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Why staining is crucial in flow decay studies.
Filters of small areas are useful as models for larger operations such as batch-scale processing. Flat disc 47-mm microporous filters are often used for this purpose, especially in flow decline (throughput) studies aimed at sizing the effective filter area (EFA) required for large-scale production.
SARTORIUS STEDIM BIOTECH
In the flow decline method of filter sizing, also known as the flow decay or total throughput method, the quantity of effluent produced using a given small filter area is extrapolated to determine the filter area needed to process the drug volume of an entire production batch. It has customarily been considered convenient and economically useful, both in terms of effort and material costs, to conduct such sizing studies with 47-mm flat discs, because their small size minimizes the quantity of fluid involved and the operational time required for the assay.
If desired, however, discs of an even lower EFA may be used. The smaller the assay filter, the less product is consumed in the testing. This can be an important consideration if the fluid being tested is an expensive drug. Because pharmaceutical filtration is a technico-economic enterprise, the expenses involved are reflected in the drug's cost of goods.
Figure 1. Variation in unspecific adsorption (mg per 10-inch cartridge) when different filter designs are used with the same filter polymer
Regrettably, extrapolating from data obtained by using smaller filter areas or over shorter test intervals tends to give less dependable results. The smaller the filter area and liquid volume used, the less accurate the extrapolation. Consequently, the use of 47-mm flat disc filters in flow decay studies sometimes produces results that are so inexact that they must be used with an extremely high safety margin. Such allowances usually are set at 15–20%, but can be as high as 150%. Such over-sizing will result in high value losses because of unnecessary large hold-up volumes and unspecific adsorption. In such circumstances, the savings achieved from the flow decay studies are lost because of batch-to-batch running costs. For example, a 10-inch filter element could adsorb an average of 200 mg of drug (Figure 1), which commonly can be doubled to 400 mg for a 20-inch filter, which could be required if the filter were improperly scaled. The per-batch value of such waste can be calculated from the dosage and value of the drug product being filtered.1,2 The importance of proper filter sizing also has received attention in the revised PDA Technical Report #26, which describes filter choice and trials in detail.3
When dealing with a "clean" liquid (i.e., a liquid devoid of suspended particles), the flow rate and the resulting throughput, per unit of time, at the applied differential pressure is straightforward to measure, based on the selected EFA. (Temperature must be kept constant, particularly because the liquid's viscosity is its reciprocal, and any increase in either factor will lead directly to a proportional rise in throughput.)
If the liquid contains suspended particulates, however, the filter's porosity will be decreased by particle retention, so additional EFA may be required to compensate for the filter area that is blocked or clogged by the particulates.4 In such cases, it will be necessary to conduct experiments to assess the EFA needed for batch processing.
Alternatively, it may be possible to restore lost throughput by using higher inlet pressures rather than increasing EFA. If filtration is already in progress, that would provide a more manageable alternative. The effectiveness of such a measure would depend, however, on the total suspended solids (TSS) in the solution. Higher differential pressures could increase the pressure drop by compacting any filter cake that may have formed. In contrast, a larger EFA would be less likely to produce such compaction under the same conditions, as there would be little or no filter cake build-up. High differential pressure also commonly leads to high flow rates, which can cause elevated fouling or adsorptive effects. Those effects, in turn, can lead to losses in product yield or the need for larger EFA.
Figure 2. Coomassie-blue stained 47-mm filter discs
If the preparation presented for filtration is relatively free from suspended particles, flat discs of even smaller diameters may suffice for filtration sizing tests because the EFA available for liquid permeation would not be diminished by particle deposits. An example of such an application would be filter sizing for deionized waters. These liquids contain so little suspended matter that the flow decline data can be secured fairly quickly from tests using a small filter. The fewer the solids in the suspension, the less demanding the mathematical extrapolation.
Nevertheless, a sizeable inaccuracy inheres to the extrapolation of results from a 47-mm disc's EFA of 1.49 in2 to the EFA expected from a 6 ft2 (or 3,864 in2) cartridge. At best, the results indicate only hypothetical, non-committal values; hence the large margins allowed for error. An assessment method that requires an EFA overdesign of as much as 1 or 1.5 times the extrapolated value does not merit endorsement.
It may well be that the extreme safety margins reported are exaggerations of the arithmetical uncertainties, reflecting the experimenter's strong fear of having to interrupt the filtration mid-stream to install new filters to allow processing of the batch to be completed. Indeed, the aseptic replacement of a filter is a risk-prone operation, so its avoidance is strongly recommended.
The flow decline, or flow decay method, is used to determine the effective filtration area required to process an entire batch of any volume in an acceptable time frame under a given delta (Δ) pressure. It is managed by a simple arithmetical proportioning. A small filter with a known effective area is used to measure the volume of a drug preparation that can be processed at a selected ?P (differential pressure), given the rate of flow and the throughput it produces. The ratio of the filtration area to the sample volume is extrapolated to the filter area required to process an entire batch. The simplicity of the test ensures its easy mastery; its performance demands only a modest technical background.
In a flow decline experiment, a small aliquot of the suspension is filtered through a small filter. The retained particulate load accumulates on the filter pores, successively diminishing filter porosity and the flow rate. At a certain point, the flow is judged to have ceased for all practical purposes; the effort and time that would be spent obtaining additional filtrate beyond that point is not economically feasible.
In these experiments, one measures the flow rate (volume over time), the throughput, and the total filtrate volume obtained over the duration of the filtration.5 The ambient temperature is also noted. The results identify the ratio between the EFA and the volume of filtrate that is produced before the filtration is terminated. The numerical value has a significance beyond that inherent in the specific filter. Rather, it is taken to quantify the capacity of filters of that type to retain the maximum amount of particulate matter present in that unique fluid preparation under the given filtration conditions. Its importance is in the nature of a validation exercise.
An arithmetical extrapolation of the experimentally obtained ratio can then be used to determine how large a filter area will be required to process a batch of product.6 The ratio that is established is:
in which X (m2 ) is the filter area required to process the batch.
The required filter area, thus calculated, can then be translated into the number and lengths of cartridges or other filtration devices one wishes to use for the batch operation, by using additional equations. The area of a flat disc filter is:
A =π ÷4 × d2
The 47-mm disc has an area of 17.4 cm2 (1.49 in2 ). A 10-in cartridge composed of the same filter polymer will be assumed to contain an area of 6 ft2 . (The areas of other filter cartridges may be calculated in the same way.) Therefore, the throughput volume of the cartridge relative to that of a 47-mm filter of the same polymeric composition is:
Thus, if the 47-mm disc yields a total throughput of 2,500 mL, then 2.5 L x 580 = 1,450 L (~383 gallons) will flow through the cartridge before it shuts down. The rate of flow measured on the 47-mm disc then indicates the number of 10-inch cartridges with an EFA of 6 ft2 that would be required to complete the batch filtration in a timely manner at the constant differential pressure, temperature being kept constant.
From the foregoing, it should be self-evident that the accuracy of the arithmetical extrapolation is highly dependent on the accuracy of the EFA attributed to the 47-mm disc. Nevertheless, it is in this very matter that many incorrect assumptions are made.
In the classical application of the flow decline method, a 47-mm membrane is removed from its package and inserted into a stainless steel holder, where it is held in position by the compressive action of an O-ring. The area of a 47-mm filter disc is 17.4 cm2 . In the holder, however, the O-ring pre-empts a certain quantity of the disc's peripheral space by clamping down on the filter disc to prevent edge-leakage. This reduces the filtration area available for filtration. Thus, the EFA of the inserted 47-mm disc is reduced to less than 17.4 cm2 .
The actual EFA of such an assembly was measured by filtering a staining solution of acridine yellow or Coomassie blue (Figure 2). The stained area identified the sealed area in the confines of the O-ring. As can be seen, the staining solution did not extend beyond or under the O-ring. The stained area measured 41 mm in diameter, which has a total area of 13.2 cm2 . Thus, of the 7.4 cm2 area of the 47-mm disc filter, 4.2 cm2 were rendered unavailable for filtration by the O-ring's preemptive sealing action.
A pre-assembled, disposable unit containing a "47-mm" diameter flat disc filter is available. The pre-assembly offers the attractive advantage of disposability. It is likely that users assume that its EFA is that of a 47-mm disc. However, the membrane used for these devices is actually 50 mm in diameter, a common filter size. The staining technique using acridine yellow reveals its effective diameter to be 48 mm (Figure 3), which means it has an EFA of 18.4 cm2 . Its throughput, however, is being ascribed to that of a smaller EFA—the assumed EFA of the 47-mm disc ordinarily used, namely, 17.4 cm2 . The discrepancy in EFA is even larger if the stained 13.2 cm2 area of an actual 47-mm diameter filter disc is used in the comparison (Figure 4).
Figure 3. Disposable test filter with 48-mm diameter or 18.4 cm2 filtration area
The flow emanating from the 18.4 cm2 EFA of the 48-mm diameter disc filter may mistakenly be ascribed to the actual 13.2 cm2 EFA of an O-ring sealed 47-mm diameter filter. If so, the extrapolation exercise will lead to a numerical multiplier that will indicate that a smaller EFA is needed for processing than would result if the multiplier were based on the smaller (stained) value of 13.2 cm2 . The result will be a low EFA that is insufficient for batch processing needs. This may cause a mid-process filter change-out, which the use of the flat disc filter sizing aims to avoid.
Figure 4. Typical 47-mm test filter used in a stainless steel holder
Even though the use of 47-mm flat disc filters is widespread in flow decline work, the operational details of these studies may differ among users, because the method is not standardized. It is not known what EFA values the many users of 47-mm disc filters actually use in their sizing protocols, because these numbers are seldom reported. Measuring filter EFAs by acridine yellow or other staining does not seem to be a widely discussed or published procedure. Clearly, its application to the use of 47-mm discs in filter sizing studies is not universal.
It is recommended that the staining technique be used to determine the exact diameter of the disc being used, so that its actual EFA can be calculated from that diameter. This practice would minimize the risk of underestimating the EFA needed for processing a batch operation, and reduce the possibility of needing mid-process filter replacements.
Even if staining techniques are used to improve the accuracy of filter sizing studies, the extrapolations from 47-mm flat disc filters to 10-inch cartridges are beset by the uncertainties derived from suspended matter. At best, flow decline assessments based on 47-mm flat disc filters constitute indicator trials.
Therefore, it is best to follow up these indicator trials with verification trials using larger-area pleated filter devices, (commonly 1.5 ft2 ).7,8 Indeed, when costly drug preparations are involved and properly defined filtration area scaling is needed, the use of full-scale filters in assurance trials is recommended. Full-scale trials ensure that the filtration system will be large enough to filter the required batch volume without being oversized, thus minimizing product yield losses.
The function of the 47-mm flat disc filter is to serve as the model for the filter to be used in batch processing. Unfortunately, filterability trials that use 47-mm flat discs can only roughly indicate which filter combination might be optimal. To perform reliably in its pilot role, the model filter should be as similar as possible in all its structural details to the production filter. Moreover, it should be tested under the conditions and in a manner as similar as possible to those of the production operation.
Batch filtration will most likely involve pleated filter cartridges. The details of pleated cartridge construction, however, are substantially different from those of flat filters. The measurement of the effective filtration area of flat disc filters is straightforward, whereas that of pleated filters is appreciably more complex. As a result, flow and retention data will not extrapolate well from flat stock to pleated filter cartridges, and EFA forecasts based on flat stock may overestimate the EFA available from pleated filter constructions. This would lead to a need for mid-process change-outs.
The differences in the EFAs of flat disc and pleated filters can result from the pleat-pack constructions and density of pleated filters. These discrepancies may derive from the flow-attenuating influences of the cartridge's support and drainage layers. In pleated filters, the entire filter area may not be available for the filtration function; a portion may serve to satisfy structural or mechanical requirements. How much of the remainder is available to the EFA function depends on how the pleating operation is conducted.
The details of the pleating operation are beyond the scope of this article.9 We can note, however, that the very nature of the pleat numbers, heights, and degrees of tightness affect the flow and retention properties of the cartridges composed of them.
The flow-attenuating influences of the cartridge's support and drainage layers, and the retention-modifying effect of pleat construction features cannot dependably be assessed from pilot studies using flat stock. Therefore, if pleated filter cartridges will be used in batch processing, indicator trials using 47-mm flat disc filters must be followed up by verification trials using miniaturized pleated filter devices.7,8 Using pleated filters in these verification trials will mean that the test results will be much more relevant. Pleated filter cartridges are available on the market in sizes as small as two inches in length.
When even more reliable EFA forecasts for batch processing are needed, assurance trials should be conducted using larger-scale cartridges. Conducting tests using filters with a larger EFA, even to the extent of using full 10-inch pleated cartridges, would proportionately increase the reliability of the extrapolated value.
As noted, the total area of filter material allocated to the pleating process is not necessarily converted to EFA usage. The uncertainty regarding the actual EFA of the pleated cartridge can, however, be resolved. A far more accurate prognostication based on the pleated model would then follow.
The exact amount of filter area used in constructing the pleated model should be known. Were it all converted to the EFA function, its extent could be calculated from flow rate studies as a function of applied differential pressures using Newtonian (clean) fluids at constant (ambient) temperature. Reverse osmosis product water could serve as the test fluid. The ideal flow rate would be the same as the flow rate if the filter matter in toto were used as EFA. That value could be obtained from the filter manufacturer, according to the filter type used, along with its porosity and thickness. The extent to which the expected flow is not realized would quantify the portion of the filter material not available as EFA as a result of pleating. The model filter's EFA would then be known with certainty, and its extrapolation could then be made with confidence.
The ultimate goal of the filter sizing exercise is to quantify the effective filter area (EFA) needed to filter an entire production batch over a set time frame. Such testing typically is done using a flat disc with a diameter of 47-mm. The proper implementation of the flow decline method of assessing EFA requirements necessitates knowing the precise EFA of the model filter, which can be measured using staining techniques.
A filter-sizing study using a 47-mm flat disc is intended as indicator trial, which is preliminary to a more meaningful verification trial, or even to a more cogent assurance trial. The latter will require filter models characterized by a pleated membrane design. The EFA assessment of the pleated filter models can be performed with confidence. This will result in dependable projections of the EFA required for the batch processing filters.
Maik W. Jornitz is the group vice president of marketing & product management, filtration and fermentation technologies, and Wayne Garafola is an application specialist, both at Sartorius Stedim Biotech, Bohemia, NY, 631.870.8463, firstname.lastname@example.orgTheodore H. Meltzer is the principal at Capitola Consulting Co., Bethesda, MD.
1. Jornitz MW, Soelkner PG, Meltzer TH. Modern sterile filtration—the economics. PharmTech Eur. 2003;15(6)29–33.
2. Priebe PM, Jornitz MW, Meltzer TH. Making an informed membrane filter choice. BioProcess Int. 2003;1(10)64–6.
3. Parenteral Drug Association. Technical report 26: Liquid sterilizing filters. Bethesda, MD: PDA; 2008.
4. Meltzer TH, Jornitz MW. Pharmaceutical filtration—The management of organism removal. PDA, Bethesda, MD: PDA and River Grove, IL: DHI Publishing LLC; 2006.
5. Jornitz MW, Meltzer TH. Comparing flow rates of microporous membranes. Eur J Parenteral Pharm Sci. 2009:14(1)19–23.
6. Jornitz MW, Meltzer TH. Filtration and purification in the biopharmaceutical industry. New York, NY: Informa; 2008
7. Jornitz MW, Meltzer TH, Bromm H, Priebe PM. Testing for the optimal filter membrane. Gen Eng News. 2004;24(13)50–51.
8. Jornitz MW, Meltzer TH, Bromm H, Priebe PM. Choosing the appropriate membrane filter—test requirements. PDA J Pharm Sci Technol. 2005 Mar–Apr;59(2)96–101.
9. Jornitz MW. Filter construction and design. In: MW Jornitz, editor. Sterile Filtration. Berlin, Heidelberg: Springer Verlag; 2006. p. 105–124.