Sterile environment

Dr Gerald McDonnell, from STERIS Corporation, discusses the use of VHP decontamination and sterilisation processes for pharmaceutical applications

A variety of methods and antimicrobial agents are used for decontamination and sterilisation.¹ For example, for periodic critical environment decontamination, conventional wipe down techniques with antimicrobial products are widely used.A variety of products are applied but range in antimicrobial efficacy and material compatibility. These include phenolic, quaternary ammonium compound and oxidising agent-based formulations.

In general, these methods are very time consuming and labour intensive. Further, although they can be validated for smaller areas, they are more difficult to ensure coverage and reproducibility over larger areas. As an alternative, traditional fumigation with formaldehyde has been used for larger areas but is uncontrolled, variable, slow acting and, more importantly, has significant health and safety concerns.²

automated methods

Alternative, automated decontamination methods are becoming widely used due to ease of use, higher levels of sterility assurance, and cost savings. These include systems based on hydrogen peroxide, ozone and chlorine dioxide. The most widely used and accepted of these are vapourised hydrogen peroxide (VHP) systems used for decontaminating a wide variety of isolated areas, including rooms and separative enclosures.³ Vapour technology was developed by STERIS Corporation (previously AMSCO) during the 1980s and was registered with the US EPA as a sterilant in 1985.

Decontamination and sterilisation processes may be classified as atmospheric or vacuum applications. Atmospheric VHP systems provide rapid, low-temperature decontamination methods of enclosed areas that may be contaminated with micro-organisms, including spore-forming bacteria. These systems are widely used for the decontamination of airborne or surface contaminants in isolators, biological safety cabinets, rooms, duct work and filters for pharmaceutical production, manufacturing, laboratory animal handling, research and biosafety laboratory facilities. Vacuum VHP systems provide greater penetration for sealed areas, including freeze-dryer, centrifuges and packaged equipment. More recently, this technology has been used for low temperature, packaged medical device sterilisation.

VHP is produced by the vapourisation (at 120°C) of 30-36% liquid hydrogen peroxide to give a mixture of VHP and water vapour. As a dry process, the concentration of VHP is maintained below a given condensation point, which is dependant on the area temperature. For room decontamination VHP is generally maintained well below the saturation concentration at 1-1.5mg/l at 25°C; for medical device sterilisation, higher concentrations can be used at higher process temperature, generally up to 60°C. When the concentration of VHP increases above the saturation point for a given temperature, hydrogen peroxide will preferentially condense out forming concentrated peroxide on a surface, as peroxide has a lower vapour pressure than water. In this situation, although the condensate may be antimicrobial, the process will become variable and damaging to surfaces. These risks are easily reduced for the hydrogen peroxide vapour process during cycle development, by considering the area temperature and volume. Alternative systems are available that use condensed hydrogen peroxide for area decontamination; these are considered 'wet' processes and present different antimicrobial compatibility and safety issues from VHP; these systems are not considered further in this article.

VHP is a broad spectrum antimicrobial with virucidal, bactericidal, fungicidal and sporicidal activity^4-6 (Figure 1). Although the exact antimicrobial mode of action of hydrogen peroxide is unknown, formation of hydroxyl radicals probably reacts with essential cell components to cause cell death.¹

bacterial resistance

In general, similar to other biocides, bacterial spores (in particular Bacillus stearothermophilus spores) have been shown to be the most resistant to VHP, even in the presence or absence of an organic soil (5% serum).^4-6 Unlike liquid peroxide, VHP is rapidly sporicidal at concentrations as low as 0.1mg/l. A comparison of the sporicidal activity of VHP and liquid hydrogen peroxide is shown in Table 1.

VHP biodecontamination is a 'dry' process and used at much lower concentrations than alternative oxidising agent-based liquids (bleach, hydrogen peroxide/peracetic acid combinations).

For these reasons, VHP is compatible and safe for use on a wide range of materials, including metals (e.g. 300 series stainless steel, aluminium and titanium), plastics (e.g. polypropylene, polyethylene, PVC, and polycarbonate), and other materials (silicones, glass, artwork, electronics etc.). For example, the VHP 1000 Biodecontamination systems has been used for 10 years in a range of pharmaceutical and industrial applications and is regarded as being safe for use for isolator/separative enclosure decontamination.^7,3

Cellulosic-based materials present the greatest challenge for decontamination due to adsorption and breakdown of peroxide. Further challenges include metals like copper and brass, which can cause peroxide degradation and, over prolonged exposure periods, may also show surface discoloration with no other changes in the material's physical properties.

environmentally safe

A further advantage is that VHP has an excellent safety and environmental profile. Unlike liquid peroxide, VHP breaks down rapidly in the environment into oxygen and water vapour, therefore presenting no significant environmental concerns. Direct contact with VHP at sporicidal concentrations (>0.1mg/l) should, however, be avoided. It is universally accepted that a safe limit for extended VHP worker exposure is 1ppm for an 8-hour time weighted average, with a short term danger level of 75ppm for 30mins. Although VHP is an odourless, colourless vapour, concentrations above 1ppm are obviously irritating and simple low and high level detection systems are available to monitor levels in a given area.

Atmospheric application, with the VHP1000 biodecontamination series has been widely used and validated in industrial applications for more than 10 years for the decontamination of sterility testing environments, production filling lines, biosafety cabinets, rooms and other enclosed areas or surfaces. A single mobile (VHP 1000, or VHP 1000ED for extended duty applications) generates, delivers, controls and removes VHP for an enclosed environment up to 8000ft3 (231m³), where multiple units can be used in tandem for larger areas. A typical decontamination application is shown in Figure 2.

Modular systems (VHP M1000) are also available for direct integration into a room. More recently systems have been developed for smaller area decontamination (given areas; VHP100 and M100) and larger area remediation/decontamination. The latter systems have been used to remediate areas greater than 200,000ft³ (>5800m³). A typical VHP atmospheric biodecontamination cycle is shown in Figure 3.

The cycle consists of four phases:

1) dehumidification,

2) conditioning,

3) sterilisation and

4) aeration,

which are controlled and monitored by the system. A given area is generally dehumidified below 40% relative humidity and then VHP introduced to achieve sporicidal conditions rapidly (conditioning).

dry process

During the sterilisation phase of the cycle, the system maintains hydrogen peroxide in a dry vapour form to maximise efficacy in given environment, and continually removes and replenishes vapour concentrations over the programmed cycle. As noted above, the VHP cycle is referred to as a 'dry' process, as the concentration is maintained below the critical condensation point of the vapour.

Finally, during aeration, VHP is no longer introduced and is rapidly removed to a safe level (<1ppm). Cycle times will vary depending on the area size, desired level of decontamination, and area contents. For example, cycle times for laboratory research rooms (>5600ft³, 162m³) for the environmental control of parvovirus have been reported up to three hours⁸ and 2.5 hours for cleanroom decontamination up to 2250ft³/65m³.⁷

Automated decontamination cycles allow for routine validation and recording of cycle parameters.

Vacuum applications have also been developed to increase the penetration of the vapour. For example, the VHP 1000DV is a mobile system for freeze dryer and aseptic filling line sterilisation.^4,9 A more recent development is the VHP MD series, based on similar vacuum technology and allowing for the terminal sterilisation of simple and complex medical devices. The series is available in a variety of customised chamber sizes, with a touch screen control and a variety of accessories.

A typical VHP vacuum sterilisation cycle includes: leak test, conditioning, sterilisation and aeration (Figure 4).

optional leak test

An optional leak test may be performed, which holds the area under vacuum for a preset time and monitors the pressure to detect leaks. The conditioning (or drying) phase uses the vacuum system to dry the chamber/load and condition the load temperature for sterilisation (generally in the range of 25-50°C). The sterilisation cycle involves drawing of a deep vacuum, injection and diffusion of VHP, and then bleeding dry air or nitrogen to the sterilisation set point pressure (0.5-700 Torr; 0.67-933mbar). VHP is prepared by direct vapourisation of a 35% hydrogen peroxide solution and, similar to the atmospheric applications, the hydrogen peroxide concentration is kept below the saturation point to prevent condensation on the device surface and maximise the antimicrobial activity and material compatibility of VHP. For medical device sterilisation, the number of sterilisation pulses for each application will vary depending on device design, load size, and materials of construction/packaging.

Finally, during aeration the vacuum system is used to remove VHP rapidly from the chamber by a series of vacuum/air pulses. No further aeration time is required in comparison to ethylene oxide sterilisation, minimising the overall time and cost for product availability. Although the cycle time may vary depending on the application, the overall total cycle time can generally be three hours or less.

The toxicity issues with VHP are significantly less than with ethylene oxide, which is carcinogenic. For example, hydrogen peroxide is used directly on the skin at 30,000-60,000mg/l (3-6%), at 7,500mg/^l on intestinal mucosa and on eye tissue at 500-1,500mg/^l. Further, unlike ethylene oxide, there are no toxic residues as hydrogen peroxide breaks down into water and oxygen.

VHP processes offer safe, effective and efficient alternatives for area decontamination and device sterilisation. As a dry biocide, VHP demonstrates rapid, broad-spectrum antimicrobial efficacy and material compatibility. In addition, these processes can minimise cycle times without any significant safety or environmental concerns. Atmospheric applications have been widely used for area decontamination and, more recently, vacuum applications have been used for increased penetration and for medical device sterilisation.