BioScience Laboratories, Inc., personnel participate in the activities of numerous national and international professional associations that focus on microbiology and infection control in the healthcare and food service industries.  As our website indicates, our interests relate to disinfectant and topical antimicrobial formulations, their importance in reducing the risk of disease transmission, and fair assessments of their antimicrobial efficacy.  Because such assessments require methods of testing that provide reliably reproducible data meaningful in the context of infection control, our personnel have, for many years, been deeply involved in method development through the American Association for Testing and Materials (ASTM), specifically, Subcommittee E 35.15 on Antimicrobials.  Four members of our staff, including myself as Subcommittee Co-Chair, serve on E 35.15.

As of the conclusion of our semiannual meeting last week, our Subcommittee has 96 members and is responsible for 45 approved methods, plus another 13 currently in the process of development.  In the interest of brevity, I will describe only two examples of the latter.

The first of special note is a modification of E 1174, the ASTM version of the FDA method specified for testing of handwash products intended for use in healthcare. The modification involves the procedure for contaminating the hands with Serratia marcescens, the indicator bacterium used to challenge product antimicrobial efficacy, and is particularly important in that the new method will be much more appropriate for testing leave-on (non-water-aided) hand sanitizers.

Another method-in-the-making is one for testing liquid microbicides versus bacterial biofilms, organized assemblages that are considerably more resistant to antibiotics, topical antimicrobials, and disinfectants than are planktonic (free-floating) bacteria.  Only in the last decade, or so, has the important role that biofilms play in disease causation and environmental fouling been understood, and colleagues from the Center for Biofilm Engineering at Montana State University here in Bozeman have been in the forefront of methods development in E 35.15.

Although I have selected for comment only these from among our many methods, I would welcome any questions you may have about testing of antimicrobial formulations and how the testing methods are created collaboratively by volunteers from industry, regulatory agencies, and CROs such as BioScience Laboratories.

– John Mitchell, Director of Quality Assurance and Chief Medical Officer

As required by <USP 1072> clean-room disinfectant validation is required “to demonstrate the efficacy of a disinfectant within a pharmaceutical manufacturing environment”.

What you must do:  Take the time to think through all the parts and pieces that make up your overall cleaning program to ensure the program is effective, practical for every day activities.

Product Selection:  

Alcohols:  Broad-spectrum efficacy against vegetative bacteria.  Typical concentration of 70% used.  Not effective against molds or spores. 

Aldehydes: Powerful and aggressive disinfectants.  However, are highly toxic to personnel and require long contact times for sporicidal claims.  

Sodium hypochlorite (NaOCl) and other chlorine compounds:  Broad-spectrum biocidal activity.  Chlorine solutions are corrosive, unstable over time, and rapidly lose activity.  Typically concentrations for sodium hypochlorite are 500 to 50,000ppm.  A low ppm will be effective against most vegetative bacteria within 10 minutes.  Unfortunately, to kill spores/molds the concentration must be greater.  Good disinfectant – Poor cleaner.  Will not remove soil load.   

Hydrogen peroxide: A potent biocide and environmentally friendly.  Peroxides are deactivated in the presence of soil loads, so pre-cleaning is required to achieve the desired reduction in the microbial population.  Typical concentrations as low as 0.5 percent.  Hydrogen peroxide can be combined with other ingredients to dramatically increase its germicidal potency and cleaning performance.

Phenolics: Broad range of disinfectants that are used on environmental surfaces. Typical concentrations are 2 to 5 percent with contact times of 5 to 10 minutes.  Added detergents are effective in removal of soil loads. 

Quaternary ammonium compounds: Non-irritating and non-corrosive to surfaces.  Typical concentrations of 0.1 to 2 percent and require 10 minutes of contact time to kill microorganisms.  However most are not effective in removing biofilms and leave surfaces with a residue that must be removed after disinfection.

Application Procedure:

A spray procedure is a quick way to effectively treat a surface.  However, with many of the disinfectants a pre-clean may be required to remove any soil load.  A wipe procedure is also very effective in mechanically removing microorganisms and is a great addition to any disinfectant cleaning program.

How can BioScience help?

We can perform the following testing (1) use-dilution tests (screening disinfectants for their efficacy at various concentrations and contact times against a wide range of standard test organisms and environmental isolates); and (2) surface challenge tests (using standard test microorganisms and microorganisms that are typical environmental isolates, applying disinfectants according to your cleaning procedures. 

– Liv Graving, Microbiologist and In-Vitro Study Director 

We have been very fortunate in the past regarding results from FDA, EPA, and Sponsor audits.  We take each one seriously, as a learning experience, and implement appropriate changes to our processes in a timely fashion.  The audits are a constant reminder for BioScience to strive to be the best that we can be.  Although we are not ISO 17025 certified, we have been audited using those guidelines, as well as Good Laboratory Practice Regulations (GLPs) and Good Clinical Practice Regulations (GCPs).  Some outcomes of the audits have included: adding “Controlled” and “Uncontrolled” stamps on copies given to auditors; adding “Obsolete” on documents once a revision has been put into place; and being more diligent in referencing other SOPs in our SOP documents.  We are accommodating in sending, prior to an auditor’s visit, such information as our current organizational chart, corporate resumes for key employees, and the index of our Standard Operating Procedures.  We have received praise on the ease with which we are able to access documentation requested by an auditor.  All in all, each audit has been a positive experience for us, and I believe for the auditor as well.  Come visit us and see for yourself.  Scientific Expertise with Montana Hospitality is a self-appraisal we take very seriously.

Amy L. Juhnke, Manager of Quality Assurance/Document Control

For those wondering about the responsibilities of a principal investigator, here is the result of my research on this topic.

The Principal Investigator receives no direct supervision and has complete authority and responsibility for the clinical trials they perform. Personal initiative is imperative. He or she must have a firm understanding of the principles of planning, organizing and management. The Principal Investigator must promote Good Clinical Practices thought the clinical research process by ensuring strict adherence to the protocol, protection of the patient’s health and welfare, integrity of the data generated, and compliance with all federal and state regulations The person who holds this position must have a current physician’s licensure to practice with five to eight years of experience in clinical research. They must have an above average ability to interface with patients, staff, sponsor, and IRB. The Principal Investigator is responsible for maintaining a cohesive team that consistently completes clinical trials on time and at budget.

Provides investigator qualifications and agreements to sponsors by maintaining current curriculum vitae, demonstrates the proper education, training and experience to conduct the clinical investigations Principal Investigator is responsible for signing Form FDA 1572, the protocol, and all sponsor contract(s), and complete disclose of any conflicts of interest

Ensures protocol compliance by possesses a thorough understanding of the protocol, determines if the inclusion/exclusion criteria are compatible with the intended study population, establishes recruitment goals and gains approval of both the IRB and the sponsor prior to any amendments or protocol deviations

Ensures continuous oversight of the clinical trial by the IRB, by providing the protocol, investigator’s brochure, informed consent, recruitment materials, protocol amendments or deviations, adverse events and any other materials regarding the study to the IRB for review

Ensures adequate facility and number of staff to conduct the study and makes adequate time to supervise and conduct the study.

Manages medical care and ensures that all medical decisions related to the clinical trial are made by a qualified physician. The Principal Investigator evaluates subject compliance and response to therapy, is responsible for adverse events, proper medical attention in the occurrence of an adverse event, and notification of primary care physician

Protects the rights and welfare of subjects ensuring the informed consent is in accordance with CFG 56 and 45 and that the subjects are aware of all aspects of the clinical trial

Oversees the quality of the trial by ensuring all case report forms are legible, complete, and accurately reflect the source documents. Reviews, understands, and approves corrections made to case report forms and ensures the proper quarantine and use of the investigational materials

Communicates effectively with subjects, the research team, the sponsor, and the IRB and is responsible for the clarity of the protocol and purpose of therapies by the research staff. The Principal Investigator will participate in study monitoring conducted by the sponsor and regulatory administrations

The Principal Investigator maintains professional knowledge and technical expertise by reviewing medical research publications and participating in professional organizations

John Dyba, Senior Account Executive

“Finally, it has arrived! The super duper doohickey with the detachable thingamajig. Hmmmm … nice box … kind of hard to get open … argh … ahhh … there … box cutter got’er done … oops … directions on the outside of the box say don’t use box cutter …oh well, it’s only a little scratch right there. Hey … it’s not assembled … dang! Guess I gotta get the instructions …  (pause while looking at instructions) … these instructions are too complicated … I can put it together without those silly things.”

And it is very possible that you are mechanically inclined enough to put this piece of equipment together. But what about that scratch that has been unintentionally placed? Unless you read and understand the instructions, you are unaware of the cautions/warnings and other pertinent information that could help avoid damage to yourself, others or the equipment.

User manual’s, technical manuals, work instructions, work documents, protocols, process instructions, policies … whew and wow! So many ways to provide information, but my personal favorite is the Standard Operating Procedure (SOP).

The standard operating procedure is a wonderful tool for controlling work. It defines those subtle details that make the difference between success and failure of the work you are performing. WIKIPEDIA (the on-line encyclopedia) defines the SOP as follows:

A standard operating procedure is a set of instructions having the force of a directive, covering those features of operations that lend themselves to a definite or standardized procedure without loss of effectiveness. Standard Operating Policies and Procedures can be effective catalysts to drive performance improvement and improving organizational results.

An Engineer reads aloud from the “Master” SOP, “Turn the nut clockwise, one revolution using the encapsulated nut rotating apparatus.” A worker crouched down over a very expensive and shiny piece of equipment makes eye contact with the engineer and repeats “Turn the nut clockwise one revolution using the encapsulated nut rotating apparatus.” The worker using the encapsulated nut rotating apparatus then turns the nut clockwise one revolution. He makes eye contact with the engineer and states, “The nut has been turned clockwise one revolution using the encapsulated nut rotating apparatus, check.” An Inspector standing next to the worker monitoring his every movement then states, “The nut has been turned clockwise one revolution using the encapsulated nut rotating apparatus, check.” The Engineer then states, “The nut has been turned clockwise, one revolution using the encapsulated nut rotating apparatus.” A second Inspector that has been standing next to the engineer and monitoring every movement in the building states, “The nut has been turned clockwise one revolution using the encapsulated nut rotating apparatus.” The Engineer and second Inspector then sign that the step has been performed.

Granted, you probably are not going to go to this length in ensuring that the written word is followed verbatim. But, hopefully, prior to starting that very important task you eventually will perform countless times, you read through the SOP relating to the task. Actually, if the work environment allows it, have an authorized copy of the SOP right at your side and follow it word-for-word. And if for some reason the SOP isn’t working for you, stop and revise it to describe clearly the correct procedure to be used in the future.

Scott D. Ferraro, Quality Control Manager

That’s a good question. Here’s another, “Is my media sterile”? Most people would answer the sterility question with an enthusiastic, “YES, of course it is, given the fact that my autoclave print out has revealed time and temperature were adequate and the biological indicator is negative.” But is your autoclave printout really telling you the truth? How about that biological indicator? Some say, “spores don’t lie.” I have personally witnessed several biological indicators placed in a liquid load, the autoclave performed a flawless sterilization cycle, BIs were incubated at 55-60˚C for 48 hours, and when inspected, were negative, but the load was not sterile. Further, the autoclave printout indicated that time, temperature and pressure were within range. So why are my autoclave and biological indicator lying to me? “I have a bad batch of biological indicators,” is probably the first thing out of your mouth. Well … more than likely it was “dog-gone it … ” and after you apologized to you co-workers for your unprofessional outburst, you immediately dialed your supplier of biological indicators and politely told them their BIs are “C_ _ P!”

So, before I inflict my wrath upon my co-workers and that nice Biological Indicator person on the other end of the phone, I should ask myself some questions:

1.                 Is the air in the autoclave chamber being replaced by steam? “Yep … when I close the door and turn or press a couple buttons, a buzzer eventually goes off, and when I open the door, everything is really hot!”

During a liquid load cycle, the steam supply enters the autoclave chamber usually from an inlet located in the upper rear of the chamber. The steam pushes the air from the top of the chamber down the drain usually located at the bottom front of the chamber. If your autoclave repair person has recently visited and ensured you that the autoclave steam traps and solenoid valves are functioning properly, then you should check to see if the load, itself, is trapping air. If a media flask is placed into a container to catch boil off from the flask, air can be trapped in the container and insulate the lower portion of the flask from heat energy transfer. Also, is the autoclave periodically calling for steam? If the autoclave is in its sterilization cycle and is not making any noise (i.e., clicking noise from activating solenoid valves or an intermittent whooshing sound of steam entering the chamber), the autoclave could be experiencing a “superheated” condition. When the chamber atmosphere is not steam-saturated, the flasks will be experiencing the same conditions as exposure to hot air and will not be sterile. 

2.                 Is this load validated? “Yep … same number of flasks and same amounts of liquid … well, maybe I did add a little extra of this and a pinch of that to the recipe.” If someone has not placed mechanical devices (i.e., thermocouples) into a load like the one you are running and determined where the hot and cold locations are for that load, you will not know where to place the BI, which flask to test for growth promotion and pH range, or how long to run the sterilization cycle. During load validations, “come-up” time and the “exhaust cycle” are considered in determining how long a sterilization cycle should be. When all of these factors are considered, risk of overcooking of media is greatly reduced, and sterility is greatly assured.

3.                 Did I place my Biological Indicator in the right place? “Yep … I followed the validation protocol map, and I know exactly where to place the BI.”

Logically, you placed the BI in the flask at the “cold” location specified by the validation study. But did you use the correct BI and did you locate it in the flask in the proper location? There are several BI manufacturers. For example, SGM Biotech manufactures a biological indicator called MagnaAmp™. Because this BI is for steam sterilization of solutions (contains the spore Geobacillus stearothermophilus) and is designed to sink. If your BI is simply floating on the surface of the liquid you are trying to sterilize, the BI will not be monitoring the cold location of the liquid (typically middle and toward the bottom of the flask), but will be subjected to, and accumulating lethality from chamber temperature.

In thermal sterilization processes, “time at temperature” is critical to achieve required product sterility.

According to the U.S. FDA, process validation is defined as: Establishing by objective evidence that a process consistently produces a result or product meeting its predetermined specifications.

Scott D. Ferraro

Quality Control Manager