Often a reason provided, in the literature, that it is a test to differentiate drug release characteristics of a product reflecting potential impact of formulation and/or manufacturing differences. This type of tests usually does not have a link or association with in vivo drug release characteristics of the products. Such tests are often referred to as “discriminatory tests”. The underlying reason for such a “discriminatory test” is that if the test relates dissolution differences to formulation/manufacturing differences, then the test will flag potential deviation from the expected behavior of the product in vivo (humans). The main assumption here is that the differences in formulation and/or manufacturing attributes would result in vivo differences in drug release characteristics which may produce a substandard product.  This unfortunately is an incorrect assumption, because the differences in formulation and/or manufacturing, on their own, do not necessarily result in differences in vivo drug release, at least not in most cases. If this assumption would have been correct then generic products, and the industry, would not have existed. The reason being, the generic products, which are based on vastly different formulation and manufacturing attributes, but are required to provide the same drug release characteristics in vivo (humans).  In addition, the generic products are also required to provide similar dissolution characteristics, thus forms the basis of pharmacopeial (USP) testing.

Therefore, developing or conducting an in vitro dissolution test just to evaluate/assess differences in formulation/manufacturing, without its link to in vivo, is of limited value or use, and in fact may be an incorrect practice.

IVIVC (In vitro-in vivo correlation) is a desired feature in the practice of drug dissolution testing. An appropriate IVIVC provides credibility to an in vitro dissolution test by avoiding false negative indications concerning quality and manufacturing of a product. It further enhances economic benefits to the manufactures by providing efficient development and modification of products, thus obtaining regulatory approvals.

Developing an IVIVC may be considered a two-part process: (1) analyzing the in vitro dissolution data and relating it to in vivo results. This has been the subject of the last few posts (1, 2, 3); (2) conducting an actual dissolution test for generating appropriate data. This post is regarding the later aspect.

For an appropriate dissolution test, in general and in particular for developing IVIVC, one requires to conduct the test selecting experimental conditions to simulate an in vivo environment as closely as possible. Commonly the following experimental conditions should be considered in this regard.

1. A dissolution medium should be an aqueous solution having a pH in the range of 5-7 and be maintained at 37C. The expected amount of the drug present in the product must be able to freely dissolve in the volume of the medium used, often 900 mL. If the drug is not freely soluble in water as such, then small amounts of solublizing agent such as SLS may be used.
2.  The dissolution medium should not be de-aerated. Preference should be given that the medium be equilibrated at 37C with dissolved air/gasses, particularly for IVIVC studies.
3.  An apparatus should be selected to have an appropriate mechanism to provide thorough but gentle mixing and stirring for an efficient product/medium interaction. Use of sinkers may be avoided as these often alter the dissolution characteristics of the test products. Paddle and basket apparatuses are known for their inefficient stirring and mixing, thus their use should be critically evaluated before use for IVIVC studies.
4. Frequent samples (8-10) should be withdrawn to obtain a smooth dissolution profile leading to complete dissolution within the dosing interval of the test product in humans.
5. If the dissolution results are not as expected, then the product/formulation should be modified to obtain the desired/expected release characteristics of the product. However, altering experimental conditions such as medium, apparatus, rpm etc. should be avoided as these are generally linked to GI physiology which remains the same for test to test or product to product. Obtaining dissolution results by altering testing (experimental) conditions may void the test for IVIVC purposes.

As explained in an earlier post, commonly used convolution/deconvolution techniques for IVIVC purposes, link three functions together. The three functions are input (absorption/dissolution results), output (plasma drug concentrations) and weighting function (usually plasma drug concentrations following an intravenous dose).

Deconvolution will be the option one would use when plasma drug concentrations of the test products are available and one would like to determine the in vivo dissolution results. These in vivo dissolution results are compared with the in vitro dissolution results.

Convolution will be the option one would use when in vitro dissolution results are available and one would like to determine plasma drug concentrations of the test product. During the product developmental stage, where a formulator likes to have an idea of potential in vivo output, a convolution technique would be the only choice. For comparison of release (dissolution) characteristics of products for generic developments or product modifications etc., again one may use the convolution technique. In this case, based on obtained dissolution results from two or more products, one would be able to obtain respective plasma drug concentration profiles, which can be compared using standard and accepted parameters of Cmax and area under the plasma drug concentration curves (AUC).

Studies have shown that the convolution technique provides better accuracy of outcome (plasma drug concentrations) than the deconvolution technique.  Moreover, computation-wise, convolution technique could be simplified and calculation may be performed using simple spreadsheet software rather complex mathematical software. Therefore, it is suggested that one may consider convolution as a first choice for developing IVIVC.

A plasma drug concentration-time profile is usually the net effect of two simultaneous processes: (1) absorption of the drug from the GI tract. As absorption is proportional to drug dissolution thus absorption and dissolution are used interchangeably; (2) elimination of the drug from the blood. These two actions, and their net effect, are represented by three profiles and are shown in the figure. In mathematical terminology, these three curves (profiles) are known as functions, dissolution or absorption as input, blood concentrations as output and the elimination as the weighting factor or function. To further simplify, in the analogy of linear regression used for calibration curves, output function may be considered as “Y or dependent variable”, “X or input function” and “M or slope/proportionality constant”. In case of linear regression analysis X, Y and M parameters have values (numbers), however, in the case of drug concentration profiles, these are functions. So solving these function based equations is a bit more complicated.

The procedure is similar to that of linear regression which is commonly used in establishing calibration curves and then using the calibration curve to determine the unknown concentrations or response (e.g. absorption or peak height/area values). So, if “Y” is known then “X” may be determined and vice versa. Similarly, if input function is known, one can determine output function and vice versa. Determining output function (plasma blood concentrations), if input function (dissolution results) is available, the procedure will be called convolution technique and inverse of it, that is obtaining input function (absorption/dissolution results) if output function is provided, the procedure will be called deconvolution.

There are computer software available which provide the capability of solving for a function when the others are available. However, the convolution approach could be simpler where use of commonly available spreadsheet software may also be used.  For further detail see, Qureshi, SA. In Vitro-In Vivo Correlation (IVIVC) and Determining Drug Concentrations in Blood from Dissolution Testing – A Simple and Practical Approach. The Open Drug Delivery Journal, 2010, 4, 38-47. (Link)

Theoretical Consideration:  The most commonly used definition of IVIVC (In Vitro/In Vivo Correlation) is the one described in one of the FDA guidance documents (link). It defines IVIVC as a predictive mathematical model describing the relationship between an in vitro property of a dosage form (usually the rate or extent of drug dissolution or release) and a relevant in vivo response, e.g., plasma drug concentration or amount of drug absorbed.

In this regard, the most sought after relationship is of “Level A”, which is further defined as a predictive mathematical model for the relationship between the entire in vitro dissolution/release time course and the entire in vivo response time course, e.g., the time course of plasma drug concentration or amount of drug absorbed.

Practical Consideration: On the practical side, the purpose of IVIVC is to use drug dissolution results from two or more products to predict similarity or dissimilarity of expected plasma drug concentration (profiles). Before one considers relating in vitro results to in vivo, one has to establish as to how one will establish similarity or dissimilarity of in vivo response i.e. plasma drug concentration profiles. The methodology of establishing similarity or dissimilarity of plasma drug concentrations profile is known as bioequivalence testing. There are very well established guidances and standards available for establishing bioequivalence between drug profiles and products.

Ideally, therefore, one should focus on predicting or calculating plasma drug concentration profiles from drug dissolution results for an appropriate IVIVC. A common mathematical technique employed for this purpose is known as convolution, which basically convolutes (combines) dissolution results (profile) with plasma concentrations following intravenous (IV) drug administration to provide expected plasma concentrations for solid oral dosage forms. In mathematical terminology, dissolution results become an input function and plasma concentrations (e.g. from IV) become a weighting factor or function resulting in an output function representing plasma concentrations for the solid oral product.

Further details about this methodology and its use will be described in future posts.

Bio-waiver is a term used for establishing and accepting quality of a pharmaceutical product (tablet and capsule) based on in vitro drug dissolution testing without corresponding bioavailability/bioequivalence data.

The scientific rationale behind this practice is that, in general, the bioavailability of a drug product depends on its release and solution formation (dissolution) characteristics. This, in vivo (physiological) dissolution is monitored or simulated in vitro by a dissolution test. Therefore, an assumption made here is that the in vitro dissolution test will appropriately and accurately reflect in vivo dissolution in humans and thus the bioavailability/bioequivalence characteristics of the products. This linkage is commonly referred to an in vitro-in vivo (co)relationship or IVIVC. Therefore, a pre-requisite for bio-waiver practices is a well established IVIVC.

It is, however, generally well recognized and documented in the literature that current practices of dissolution testing do not relate well to in vivo outcomes or often lack IVIVC. Therefore, presently, a bio-waiver should be considered as a scientifically weak case.

There are, however, guidances available for considering bio-waivers in certain, very specific, cases.  For example, bio-waivers are considered for products having drugs with high aqueous solubilities and absorbabilities from the GI tract in humans. In addition, the products should show fast dissolution. The underlying thought is that if drugs are highly soluble and absorbable, and the products show fast and complete in vitro dissolution (usually in less than 30 minutes), then the products are not expected to pose any potential problems in vivo, and thus they may be considered for bio-waivers. Therefore, it may be assumed that current practices of bio-waivers are based on faith rather their scientific merits.

In short, for an appropriate and scientifically valid bio-waiver inference, one has to establish IVIVC, with evidence that the techniques and methodologies (dissolution apparatus, medium and associated experimental conditions) employed are indeed capable of providing physiologically relevant results.

In continuation of the earlier post (link), this post describes some of the steps which are commonly described in the literature as method development, but should not be considered as method development steps. These steps are usually variations in experimental conditions to achieve certain desired characteristics of a dissolution test such as discriminatory, improved reproducibility and/or bio-relevancy. The variations can be are numerous, such as choice of apparatus (paddle or basket), spindle rpm (50, 75, or 100), media (water or buffer), pH (between 1 and 6), choice of de-aeration or its technique, use and choice of a sinker, etc.

Considering these steps or practices as method development is incorrect because of two reasons:

1. These practices are commonly used during product development stage. At this stage an analyst is working with the variations in the product (formulation and manufacturing attributes) which requires a fixed dissolution method, not variations in the method itself, to evaluate the impact of product variations.
2. The products are developed for human use. The drug is expected to be released from the product in the GI tract environment, which remains constant, from product to product. The in vitro dissolution testing conditions simulate this GI tract environment, therefore, these conditions should also remain constant.

It is, therefore, critical that a dissolution test method should be decided and fixed, reflecting GI tract environment, prior to working on the development of a product.

The role of an (or any) analytical method validation (chromatographic/spectrophotometric) is to demonstrate that the method is capable of measuring an analyte accurately (accuracy, which includes specificity) and reliably (precision, which includes repeatability and reproducibility). In addition, if the analyte is expected to be in a wider range e.g. zero to 100 %, which is usually the case in dissolution testing, then one has to establish that concentrations and responses have a linear relationship (linearity), by measuring responses at different concentrations.

All of the above mentioned practices (tests) boil down to determining responses (UV absorptions or peak height/area for chromatography) against concentrations and are to be done in replicate (5/6 times) to be able to determine a standard deviation (variance) to establish confidence in the results. In short, if one has different solutions of 100, 50 and 25% concentrations (strength) of drug and measure their responses, which come out in the ratios as the concentrations (by doing it repeatedly 5/6 times), then the method has been validated.

For the purpose of drug dissolution testing, one has to demonstrate that if the drug is in solution then the analytical method is capable of measuring it accurately and reliably. Therefore, for validation of such methods, one needs to add the drug (“spiking”) in solution form to a dissolution testing apparatus  i.e. vessel containing required volume of medium maintained at 37 ºC and spindle rotating. Samples are withdrawn and processed exactly as if these were from a product (filtration, dilution/concentration, extraction etc) and responses are measured accordingly. If responses and concentrations are as one would expect (as explained above), then that dissolution method has been validated.

One should, then, be able to use this method (validated) for measuring dissolution of a drug from a product. Method validation steps are independent of drugs and products.

On the other hand, a method development exercise is drug and product dependent. In (analytical) method development, an analyst needs to select appropriate parameters such as wavelength, chromatographic column, dilution, extraction steps, filters etc. Once such parameters are established, one then moves to the method validation exercise as a second step.