

Summary

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ResultsObjective of material study:

Examine influence of differentconstruction material surfaceson inactivation of spores

Determine variable kill kinetics(D-Value)

Test organism: spores of Geo-bacillus stearothermophilusATCC 12980 with population of > 1*106 spores per BI.

----++

Results

KillingFractional

Survival

----++

---+++

2520151050Exposure time [min]

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ResultsResults on some materials (50 different surfaces tested)

D-Values of tested Materials in Pre-, Main- and Poststudy

00,5

11,5

22,5

33,5

44,5

5

avera

ge s

tanda

rd co

ntrols

(ES-1)

(ES-F

G)

(ES-E

P)

(ES-G

)

(ES-44

35)

(ES-43

01)

(Alum

inium

)

(POM)

(EPDM)

(PVDF)

(FPM)

(HYPA)

(PA-12

GF)

(PVC-T

R)

(CG-T

ISS)

(PVC-S

CH)(P

C)

(PEEK-N

at)

(PETP)

(PVC-U

)

(PEEK-P

VX)

(PE-U

HMW)

(VMQ)

(TRFX)

Material

D-V

alue

in m

in

Prestudy Mainstudy Poststudy * No reliable kill

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ResultsResults on standard control (commercial Bio indicator made from SSL)

D-Values of Standard Controls and average Hydrogen Peroxide Concentration during the Cycles

0,0

0,2

0,4

0,6

0,8

1,0

1,2

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27

Number of Cycle for Material Study

D-V

alue

of

Stan

dard

C

onto

rls in

min

500

600

700

800

900

1000

1100

Ave

rage

C

once

ntra

tion

of H

2O2

in p

pm

D-Value Standard Controls Concentration of Hydrogen Peroxide

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ResultsSummary of the study:

Most materials tested show good inactivation properties.

Plastic/elastomeric materials have generally higher D-values compared with stainless steel and glass.

Standard commercial available BI’s show a certain spread of D-values (0,2 min to 0,9 min – late positives included in the calculation), which represents microbiobolgical accuracy.

A correlation between wetting properties, surface roughness and the entire D-values can be recognized.

Critical materials of construction should either be replaced or need prolonging of the cycle time.

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ResultsThe results were recently published:

PDA journal, Vol. 61, No. 4, July-August 2007

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ResultsProcedure for Humidity study:

Separate evaporation of waterand hydrogen peroxide (35%)

Control of humidity level withNIR probe, dew point sensorand standard humidity sensor

D - value determination of commercial BI’s (SSL)

Investigation on inactivationin hindered surfaces withhigh aspect ratio

35% liq. H2O2

H2O

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ResultsDetails:

300

500

700

900

1100

1300

1500

1700

1900

2100

00:00 00:02 00:05 00:08 00:11 00:14 00:17

Duration of bio-decontamination phase [hh:mm]

Hyd

roge

n pe

roxi

de c

once

ntra

tion

[ppm

]

0

5000

10000

15000

20000

Wat

er c

once

ntra

tion

[ppm

]

Water concentration (humidity level 4)Water concentration (humidity level 3)Water concentration (humidity level 2)Water concentration (humidity level 1)Hydrogen peroxide concentration

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Results400 ppm:

0

5000

10000

15000

20000

25000

0 2 4 6 8 10 12 14 16 18 20

D-value [min]

Wat

er c

once

ntra

tion

[ppm

]

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Results600 ppm

0

5000

10000

15000

20000

25000

0 2 4 6 8 10 12 14 16 18 20

D-value [min]

Wat

er c

once

ntra

tion

[ppm

]

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Results800 ppm:

0

5000

10000

15000

20000

25000

0,0 0,5 1,0 1,5 2,0 2,5 3,0 3,5D-value [min]

Wat

er c

once

ntra

tion

[ppm

]

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ResultsComparison

Wrapped vs.Nude BI’s :

0,0

0,5

1,0

1,5

2,0

2,5

3,0

Humiditylevel 1

Humiditylevel 2

Humiditylevel 3

Humiditylevel 4

Humidity level

D-v

alue

[min

]

BIs wrapped in Tyvek unpacked BIs

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ResultsCorrelation of injected H2O2-amount during Cond.phase with D

values (mean average)

0,00

2,00

4,00

6,00

8,00

10,00

12,00

14,00

0 100 200 300 400 500 600 700

Injected H2O2 amount [g]

D-W

ert [

min

]

Mittelwert 800ppm 1. FLMittelwert 800ppm 2. FLMittelwert 800ppm 3. FL Mittelwert 800ppm 4.FLMittelwert 600ppm 1. FLMittelwert 600ppm 2. FLMittelwert 600ppm 3. FL Mittelwert 600ppm 4. FLMittelwert 400ppm 1. FLMittelwert 400ppm 2. FLMittelwert 400ppm 3. FLMittelwert 400ppm 4. FL

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ResultsSummary:

The higher the cgas phase concentration, the more independent is the inactivation effect from the humidity level.

At lower concentrations the same kill as at higher concentration can be achieved with higher humidity (condensation)

The faster the concentration rises during conditioning phase (flash evaporation), the less peroxide is needed to achieve a good kill.

(Nano/micro) condensation occurs at all humidity levels. Visibleconcentration is not needed to get short inactivation times.

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Results

More Applications

Summary

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More ApplicationsVaporized H2O2 is applied with all kindsof isolators: fill/finish isolators, sterilitytesting isolators, hospital isolators…

A new trend is fumigation of cleanroomsand animal testing facilities with hydrogenperoxide vapor.

H2O2 in vapor state could potentially replace alcoholic spray&pray procedures in materialtransfer chambers and material locks for thermolabile products.

Bio-decontamination of tubs with presterilized,nested syringes can also be performed with H2O2.

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Results

More Applications

Summary

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SummaryHydrogen peroxide vapor is widely used in pharmaceutical industry. One major application is disinfection of isolator interior.

Closed and open loop systems are established in the market, which are technically robust and validatable.

Inactivation effect on bioburden differs depending on the material of construction (material and finish).

Gas concentration and humidity effect dominate the kill efficacy: high gas concentrations through flash evaporation help to achieve efficient processes.

New applications of H2O2 vapor include cleanroom fumigation and decontamination of thermolabile materials in transfer chambers,

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Thank you for your attention !

[email protected]

bio-decontamination with h2o2 – vapor presentation …€¦ · bio-decontamination with h2o2 –...

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