Question and Answer Section — May 2014

William Brown and Margareth Marques
The following questions have been submitted by readers of Dissolution Technologies. Margareth Marques, Ph. D. and Will Brown, United States Phamacopeia, authored responses to each of the questions.
*Note: These are opinions and interpretations of the authors, and are not necessarily the official viewpoints of the USP

Email for correspondence: web@usp.org

Q What is the difference between dissolution and drug release?

The differences are historical, and the procedures are basi-cally the same. The initial thought was that dissolution would be applied to immediate-release dosage forms and drug release for modified-release dosage forms, but this concept is not valid anymore. Drug release is currently reserved for transdermal systems and osmotic-pump tablets.

Q The dissolution test in the USP monograph does not specify the concentration of the standard solution. Which concentration should be used?

In most cases, the concentration of the standard solution is obtaining by diluting an amount of drug substance equivalent to the product label claim in the total volume of dissolution medium used in the procedure. If the final concentration of this solution is outside the linearity range of your instrument, you can make any additional dilutions to have the final standard solution concentration within the linearity range of your equipment.

Q We demonstrated in vivo bioequivalence between the pilot batch and the reference product. We did the dissolution test of the pilot batch using the dissolution conditions specified in the FDA database, and we obtained slightly lower results. When we run the dissolution test with peak vessels, the results show some improvement. Can we use peak vessels in this case?

You need to evaluate what you are defining as slightly lower dissolution values. The dissolution methods in the FDA database are recommendations, and they are not mandatory. They are a very good starting point to help in the development of a dissolution test that is discriminative for your formulation. The FDA database does not give any information about acceptance criteria, so it is difficult to understand what you mean by slightly lower results. The dissolution test must be discriminative for the critical quality attributes of your product. The dissolution test is not an assay of the contents of the dosage unit. Its purpose is to investigate limitations to release, and therefore testing typically ends before 100% release. Peak vessels can only be used with appropriate justification. In most cases, you can get the same results just by increasing the rotation speed of the paddle.

Q How is the expiration date of vessels determined? Answer provided by Erika Stippler, Ph.D., ESS@usp.org.

There is no specific instruction related to age or frequency of use for dissolution vessels. However, the user has to be sure that the vessels in use comply with the pharmacopeial requirements, are intact, have undamaged inner surfaces, and are clean. The recommendation is to keep the vessels in the same position within the qualification interval. If a vessel needs to be replaced, a requalification of the instrument should be conducted. Several papers have been published regarding dissolution vessels:

Mark R. Liddell et al. Evaluation of Dissolution Vessel Dimensions and Irregularities. Dissolution Technol. 2007, 14 (1), 28-33.
Tanaka et al. Effect of Irregular Inner Shape of a Glass Vessel on Prednisone Dissolution Results. Dissolution Technol. 2005, 12 (4), 15-19.
Scott, P. Geometric Irregularities Common to Dissolution Vessels. Dissolution Technol. 2005, 12 (1), 18-21.

Q We are developing a dry powder for oral suspension, and during the accelerated stability study of this product, we have observed that the dissolution results at two months were on the lower side but within the specification limits, whereas for assay and other tests, the results were within the limits. At three months, the results for dissolution were passing but at S2 criteria, and the results for assay and other tests were within the limits. We studied the dissolution profile up to the infinity point (200 rpm for 10 min), and we found complete release of the drug. We found that the viscosity of the product had increased, and an undissolved mass of the suspension was observed at the bottom of the dissolution vessel. To overcome the lower release profile of the product, we evaluated two different approaches-increased agitation speed and peak vessels-and the percentage of drug released increased to 100% for the same batch. Do you have any recommendations in a case like this?

The dissolution test for suspensions is formulation dependent. The acceptance criteria for dissolution are defined using dissolution profiles of samples prepared during the development of the product, including the profiles obtained with the samples under accelerated stability studies. The acceptance criteria are going to be defined using all the dissolution profiles as well as the behavior of the product during the stability studies. The product is considered out of specification only if it fails S3. You need to discuss the changes in viscosity and any other product behavior with your formulation development group because it is important information that can help in improving the product. In the case of suspensions, the sample to be transferred to the dissolution vessel should be prepared according to the instructions to the practitioner and patient. It is advisable to review the sample preparation and the instructions to the patient. The objective of the dissolution testing is not to release 100% of the drug present in the dosage form. The dissolution test should be discriminative for the critical quality attributes of your product. Your comments indicate that the viscosity of the product changes with time. It will be very useful to evaluate if the dissolution test is discriminative for this parameter and assures that the appropriate dose is available to the patient over the treatment period. Peak vessels can only be used with appropriate justification. In most cases, you can get the same results by increasing the agitation speed of the paddle.

Q We are performing dissolution in water for a particular gelatin capsule product, and the product displays cross-linking. Based on the USP General Chapter <711> Dissolution, we need to follow the dissolution method and use pepsin at the specified medium at pH 7.0. It appears that these instructions are in conflict with the enzyme activity level and the required pH. Do you have any suggestions on how to proceed with this test?

The current and official test in <711> Dissolution states that pepsin can be added to the dissolution medium if the dissolution medium is water or has a pH below 6.8. Pancreatin can be added to the dissolution medium that has a pH above 6.8. As this is the official text, you need to start your evaluation using the conditions specified in the chapter. If they are not appropriate for your situation, you can consider modifications or use other conditions with appropriate justification. You need to have data showing that the conditions in <711> are not suitable for your product to justify any deviations from the official text. Water has a pH of about 6. Pepsin has good protease activity up to about pH 4 (see Piper, D. W.; Fenton, B. H. pH stability and activity curves of pepsin with special reference to their clinical importance. Gut 1965, 6 (5), 506-508). USP is aware of the deficiencies in the chapter <711>, and we have an expert panel working on improving and clarifying the text. Our experts are evaluating preliminary data showing that papain or bromelain may be appropriate proteases for the pH range from 4.0 to 6.8. Any revisions to <711> will be published in a future issue of Pharmacopeial Forum, available free of charge at www.usppf.com.

Q Does the release of the fill material have to be observed during the disintegration test? If it is not possible, as in the case of capsules containing a semisolid fill material, must the shell contain a hole in some area? Is it enough for the shell just to get soft even though it does not show a hole anywhere?

The capsule shell must open during the disintegration test. However, dissolution is seen as incorporating the disintegration process. Typically, a dissolution test is developed first, and then depending on the physical-chemical characteristics of the product and its behavior during the dissolution test and with appropriate justification, it might be replaced by the disintegration test. See the USP General Chapter <1094> Capsules-Dissolution Testing and Related Quality Attributes in the First Supplement of USP 37.

Q What is Q value in dissolution, and how is it determined? Is there any common Q value specified for all products? Does it change from product to product? How is Q used as an acceptance criterion?

Q is the amount of drug substance released from the dosage form into the dissolution medium over the specified test time. Q is in units of percentage of product label claim. Typically, Q is selected using dissolution profiles obtained during product development including the dissolution profiles of the samples under accelerated stability studies. The Q value varies from product to product because it depends on the release mechanism of the dosage form, on the solubility of the drug substance, and on the discriminative power of the dissolution test. Results are typically interpreted from an acceptance table. The acceptance tables have Q as a variable, and the value is given in the monograph.

Q USP General Chapter <711> Dissolution recommends using either pepsin or pancreatin depending on the pH of the medium. However, no significant enzyme activity is reported for any of those enzymes at pH values between 3 and 5. Is USP looking for alternatives to fix this situation?

Pepsin has effective protease activity up to pH 4 (see Piper et al., above), and pancreatin has good protease activity from pH 6.8 to about 10. We received information that papain or bromelain may be suitable proteases for the pH range 4-6.8. A revision to USP General Chapter <711> Dissolution is under development and will be published in a future issue of Pharmacopeial Forum (available free of charge at www.usppf.com).

Q How were the USP General Chapter <711> Dissolution limits selected for either pepsin or pancreatin? Is there any situation where higher amounts of enzyme can be used?

The amounts of not more than 750,000 Units per 1000 mL for pepsin and not more than 1750 Units of protease activity per 1000 mL for pancreatin were selected based on the results of the Gelatin Capsule Working Group organized by FDA in the early 90s (see Aikman et al. Collaborative development of two-tier dissolution testing for gelatin capsules and gelatin-coated tablets using enzyme-containing media. Pharm. Forum 1998, 24 (5), 7045-7050). The activity of pepsin should be determined by the procedure employing hemoglobin under Pepsin, Purified in the Reagent Specifications section of USP-NF. This procedure has been revised, and the revisions were published in Pharmacopeial Forum (PF) 38 (3) [May-June 2012] and PF 39 (6) [Nov-Dec 2013]. Extensive comments on these revisions were received, and a new version of the Pepsin activity determination procedure will be published in a future issue of PF (available free of charge at www.usppf.com).

Enzymatic activity depends strongly on the type of substrate and on the conditions of the test. Activity values obtained by procedures using different substrates, different test conditions, or both are neither equivalent nor interchangeable. The activity of pepsin and pancreatin used in dissolution testing must be determined by the procedures stated in USP. Before any decisions are made on changing the amounts of enzyme to be added to the dissolution medium, it is necessary to verify that the dissolution failure is caused by cross-linking. This can be done by visual observation (the capsule swells but does not rupture or open), by an instrumental technique (FTIR, DSC, NMR, etc.), or by transferring the capsule contents to a capsule that does not have cross-linking. In addition, it is necessary to confirm that the enzyme activity was determined by the procedure recommended by USP.

Q In the case of a capsule that fails S1, what is the right order for the following events: OOS investigation, S2 and S3 stages, S1 plus enzymes? Can I test S2 or S3 with enzymes?

Dissolution failure is considered only after the product has failed S3 or L3 (see the appropriate Acceptance Table in <711> Dissolution). Therefore, out-of-specification (OOS) investigations should be carried out only if the product failed L3 or S3. The text in <711> Dissolution is not very clear, and a revision is under development. If the product failed at S1 or L1 and the failure is due to cross-linking in the gelatin capsule, our recommendation is that the test can be repeated with the addition of the appropriate enzyme to the dissolution medium with six units at S1 or L1 stage. The appropriate acceptance table is used.

Q If I need to use enzymes during a stability study, are enzymes then used at subsequent sample points?

The text in <711> Dissolution is not very clear, and a revision is under development. First, you need to be sure that any failures are due to cross-linking in the gelatin capsule. Once started, the cross-linking process continues regardless of the action of a cross-linking agent. If the failure is due to cross-linking, our recommendation is to add the enzyme at S1 for all the remaining sample points in the stability study.

Q ICH guidance Q1E establishes that the following situation is not a significant change during stability studies at the accelerated condition: ".failure to meet acceptance criteria for dissolution for 12 units of a gelatin capsules or gel-coated tablet if the failure can be unequivocally attributed to cross-linking." How is the USP General Chapter <711> Dissolution in agreement with this statement? How can we consider unequivocally that a dissolution failure is due to cross-linking in the gelatin capsules?

The current version of <711> Dissolution does not explicitly state that the failure must be due to cross-linking in the gelatin capsule or gelatin-coated tablet. We are aware of this deficiency, and a revision of the chapter is under development. Evidence of cross-linking in gelatin can be by visual observation (the capsule hydrates, swells but does not open or rupture), by an instrumental technique (FTIR, NMR, DSC, etc.), or by transferring the contents to a non-cross-linked capsule.

Q Is there any protease activity test for pancreatin?

For dissolution applications, the protease activity of pancreatin should be determined by the Assay for Protease Activity in the USP monograph for Pancreatin.

Q Does USP have any data regarding stability of open containers of enzymes? Should I test enzyme stability and activity of in each dissolution medium to be used for my product?

Pepsin as a dry powder is stable for three years at room temperature and for several years when stored at -10 to -25 °C (information from one of the major manufacturers of pepsin). Pepsin in solution at pH 4.4 is stable at -20 °C for about 2-3 months (Rajagopalan, T. G. et. al. Pepsin from pepsinogen. Preparation and properties. J. Biol. Chem. 1966, 241 (21), 4940-4950). At pH 1.5, pepsin exhibits about 90% of maximum activity, and at pH 4.5 about 35% of maximum activity (Bohak, Z. Purification and characterization of chicken pepsinogen and chicken pepsin. J. Biol. Chem. 1969, 244 (17), 4638-4648).

Pancreatin as a dry powder has stable protease activity at room temperature for two years; the lipase and amylase activities are not as stable. In solution, the protease activity of pancreatin is stable for 16 h at pH 4-9 (information from one of the major manufacturers of pancreatin).

As the activity determination depends strongly on the type of substrate and on the test conditions, the activities of pepsin and pancreatin should be determined using the procedures recommended in USP-NF.

Q Should enzymes be added to the dissolution medium before or after degassing?

Although this needs experimental verification, the enzyme is typically added after the degassing procedure.

Q Are there any guidances for dissolution methods for chewable tablets?

We are not aware of any specific guidance on dissolution testing of chewable tablets.

Q Should the dissolution procedure and acceptance criteria be the same for an immediate-release dosage form and chewable tablets?

There are no general acceptance criteria for dissolution of any type of tablets. The acceptance criteria are defined in a case-by-case approach by evaluating the dissolution profiles of all the samples tested during product development including the samples under stability studies. Typically, chewable tablets are harder than ordinary immediate-release tablets. Consequently for them, the dissolution test may be longer or have a higher agitation speed when compared with the immediate-release tablets containing the same drug substance.

Q The USP monograph for Norethindrone Acetate and Ethinyl Estradiol Tablets states that the sample solution should be centrifugated and the supernatant used for the quantitation procedure. Is centrifugation against the principle of dissolution as per <711>? Should the solutions be filtered even if both active ingredients are adsorbed in the filter?

The General Chapter <711> Dissolution does not specify if the sample should be filtered or centrifuged or treated in any other way. The general chapter says to treat the sample as directed in the monograph because the treatment will depend on the drug substance, on the dosage form, and on the quantitation procedure. The monograph conditions supersede statements in USP general chapters. The dissolution test in the USP monograph for Norethindrome Acetate and Ethinyl Estradiol Tablets is the test approved by FDA for some products available in the United States market. The text is based on the validation report submitted by the monograph sponsor. Any changes to the text would only be made based on review and approval from FDA following requests for revision.

Q I am developing a generic version of glimepiride tablets, and the product does not meet the acceptance criteria for Dissolution Test 1 in the USP monograph for this product. The disintegration time for each pilot batch was good (30-40 s), but the product did not meet the dissolution acceptance criteria. Increasing the hardness of the tablet increased the disintegration time but produced no changes in the dissolution results. Do you have any suggestions on how to improve the dissolution results of this product?

Glimepiride is practically insoluble in aqueous media. Any drug must be in solution to be absorbed. For a number of reasons, each manufacturer will use a different formulation strategy to increase the solubility and the absorption of glimepiride. Consequently, the dissolution test for glimepiride tablets reflects these formulation differences. You will need to develop a dissolution test that is discriminative for the critical quality attributes of your formulation. Hardness may not be the most critical parameter. Glimepiride has two polymorphic forms, and the proportion of the two polymorphic forms is critical. Other parameters that may be critical for poorly soluble drugs are particle size and particle size distribution.

Q Why are the coefficient of variation limits different for the performance verification test for baskets and paddles (NMT 7.2% for baskets and NMT 4.9% for paddles)? How do hydrodynamics with the paddle or any other physical parameter impact the coefficient of variation in performance verification testing for baskets and paddles? Answer provided by Erika Stippler, Ph.D., ESS@usp.org.

The acceptance ranges for the Performance Verification Test (PVT) for both USP Apparatus 1 and 2 are established statistically from a pool of experimental data generated by an international, multi-lab collaborative study. Generally, the results of the collaborative study and the approach used to establish the acceptance limits are published and readily available. It is well known that the hydrodynamic conditions with the paddle are different from those with the basket. The tablet surface experiences different shear in the two apparatus. There are several published papers that discuss this topic.

Q The USP General Chapter <701> Disintegration does not specify limits for the disintegration time of uncoated tablets. If the disintegration time is not specified in the USP monograph, what is the disintegration time to be considered for uncoated tablets?

The acceptance criterion for a disintegration test is defined in a case-by-case approach. If the USP monograph has a disintegration test, it will have acceptance criteria for it. If the USP monograph does not have a disintegration test, it will have a dissolution test. Orally disintegrating tablets are the only dosage form that may require both tests. A disintegration test is not a mandatory part of a monograph. First, develop a dissolution test. Depending on the characteristics of the product and its behavior during the dissolution test, the dissolution test may sometimes be replaced by a disintegration test with proper justification.

Q What dissolution medium can be used with poorly water-soluble drugs (BCS Class 2)? If the drug substance is not soluble in that medium, how should the standard solutions for the calibration curve be prepared?

Consider the solubility of the drug substance in several dissolution media within the physiological pH range at 37 °C and the composition of the formulation. Several papers and book chapters explain how to make the selection of the dissolution medium. Depending on the composition of your formulation, a surfactant may be added to the dissolution medium. See the papers:

Noory, C.; Tran, N.; Ouderkirk, L.; Shah, V. Steps for Development of a Dissolution Test for Sparingly Water-Soluble Drug Products. Dissolution Technol. 2000, 7 (1), 16-18. Rohrs, B. R. Dissolution Method Development for Poorly Soluble Compounds. Dissolution Technol. 2001, 8 (3), 6-12.

If the drug has low solubility in the dissolution medium, a stock solution may be prepared in an organic solvent and then further diluted with dissolution medium to the appropriate concentration.

Q We have a dissolution method where the duration of the test is 30 min. We are going to change this time to 45 min. Is it necessary to revalidate the dissolution method? If yes, should it be a complete or partial revalidation?

This type of modification in a dissolution test is critical and requires proper justification. Dissolution profiles are obtained during the development and validation of a dissolution method. Run the dissolution profile until it reaches a plateau or until around 85% of the product label claim has been released. Depending on how the original method was validated, the 45-min time point may have already been included in the dissolution profile.