With all of the work and focus on cleaning validation,1-7
one facet of the process often overlooked or not understood
is the aspect of Hold Time Studies. So, what exactly are
Hold Time Studies? There are two aspects of “hold times”
generally evaluated for validated cleaning processes in
pharmaceutical manufacturing:
- Clean Hold Times:
The time elapsed from the end of the cleaning
process until the beginning of the use of the cleaned
equipment for manufacture of the next product - Dirty Hold Times:
The time elapsed from the end of manufacturing
until the beginning of the cleaning process
Either of these two time periods can have a significant
impact upon the cleaning process. Keep in mind that a
cleaning validation study is a reflection of the process(es)
used for cleaning and maintaining items in a cleaned state.
An investigator may note two different types of situa-
tions during an audit. One is that equipment is stored with a
status of “Clean – Ready for Use” for perpetuity. That is, no
time limit has been placed on when the item is to be re-
cleaned. Questions that are asked include: - Is this equipment really clean now?
- Do Standard Operating Procedures (SOPs) indicate
how long the cleaned equipment can be stored prior
to re-cleaning?
Does adequate data exist to support the storage of the cleaned equipment?
A second observation regards dirty equipment held over a period of days –sometimes longer – prior to cleaning.
This can be problematic for cleaning – especially if you deal with liquids, which dry and crystallize onto the surface, or if you handle powder, which is hygroscopic and soon trans-
forms into a paste. Questions for these situations include:
- What is the impact on the surface of the equipment
being held for that period of time? - How is the dirty equipment held so as to not contaminate other items?
- Is there data to support the cleaning of the dirty equipment after it has been held for long periods of time? (To illustrate: cleaning off a powder or liquid
is handled much differently from cleaning off a paste or a crystallized substance.)
This article is presented as a guide to understanding the approach to conducting validation studies which compile
data in support of typical operational practices.
CLEANED EQUIPMENT HOLD TIME
VALIDATION
This section covers issues related to the “Cleaned Equipment Hold Time” (which will be abbreviated CEHT).
CEHT is sometimes called the expiration or expiry period for the cleaned equipment.
Why CEHT Is Important for the Control of a Cleaning Process
Essentially, the best reason to conduct such a study is that nothing stays clean forever, no matter how well protected it is in storage. Re-contamination of the equipment may be an event-related phenomenon. Heating, Ventilation,
and Air Conditioning (HVAC), personnel, storage areas,etc., may all have an impact upon the clean status of equipment. In dealing with possible ways the equipment could be
re-contaminated, one should consider both endogenous sources (such as the growth of microorganisms already within the equipment) and exogenous sources (such as the
entry of external contaminants into the equipment).
Equipment Storage Concerns
A major issue related to the characteristics of the cleaned equipment is whether the equipment is stored wet or in a manner such that microbial proliferation is a possibility. This is covered in the Food and Drug Administration’s (FDA’s)
Cleaning Validation Guidance 1
. If the equipment is stored
wet for a sufficient length of time, there is reasonable prob- ability that microbes will grow to an unacceptable level.
Another issue related to the characteristics of the equipment involves possible routes of entry for external contaminants. For small pieces of equipment (scoops, for example),this involves exposure of the item itself. For larger pieces of
equipment (such as tanks), this involves any opening in the equipment.
Objective of CEHT
The purpose of the CEHT is to adequately protect the equipment during the designated time period including a conservative safety margin. Another purpose is to provide
ample justification that equipment remains safe to use throughout that time period.
Analyze the Storage Situation
Study the storage situation closely and consider factors such as:
- Characteristics of the cleaned equipment
- The nature of possible re-contaminants
- The storage conditions
Be Proactive While considering the situation and factors unique to
your storage circumstances, verify what can be done to adequately protect the equipment from re-contamination.
Justify the Holding Time of Cleaned Equipment
A professional judgment should be made based on an analysis of the storage situation. If equipment is ade-
quately cleaned and then stored dry, all external ports or openings to the storage room environment are closed or
covered with a plastic film, and the storage room is controlled, it is feasible to justify a CEHT of several weeks or months. It should be noted that this approach has its lim-
its; for example, justifying a CEHT of one year by this method would be unreasonable.
➣ Develop Data
This involves holding the cleaned equipment for the maximum CEHT and then sampling surfaces and analyzing those samples for possible re-conta-
minating residues. The possible re-contaminating species or material must be determined. As a general rule, these are limited to bioburden and dust.
Testing procedures for bioburden are relatively straightforward using accepted microbiological techniques. For dust, consider either a particle counter, Total Organic Carbon (TOC), or perhaps an
enhanced visual inspection, such as a white or black wipe cloth.
For sampling sites, do not automatically sample the same sites that were sampled in the cleaning validation protocol. Consider which sites are most
likely to be re-contaminated during storage. For example, for agitator blades, the bottom of the agitator blade (a worst-case cleaning location) may be the appropriate sampling point as part of the clean-
ing validation protocol; however, for determination of the CEHT, the top of the agitator blade may be the appropriate sampling point for dust, since dust
is more likely to settle on the top of the blades.
DIRTY EQUIPMENT HOLD TIME
VALIDATION
This section covers issues related to the “Dirty Equipment Hold Time” (which will be abbreviated DEHT).
Why DEHT Is Important for the Control of a Cleaning Process
The nature of the “dirt” on the equipment surfaces may change over time. This may include drying of the dirt and/or
microbial proliferation. Such changes may make the dirt more difficult to remove by the standard cleaning process.
The cleaning process should be designed to adequately clean the equipment under the worst cases of normal oper-
ating conditions. A demonstration of effectiveness under the worst-case, usually the longest, DEHT should be con-
ducted. It should be noted that there may be some cases where the hold time will have no effect on the ease or difficulty of cleaning. In such cases, establishing a maximum.
DEHT, and demonstrating effectiveness at that maximum time is recommended.
Establish a Dirty Equipment Hold Time Limit ,The most important considerations are that holding times be achievable and practical for the manufacturing en-
vironment and its capabilities. For the maximum DEHT, ask the operations departments about the longest time the equipment would be dirty before the cleaning process would be started. Be conservative and add some extra time to what the Manufacturing and Production Department specified.
For the minimum DEHT, a similar approach is used.
Operations should be asked for the shortest time practical between the end of manufacturing and the beginning of
cleaning. Documentation
If a DEHT is specified for a cleaning process, and that DEHT is critical for control, then it is important to docu-
ment that time. The term, documenting, means that the time of the end of the manufacturing procedures and the time of
the beginning of the cleaning procedures are to be recorded.
The beginning of cleaning time should be defined based on the cleaning process. It should not be based on a Quality Control (QC) release time. The end of the manufacturing time is defined based on the manufacturing process.
Dirty Hold Time (DHT) is the duration of time your equipment sits in a soiled state before cleaning.
Best practice is to clean equipment as soon as processing is complete, while the soil is the easiest to remove. If the soil sits on the equipment surface it not only dries out, but becomes a host for microbial growth.
Dirty Hold time (DHT) – the time from the end of manufacture until the beginning of the cleaning process (also called things like “soiled hold time”) Clean Hold Time (CHT) – the time from the end of cleaning until subsequent use of the equipment (subsequent use includes product manufacture or a SIP cycle)
Hold‑time studies establish the time limits for holding the materials at different stages of production to ensure that the quality of the product does not produce results outside the acceptance criteria during the hold time. The design of the study should reflect the holding time at each stage.
Clean hold time is generally considered to be the time between the completion of cleaning and the initiation of the subsequent manufacturing operation. The main factors affecting the clean hold time are the storage conditions and the room classification / qualification.
Pharma Qualification
Dirty Equipment Hold Time Study Protocol
QUALIFICATION TEST METHOD
Pr-requisite:
Prior of conducting/ executing the protocol, following pre requisites must be full filled.
Environmental conditions should be maintained within the limit.
Personal involved in the handling, storage and testing of sample should be trained.
Qualification test program:
Following procedure should be followed for determining the hold time of the equipment’s.
Dirty equipment holding, sampling and testing for change in microbiological properties.
Swab sampling method:
Two types of parameters will be used to evaluate the samples.
Chemical parameter (UV absorbance)
Microbiological parameters (Total aerobic microbial count).
Swabbing method shall be used for the collection of chemical/ microbiological samples.
Chemical:
For chemical swab sampling cotton swab bud shall be used for swabbing.
The swab head should be moist with purified water and then swabbed on 5cm x 5cm (25cm2).
In case of circular location cover the maximum possible area since the swabbing can’t be done using the standard size of 5cm x 5cm (25cm2).
The swabbing shall be done using approx. 5 horizontals 5 parallel vertical strokes.
Dip the swab in 5 ml of methanol and sent to the QC for analysis for the analysis.
Microbiological Swab:
Sterile cotton bud shall be used for swabbing site where the chances of microbial contamination. Pre moisten the swab in sterile 0.9% saline solution
The swabbing shall be done using approx. 5 horizontals 5 vertical strokes to take swab by assuming that the entire area is swabbed to get the maximum recovery and immediately place in labelled test tube.
Analyze each swab sample as per defined procedure in cleaning validation protocol.
Procedure for Microbial Analysis
Take 4 sterile Petri dishes and aseptically, Transfer 1 ml of swab sample into each Petri dish.
Add 20 ml sterile soybean Casein Digest Agar to two plates and 20 ml sabouraud dextrose agar to remaining two plates
Incubate SCDA 30-35 for 5 days (bacteria) and SDA 20-25 for 5 days (fungus).
At the end of incubation period, count the No. of colonies formed and report the results obtained.
Limit: TMC should not be more than 50 CFU/25 cm2.
Note:
Use sterile hand gloves and mask during swabbing.
Swabbing site shall not be repeated as previous location.
Microbiological swabbing should be always performed prior to chemical swabbing as per the sampling plan to negate risk of contamination.
EXPERIMENTAL PLAN:
After completion of the batch keep the equipment without cleaning for 3 days initially and collect the samples daily without cleaning as per the time mentioned in this protocol. After the final sample cleaning of the tank shall be performed and samples shall be collected and send to QC for analysis, samples shall be collected as per the sampling plan.
After cleaning samples shall be collected as per the sampling plan.
Storage: During the study the equipment shall be maintained in closed condition.
Sampling intervals and locations:
After completion of the batch tank shall be kept on hold without cleaning for 3 days initially.
After that microbial sampling shall be carried from designated locations daily for 3 days.
The chemical swab samples shall be collected on final day after the cleaning activity.
Chemical swab locations: (Final Day)
S. No.
Location
Justification
Sample from out let
Surface of the Lid
Difficult to clean
Inner surface of the tank
Difficult to clean
Microbial swab locations: (initial to third day)
S. No.
Location
Justification
Inner surface of the tank
Difficult to clean
Surface of the Lid
ACCEPTANCE CRITERIA:
This study is carried out to establish the hold time of equipment’s without cleaning. Following acceptance criteria shall be followed to evaluate the efficacy of hold time before cleaning.
Microbial Determination:
Acceptance limit for total microbial count is based on in house specification for bioburden of equipment and pathogen, it is based on in house specification for purified water.
S. No.
Swab sample
Acceptance limit
1
Total microbial count (TBC & TFC)
NMT 50 cfu/25sq.cm.
Acceptance limit for chemical testing is based on our in house specification for purified water.
Acceptance criteria:
Residual content NMT 78.749 mcg/ml at 282 nm by UV photometry (UV absorbance NMT 2.798)
EVALUATION OF THE RESULTS:
Results shall be documented as an annexure and shall be attached with final report. Evaluation of results shall be carried out to derive a time limit for the clean equipment hold time study.
DOCUMENTATION:
Results and reports shall be compiled which shall contain the following:
Summary report
Validation protocol
Test reports
Summary report shall contain the following:
Approval
Objective
Acceptance criteria
Brief validation methodology
Evaluation of results
Final result summary
Deviation, failure investigation reports and corrective actions (if any)