Microbiological Best Laboratory Practices

Introduction

The USP finalized a series of chapters this summer. Past issues of the newsletter have discussed four of these:

• <61> PMF Newsletter vol 12 no3 (March, 2006)
• <62> PMF Newsletter vol 12 no4 (April, 2006)
• <1111> PMF Newsletter vol 12 no6 (June, 2006)
• <1072> PMF Newsletter vol 13 no2 (Feb, 2007)

There is a need to set some base level of microbiological laboratory standards. Therefore, after several rounds of discussion (stretching over a couple years), the AMB has presented a draft chapter entitled “<1117> Good Microbiological Laboratory Practices” (1). The response to this initial proposal was spirited. Comments can in from industry and FDA on this initial proposal, and a draft revision was published in 2004 (2). This revision was well-received (few comments) and made effective (with minor changes) in August of 2006 (3).

Chapter Organization

The chapter is meant to provide minimum guidance for microbiological practices in the pharmaceutical environment. As such, it is organized into topics of importance to the microbiologist:

• Media Preparation and Quality Control
• Maintenance of Microbiological Cultures
• Maintenance of Laboratory Equipment
• Laboratory Layout and Operations
• Training of Personnel
• Documentation
• Maintenance of Laboratory Records
• Interpretation of Assay Results

Media Preparation and Quality Control

The quality of work in a microbiological laboratory depends on the quality of the culture media. It is essential to use the correct media for the purpose at hand, although the correct media is not always obvious. For example, water testing is commonly done with R2A agar, but many facilities use TSA for this purpose. The recommendation is provided that the choice of media should be consistent, appropriate and justified.
The chapter spends some time on media preparation as well. The recommendations include accurate weighing of dehydrated components, the use of high quality (USP Purified) water, completely dissolving the dehydrated media or individual ingredients, and the need to control the heating of the media to avoid damaging heat-labile components of the media. Some recommendations on the labeling and packaging of media are also provided.
An entire section is devoted to the question of media storage and the effects this might have on the media quality. Excesses of heat and cold are to be guarded against, as is the potential for dehydration of poured plates. Some guidance is also provided in quality control for molten media used in pour plates.
The quality control of the media is a critical concern. Interestingly, initially some of the most passionate commentary on the chapter dealt with the “excessive” amount of space provided to media quality checks. Since the initial release in 2003, however, the harmonized Sterility Tests and the harmonized Microbial Limits Tests have both incorporated stringent media quality checks.

Maintenance of Microbiological Cultures

Second only to media, safeguarding the stock cultures is the most important component of a successful microbiology laboratory. These must be handled carefully at all times to avoid contamination.
The care of the cultures starts upon receipt. A careful stock culture curator will confirm the identity of the received cultures, even if they come from as respected a source as a national culture collection. Mistakes can happen. The use of an incorrect strain in a compendial test could bring the results of weeks or months of work into question.
The chapter reinforces the compendial preference for the “seed lot technique” in culture maintenance. Critical to this is the need to go into your containers of stock culture only once, and in restricting the number of passages. Now, it must be stated that there is nothing magic about the number 5. This number gained popularity in the compendia through its use in the Sterility Test, and has been maintained for consistency. The point to the practice is that a careful lab will safeguard the purity and identity of their stock cultures by limiting the potential for “drift” due to excessive transfers.

Maintenance of Laboratory Equipment

This section was included more for the sake of completeness than because of concerns peculiar to the microbiology laboratory. Most lab equipment in the microbiology laboratory is subject to the standard validation practices of IQ, OQ, and PQ. As is common, periodic calibration/maintenance may be required for the particular equipment based on its nature, and performance verification checks should also be performed regularly. The frequency will depend on characteristics and use of the equipment.

Laboratory Layout and Operations

The need for this section stems from the concern that too few facilities understand or plan for the separation of samples from a microbiological perspective. The success of a laboratory can be enhanced by the thoughtful separation of samples likely to have contamination from those that are expected to be sterile.

Training of Personnel

The chapter states plainly what should be common sense in recommending that microbiologists and managers in the pharmaceutical support lab should have academic training in microbiology or allied health sciences. This recommendation is in line with current best practice for biosafety as laid out in the 5th Edition of the Center for Disease Control’s (CDC) manual “Biosafety in Microbiological and Biomedical Laboratories (BMBL).” (4)
In addition to the recommendation that the microbiology staff have studied a relevant subject while in school, the proposed guidance chapter points out a fundamental link between training and the unit’s SOP system. It recommends that the SOP system should be comprehensive and serve as basis of the training program. This proposal also recommends that performance assessments be done periodically and should demonstrate competency in core activities of the lab.

Documentation and Maintenance of Laboratory Records

Like the section on equipment maintenance, this section was included only for the sake of completeness.

Interpretation of Assay Results

This section was initially entitled “OOS Investigations” it was renamed “Microbial Data Deviation Investigations” out of deference to the work underway by a PDA task force. However, during the writing process it became clear that the scope of this section was broader than merely investigations, and so the current title was settled upon almost by the process of elimination. A discussion of the inherent variability of microbiological data was necessary in this chapter. One view of good laboratory practices could be structured around determining practices that minimize variability in the microbiology lab. However, because we are dealing with such low numbers on plates (frequently less than 20 CFU/plate) and the real opportunities for human error in tests that may run over a month to completion, the microbiologist must always be aware of the role that random chance has in the data and be on guard against over-interpreting the results of a study.
This section of the proposed guidance document is intended to be both a discussion of the limitations of compendial test methodologies and a guide to developing methods of investigating test failures. It discusses the difference between a test that has failed, a test that should be invalidated and a test that should be repeated for confirmation.

Summary

The proposed chapter <1117> Good Microbiological Laboratory Practices” was developed in response to repeated requests from industry for guidance in this area. This chapter is meant to provide guidance to workers and to regulators in evaluating the operations of the QC microbiology lab.
This article is an updated version of an article by the author that originally appeared in the PMF Newsletter vol. 11 no.2 (2004).

References

1. USP. 2006. <1117> Microbiological Best Laboratory Practices. USP 29 Suppl 2 pp. 3804-3807.
2. USP. 2003. <1117> Good Microbiological Laboratory Practices. Pharmacopeial Forum 29(3):842- 850.
3. USP. 2004. <1117> Microbiological Best Laboratory Practices. Pharmacopeial Forum 29(3):1713- 1721.
4. CDC. 2007. Biosafety in Microbiological and Biomedical Laboratories (BMBL) 5th Edition http://www.cdc.gov/od/ohs/biosfty/bmbl5/ bmbl5toc.htm released to the web in February, 2007.

SOPs: Periodic Review AND Process Improvement

The purpose of Standard Operating Procedures (SOPs) is to achieve consistency and uniformity in the way processes are conducted and to ensure that these processes are conducted in a manner compliant with industry and government regulations. Once an SOP is in place, it is equally important to strive for process improvement. Whether called for by changes in regulations or discovery of a “better way,” the process of updating SOPs ultimately contributes to the integrity of a pharmaceutical company and its products.

This is where periodic reviews of SOPs come into play and have a highly significant role. Regulations evolve and change and, therefore, processes must change accordingly. Reviews of SOPs (whether annual, biennial, etc.) are an opportunity to review the most current and updated regulations and incorporate changes to processes as required. It is the periodic reviews of SOPs that ensure continued compliance and contribute to a culture of ensuring patient safety, which should, of course, always be the number one goal.

Apart from regulation-based changes in processes, periodic reviews also provide the opportunity to re-evaluate a current process and seek improvement. Often times in a review of an SOP, people focus on documenting the “current” process. Often one hears much of “this is how we do it.” The question should not just be “what do we do now;” rather, the right question is really “can we do this better.” A current process may be compliant, but there may be ways to streamline it and make it more efficient, allowing the individuals and departments involved to function in a more productive manner.

Periodic reviews of SOPs are all too often viewed as cumbersome. However, they really should be viewed in a positive light as they are an opportunity to not only ensure that a process is correct, but to also determine if the process can be improved. It will always be important to keep informed of the most current regulations and change processes accordingly. However, pharmaceutical and other healthcare companies can remain ahead of the game and have a greater positive impact on patients by exceeding what is merely required by regulations and focusing on process improvement overall.

Writing Standard Operating Procedures (SOPs)

SOPs are living documents that detail written instructions describing specific steps to follow in all activities under defined conditions. SOPs are necessary to ensure the continuity of processes to achieve quality performance and quality products/preparations. They form part of clinical governance, and in particular, show that pharmacists are putting in place strategies for risk management and harm minimisation.
There are different ways and formats of writing SOPs but the main aim is to get pharmacy staff to write down what they do and, at the same time, to have an opportunity to review what they are doing. The main benefits of SOPs are seen as ensuring that good practice is achieved at all times, clarifying who does what and giving guidance for locums, part time staff and new employees.
SOPs should allow for the continual improvement of standards of service, and provide evidence of commitment towards protecting patients. Additional benefits are:

• Help to assure quality and consistency of service;
• Help to ensure that good practice is achieved at all times;
• Provide an opportunity to fully utilise the expertise of all team members;
• Enable pharmacists to delegate;
• Help to avoid confusion over who does what (role clarification);
• Provide advice and guidance to locums and part-time staff;
• They are useful tools for training new members of staff;
• Provide a contribution to the audit process.

All pharmacies operate differently, and SOPs need to reflect this. However, there are some general principles that will apply. SOPs should:

• Be pharmacy specific;
• Be dependent on the competence of the staff working in that pharmacy;
• Under normal circumstances, be applicable at all times, i.e. not dependent on the presence of the pharmacist under whose authority the procedure was prepared.

SOPs should help to ensure that, other than in exceptional circumstances, recommended procedures are followed at all times. Their introduction provides an opportunity for pharmacists to define and assess their own practice, to communicate this to staff and help to improve team-work within the pharmacy.

Who should write SOPs?

Ideally, SOPs should be written by "teams" as it is a good idea to involve staff in developing, reading and commenting on SOPs so they are happy to use them. Getting staff who are involved in specialist tasks to do the first draft of procedures can help, especially if the Charge Pharmacist has less experience in that area. Team writing accomplishes several goals in addition to producing an SOP:

• It ensures that comprehensive knowledge acquired from different perspectives is applied to the SOP;
• It creates "buy-in," which increases the likelihood that the SOPs will be implemented under the guidance of the writers;
• Team-work trains trainers. Having participated in in-depth decision making about the SOP, writers know it intimately and are more likely to be effective trainers (coaches);
• It involves people with differing roles, which helps ensure that when new and modified processes are implemented, someone goes back and updates the SOP.

SOPs should not be wordy and complicated; if short sentences or bullet points will do the trick, use them. The step-by-step parts of an SOP can be described as a numbered list or by using flow charts.

What to include

1. Write a title (with a descriptive verb) that defines the purpose of the SOP.
2. Use document reference numbers if necessary to link SOPs and add revision dates on the title or cover page and the first page of text.
3. Identify the specific points of activity for which the SOP has been written.

4. State the purpose of the SOP. Include information about process and regulatory standards. Some SOPs include both desirable and undesirable consequences.

5. Job descriptions e.g. technician, can be used to describe levels of responsibility if there is a high turnover of staff, or there are a number of part-time staff, so locums know who is responsible for what activities. Using names instead of descriptions may mean the SOP will need to be amended each time a member of staff leaves.
6. Write a "scope" statement that tells what related subjects the SOP will not cover if there is any chance someone will be confused and make a mistake. Scope statements are useful for two reasons: to focus your attention as a writer and to clarify things for a reader.
7. If any "tools" are required for following the SOP e.g. extemporaneous compounding, list these where they apply (possibly in a table).
8. Give an overview of the steps in the SOP that describes the process in terms of its major functions. Include anticipated safety, health, environmental and operational issues.
9. Define terms and concepts. If the SOP contains terms and concepts that readers may be unfamiliar with e.g. in aseptic dispensing, define these in their own paragraph so that readers (a) know that there are unusual words or concepts, and (b) can find them easily for use when needed. A long list of terms may fit better in a glossary at the beginning of a document. If you decide that a simple list of terms and definitions is better, include the list within the write up, perhaps right before the list of steps to be performed.
10. Place safety warnings, cautions and notes prominently within the SOP before the actual step to be described. Never place safety items at the end of a step.

11. List and explain the process steps in sequential order in which a SOP user should perform the steps.
For each procedure, think about the following:

Dispensing SOPs

Pharmacists are accountable for the dispensing process, but in developing and working to SOPs, should be able to benchmark current practice and ensure that systems of practice operating within their pharmacy is safe. SOPs should cover all aspects of the dispensing process, including the delivery of the medicine or product to the patient, and must comply with professional requirements applying to the dispensing process. The added-value of the pharmaceutical service i.e. the pharmacist's professional input into the assessment of the safety and appropriateness of a prescription and, in the provision of information and counselling when completed prescriptions are transferred to patients, should be explicit.
The dispensing process should be clearly defined in the SOP and it should specify which activities must be carried out personally by a pharmacist, including the clinical check (see Council Newsletter October 2006), which activities can be delegated to identified competent support staff and how the checks for accuracy are to be carried out. It is good practice for SOPs to incorporate an audit trail so that the pharmacist can determine who is responsible for each aspect of the process i.e. for each item on the prescription, the dispenser and the checking pharmacist should be clearly identified.

Keeping SOPs current

All SOPs should be numbered and should clearly marked with the date of preparation and/or date of review/amendment. They should be kept up to date and relevant at all times and should be regularly reviewed to allow for changes in practice or circumstances, for example, legislative changes or changes of staff. In the absence of any obvious changes, reviews should be undertaken at least once every two years.
When SOPs are first drafted, or when new members of staff are appointed, it is good practice to ask staff to sign to say they have read and understood them. As well as clarifying staff roles, this can also offer an opportunity for staff training and development. Pharmacists should ensure that any changes to SOPs are brought to the attention of relevant staff.

Conclusion

Writing detailed procedures and developing document management systems seem tedious and, for many, are not core competencies. However, procedures are the cornerstone of a strong quality system; they provide infrastructure; and allow for audit processes against procedures.
Creating appropriate procedures eliminates the need for interpretation by employees and ensures that procedures are being followed as intended. If developed appropriately, well-written procedures demonstrate that a pharmacy is conducting business in a controlled quality system environment. It also supports meeting the overall goal of providing the public with safe and effective medical products, which, at the end of the day, is what all pharmacies strive to achieve.
Acknowledgement: The Pharmacy Council of New Zealand kindly acknowledges the permission granted by the Royal Pharmaceutical Society of Great Britain for allowing a link to the document "Developing and implementing standard operating procedures for dispensing". This document can be viewed on http://www.rpsgb.org/pdfs/sops.pdf and PCNZ recommends visiting the website for the most up to date version.

SOPs and KISS theory can enable organisations achieve targets

A LOT of advantages and merits can be achieved by having Standard Operating Procedures (SOPs) with the golden rule of Keeping It Short and Simple (KISS) in every organisation. In organisations where work is of a repetitive nature, SOPs can come in very handy and prove to be more than effective.

 

For example, in the aviation world, the safety and security of a flight depends highly on the crew ability to use SOPs. Theoretically, every time a pilot flies with a different pilot, there should be no difference in the way the flight was accomplished as all follow the same SOPs. In other words, aviation industry can sustain and flourish only because of SOPs.

The basic purpose of having a SOP is to break down all the tasks done in a division from simplification view point ensuring no errors are in place. Care should be taken that every member of the concerned division is well trained on understanding the SOP and adhering to the standards set out there in (this will ensure that SOPs are not merely prepared as a book keeping exercise to be kept in files without anyone reading it). Even periodic quiz should take place through inter departmental get-togethers on topics outlined in SOPs.

In organisations like military, aviation and pharmaceutical, SOPs become indispensable because every task is outlined as to how it will be done and who is going to do it.

Some of the features and big merits derived out of SOPs are as follows:

-Administration and Security of an organisation can be very effective if simplified SOPs are in place for every division and organisation listing out very clearly the numerous tasks performed and by whom.

-Modification of SOPs should be a recurring exercise so that every time a lapse or an error takes place, suitable remedial measures are taken to avoid recurrence of the same in future. Even otherwise, creeping of an error in an organisation system can be found out very easily through the SOPs listing out tasks.

-Confusion and chaos in an organisation will be to the bare minimum if not nil once all the job listings are clearly spelt out in the form of SOPs and the staff have been trained properly to understand as to which task is whose responsibility.

-Chain of tasks depending on one after another can be accomplished successfully leaving no scope for any error creeping in whatsoever.

-While framing out SOPs, care should be taken to ensure that it is written simple and not to show the writer’s skills.

Whenever Total Quality Management (TQM) matter is raised in an organisation, SOPs will always be present in the forefront. It would be a good idea if SOPs are also present or made mandatory in organisations and institutions where work carried out may not be repetitive in nature; care is needed to ensure that job listings done cover every task performed.

We often hear, due to bureaucratic delays, in Government departments work could not be done.  This is mainly because there is no presence of clear cut SOPs.

 

Therefore, Government can consider making SOPs mandatory for every department/division/wing howsoever big or small they may be because SOPs guarantee more and more advantages with simplification of the whole processes.

Which Cleaner Should I Use?

Before: Reverse osmosis membranes must be cleaned to yield required flowthrough rates - without contaminating food products. After: After cleaning with TERGAZYME brand, diaphragm shows required drop in pressure differential - without an increase in the permeation flow.

Which Cleaner Should I Use? Use this chart to determine which cleaner is recommended for your cleaning application.
Membrane	Soil Removed/Foulant Cleaned
	Iron Oxide, Metals, Aluminum, Carbonate Scale, Some Polymer, Hydroxides, Other Acid Solubles	Silt, Clay, Organic Material, Aluminum, Bio-Film, Some Polymer, Oil & Grease, Alkaline Solubles	Silt, Clay, Organic Material, Aluminum, Bio-Film, Some Polymer, Oil & Grease, Alkaline Solubles	Silt, Clay, Organic Material, Aluminum, Bio-Film, Blood, Protein Based Soils, Oil & Grease, Bio-Mass	Silt, Clay, Organic Material, Aluminum, Heavy Bio-Film, some Polymer, Oil & Grease, Alkaline Solubles
Cellulose, Acetate, & Tri-AC RO/UF	Citranox, Citrajet	Alconox*	Liquinox*	Tergazyme*	n/a
Polyamide/aramid R/O / NF	Citranox, Citrajet	Alconox	Liquinox	Tergazyme	Alcojet**
Polysulfone & Derivative U/F M/F	Citranox, Citrajet	Alconox	Liquinox	Tergazyme	Alcojet
PVDF & PTFE UF/MF	Citranox, Citrajet	Alconox	Liquinox	Tergazyme	Alcojet*
Ceramic & Stainless UF/MF	Citranox, Citrajet	Alconox	Liquinox	Tergazyme	Alcojet
All Porous Plastic MF	Citranox, Citrajet	Alconox	Liquinox	Tergazyme	Alcojet
Polyvinyl Alcohol Derivative NF	Citranox, Citrajet	Alconox	Liquinox	Tergazyme	Alcojet*
Polyolefin UF/MF	Citranox, Citrajet	Alconox	Liquinox	Tergazyme	Alcojet
* Adjust pH with Citric Acid to membrane manufacturer tolerances. ** Check with membrane manufacturer for approval. Approved membrane configurations are: Disc & Tube, Spiral Wound, Plate & Frame and Tubular styles. RO = Reverse Osmosis, UF = Ultrafiltration, NF = Nanofiltration, MF = Microfiltration

Pharma Clean In Place (CIP) and Standard Operating Procedures (SOPs)

CIP stands for Clean-In-Place and is a method of cleaning the interior surfaces of closed systems and process equipment without dismantling the equipment. There are different types of CIP including, single pass systems and recirculation system.
The pharmaceutical industry heavily relies on clean in place (CIP) system because operators are not required to enter plant to clean, difficult to access areas can be cleaned, production time between production runs is minimized, and recycling the cleaner can reduce cost. Equipment used in pharmaceutical manufacturing must be cleaned before each use, and the cleaning procedure used must be in accordance with good manufacturing practices (GMPs). Alconox provides support to meet cGMP regulations. In fact, Alconox, Inc technical team has written The Aqueous Cleaning Handbook, where Standard Operating Procedures for CIP systems are discussed (see Chapter 7)

Zeta Potential Determination for Macroscopic Solid Samples

Problem: The majority of the zetameters are based on the electrophoresis measuring principle where the zeta potential for a colloidal system is analyzed by measurement of the electrophoretic mobility of the particles. However, the measurement of the charged macroscopic surfaces is not possible with electrophoresis. Electrokinetic measurements will enable users to analyze the charged macroscopic surfaces where the zeta potential will give information regarding the adsorption and adhesion processes as well as hydrophobic and hydrophilic nature of these surfaces.

SurPASS (Surface Potential Analyzer for solid samples)

Solution: With the SurPASS, the zeta potential becomes accessible by the streaming potential or (alternatively) the streaming current method. A dilute aqueous electrolyte solution (such as 1mM KCl) is circulated through the measuring cell that contains the sample, which may be a porous sample (permeation method) or a pair of planar surfaces separated by a small gap (tangential method). The sample arrangement represents a mechanical resistance and the liquid flow generates a differential pressure between the inlet and outlet of the measuring cell. Electrical charges, which are accumulated at the interface between the solid surface and the surrounding liquid, are sheared off and accumulated on one end of the measuring cell. The generated potential difference is detected by the Ag/AgCl electrodes.

Electrochemical double layer

The zeta potential itself is defined as the electrical potential at the interface between the immobile layer of solid surface charges and the diffuse layer of counter ions and is proportional to the streaming potential coefficient dU/ dp. The complete equation for the zeta potential ζ reads

Besides the solid surface properties, the viscosity η, the dielectric coefficient ε ε0 and the electrical conductivity κ of the liquid phase contribute to the magnitude and sign of the zeta potential. The conductivity and the pH of the electrolyte are measured externally in the electrolyte reservoir. SurPASS also features a built in automatic titrator that ensures that the zeta potential can be measured as a function of varying pH or conductivity of the electrolyte. It is also possible to add small quantities of additives to detect the changes to the sample surface due to adsorption processes.
This electrokinetic analyzer enables the measurement of the zeta potential for a variety of solid samples that can range from powder, fibers and planar samples such as membranes, filters, polymers, biomaterials, silicone nitride wafers etc. These samples can be mounted into the SurPASS by using three types of measuring cells.