Chapter 18
`Bioavailability and
`Bioequivalence Testing
`
`Steven B. Johnson, PharmD
`
`INTRODUCTION 349
`DISSOLUTION Jc19
`ABSORPTION FACTORS 350
`BIOEQUIVALENCE TESTING 351
`METHODS FOR DETERMINING BIOEQUIVALENCE 351
`
`MINIMIZING THE NEED FOR BIOEQUIVALENCE
`STUDIES 352
`
`EVALUATION OF BIOEQUIVALENCE DATA 352
`BIOEQUIVALENCE ASSESSMENT AND DATA
`EVALUATION 352
`
`CRITERIA FOR BIOEQUIVALENCE 35,~
`
`STUDY DESIGN 356
`
`THERAPEUTIC EQUIVALENCE EVALUATIONS 358
`
`INTRODUCTION
`Understanding the concepts of bioavailability and bioequiva-
`lence testing is essential in the drug development process be-
`cause they create the foundation for regulatory decision making
`when evaluating formulation changes and lot-to-lot consistency
`in innovator products. They also serve as the primary compo-
`nents to demonstrate therapeutic equivalence between generic
`·
`products and the reference innovator product.
`Changes in bioavailabiJity can be thought of in terms of chang-
`es in exposure to the drug. If these changes are substantial, then
`they can alter the safety and efficacy profile of the compound in
`question. The bioavailability of orally administered drugs can be
`affected by numerous factors. These include food or fed state,
`differences in drug metabolism, drug-drug interactions, gastro-
`intestinal transit time, and changes in dosage form release char-
`acteristics (especially for modified release products).
`Bioequivalence is an important consideration in ensuring lot-
`to-lot consistency, including whenever evaluating changes in a
`marketed product's formulation, manufacturing process, and dos-
`age strength. Bioequivalence is also critical in regulatory author-
`ity decision malting when determining whether a generic product
`is therapeutically equivalent to the original innovator product.
`In addition, chemical equivalence, lot-to-lot uniformity of
`physicochemical characteristics, and stability equivalence are
`other factors that are important, as they too can affect product
`quality. In this chapter, bioavailability and bioequivalence top-
`ics are emphasized for solid oral dosage forms. However, many
`of the general concepts can also be applied to other dosage
`forms, including biologics.
`
`GENERAL CONCEPTS
`The terms used in this chapter require careful definition, since,
`as in any area, some terms have been used in different contexts
`by different authors.
`Bioa'Vailability is a term that indicates measurement of both
`the rate of drug absorption and the total amount (extent) of
`drug that reaches the systemic circulation from an adminis-
`tered dosage form. It is specific to the parent or active drug
`substance as contrasted to metabolites.
`Equi'Valence is more a general and relative term that indi-
`cates a comparison of one drug product with another. Equiva-
`lence may be defined in several ways:
`
`• Bioequi'Valence indicates that a drug in two or more
`similar dosage forms reaches the systemic circulation at
`the same relative rate and extent (i.e., the plasma level
`
`profiles of the drug obtained using the two dosage forms
`are the same).
`• Chemical equi'Valence considers that two or more dosage
`forms contain the same labeled quantities (within speci-
`fied limits) of the drug.
`• Clinical equi'Valence occurs when the same drug from
`two or more dosage forms gives identical in 'Vi'Vo effects as
`measured by a pharmacological response or by control of
`a symptom or disease.
`• Pharmaceutical equi'Valence refers to two drug products
`with the same dosage form and same strength.
`• Therapeutic equi'Valence implies that two brands of a
`drug product are expected to yield the same clinical
`result. The FDA specifically uses the term "therapeutic
`equivalence" in the evaluation of multi-source prescrip-
`tion drug products (generic drugs).
`
`Area under the Concentration-Time Curve (AUG) is the inte-
`gral of the concentration- time curve after administration of a single
`dose of drug or after achieving a steady state. The calculated area
`under the serum, blood, or plasma concentration-time curve is re-
`ported in amount/volume multiplied by time (e.g., IW'mL x h or
`!1/100 mL x h) and can be considered representative of the amount
`of drug absorbed. Several variants of AUC exist, including AU Co.,;
`AUC0_; and AU~, SS• corresponding to the calculated area from
`time zero to a truncated time point (e.g., AUC048), the total area
`under the curve, and the area when steady state has been achieved.
`Peak-height Concentration (Cmax) is the peal' of the blood
`level-time curve and represents the highest drug concentration
`achieved after drug administration.
`Time of Peak Concentration (Tmax) is the measured length of
`time necessary to achieve the maximum concentration (~")
`after drug administration.
`
`DISSOLUTION
`For a drug to be absorbed, it must first go into solution. In Figure
`18-1 the steps in the dissolution and subsequent absorption
`process of a tablet or capsule dosage form are outlined. Simi-
`lar profiles could be obtained for any solid or semisolid dosage
`form, including oral suspensions, parenteral suspensions, and
`suppositories. The theory and mechanics of drug dissolution
`rate are described in detail in Chapter 22. The physical charac-
`teristics of the drug and the composition of the tablet (dosage
`form) can have an effect on the rates of disintegration, deaggre-
`gation, and dissolution of the drug. As such, these can affect the
`rate of absorption and resultant blood levels of the drug.
`
`349
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`350
`
`Solid dosage form
`Dissoluti~n
`of
`drug
`
`°• IARMfiCEUTICAl_ ANALYSIS AND QUALITY CONTROL
`Disintegration
`Deaggregation
`Granules
`
`Small particles
`
`drug
`
`j Diss~~ution/issolution
`Drug in solution ! Drug absorption
`
`of
`drug
`
`Drug in blood
`Figure 18-1. Sequence of events involved in the dissolution and
`absorption of a drug from a solid oral dosage form.
`
`An important aspect of product quality for marl<eted oral solid
`dosage forms relates to dissolution testing. The dosage forms ac-
`tually used by patients will be from lots that have not directly
`undergone human bioavailability testing. Once adequate product
`quality has been established by bioavailability testing, subsequent
`batches manufactured using the same formulation, equipment,
`and process are likely to be bioequivalent to the original batch
`tested in humans. This is an important concept in the regulatory
`control of product quality and is where in vitro testing such as
`assay, content unifom1ity, tablet hardness, and dissolution are
`important. Among these several in vitro tests, dissolution test-
`ing is probably the most important, related to bioavailability. As
`part of the drug approval process, a dissolution test procedure is
`established for all oral solid dosage forms. These dissolution tests
`are incorporated into the United States Pharmacopeia (USP) and
`apply both to innovator and generic drug products. All marketed
`batches of these drug products must meet the Abbreviated New
`Drug Application (ANDA)/New Drug Application (NDA)IUSP dis-
`solution tests throughout the shelf-life of tl1e product. Products
`failing their approved dissolution test and/or a USP dissolution
`test must be removed from the market.
`PROPERTIES OF THE DRUG
`The physical characteristics of the drug that can alter bioavail-
`ability are discussed in Chapters 9 and 54 and consist of the
`polymorphic crystal form, choice of the salt form, particle size,
`hydrated or anhydrous form, wettability, and solubility of the
`drug. Chapter 9 also discusses several other properties that can
`adversely affect drug product quality. These factors should be
`investigated during product development and should not, there-
`fore, affect the bioavailability of the drug product.
`PROPERTIES OF THE DOSAGE FORM
`The various components of the solid or semisolid dosage form,
`other than the active ingredient, are discussed in Chapter 45.
`Only an overview, for tablet dosage forms, is given here. In ad-
`dition to the active ingredient, a tablet product usually will con-
`tain the following types of inactive ingredients:
`• Glidants are used to provide a free-flowing powder from
`the mix of tablet ingredients so that the material will flow
`when used on a tablet machine.
`• Binders provide cohesiveness to the tablet. Too little
`binder will produce tablets that do not maintain ilieir
`integrity; too much may affect adversely the release (dis-
`solution rate) of the drug from the tablet.
`• Fillers are used to give the powder bulk so that an accept-
`ably sized tablet is produced. Most commercial tablets
`weigh from 100 mg to 500 mg, so it is obvious that for
`many potent drugs the filler constitutes a large portion
`of the tablet. Binding of drug to the fillers may occur and
`affect bioavailability.
`• Disintegrants are used to cause the tablets to disintegrate
`when exposed to an aqueous environment. Too much
`will produce tablets that may disintegrate in the bottle
`because of atmospheric moisture, and too little may be
`insufficient for disintegration to occur and may therefore
`
`alter the rate and extent of release of the drug from the
`dosage form.
`• Lubricants are used to enhance the flow of the powder
`through the tablet machine and to prevent sticking of the
`tablet in the die of the tablet machine after the tablet is
`compressed. Lubricants are usually hydrophobic materi-
`als such as stearic acid, magnesium, or calcium stearate.
`Too little lubricant will not permit satisfactory tablets to
`be made; too much may produce a tablet with a water-
`impervious hydrophobic coat, which can inhibit the
`disintegration of the tablet and dissolution of the drug.
`
`ABSORPTION FACTORS
`A significant factor related to drug bioavailability is the fact
`that many drugs are administered, not as a solution, but as a
`solid dosage form. Optimal bioavailability might be expected
`from a solution, since a solid drug must first dissolve to be ab-
`sorbed, but considerations such as drug stability, unpalatable
`taste, and the desired duration of action (for controlled-release
`drug products) may prevent tl1e development of solution-based
`dosage form.
`In the dose titration of any patiem the objective is, in con-
`ceptual terms, to attain and maintain a blood level that exceeds
`the minimum effective level required for response but does not
`exceed the minimum toxic (side-effect) level. This is shown
`graphically in Figure 18-2. There are several absorption factors
`that can affect the general shape of this blood-level curve and
`thus drug response.
`DOSE ADMINISTERED
`The blood levels wi!J rise and fall in proportion to the dose
`administered.
`AMOUNT OF DRUG ABSORBED
`Blood levels achieved are also dependent on the amount of drug
`absorbed. For example, the effect of having only one-half of the
`drug absorbed from a dosage form is equivalent to lowering the
`dose (Figure 18-3).
`RATE OF ABSORPTION
`If absorption from tile dosage form is more rapid than the rate
`of absorption that gave profi.le C in Figure 18-4, minimum toxic
`(side-effect) levels may be exceeded. If absorption from the
`dosage form is sufficiently slow, minimum effective levels may
`never be attained, as in profi.le B in Figure 18-4.
`In either instance, the time course and extent of clinical re-
`sponse to the drug may be altered because of changes in dose or
`rate and extent of absorption.
`Both factors, rate and extent of drug absorption, can be
`affected by the dosage form in which the drug is contained. The
`
`Minimum toxic level
`
`c
`0
`:;::>
`~ c Q)
`0 c
`0
`0
`«l E 1/)
`«l a:
`
`Time
`Figure 18-2. Typical plasma-level curve of a drug with effective and
`toxic (side-effect) profile levels defined.
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`Minimum toxic level
`---------------------------------------------------·
`
`Minimum effective level
`
`c
`.2
`~
`'E Q) u
`c
`0 u
`ro
`~
`ro a:
`
`nme
`Figure 18-3. Effect of the extent of drug absorption from a dosage
`form on drug-plasma levels and efficacy. The extent of absorption
`from dosage form B is 50% of that from dosage form A.
`
`Minimum toxic level
`
`c
`
`c
`0
`~
`'E
`Q) u c
`8
`ro
`E (/) ro a:
`
`Time
`Figure 18-4. Effect of the rate of drug absorption from a dosage form
`on the plasma-level profile and efficacy. The rates of absorption from
`dosage forms Band Care 1/10 and 10 times those from dosage form A.
`
`effect of rate of absorption may be intentional, as in controlled-
`release products, or unintentional, for example, as brought
`about by a change in the composition and/or method of manu-
`facture of the dosage form.
`The choice of the inactive ingredients (excipients) used to
`prepare a dosage form is up to the individual manufacturer. It is
`through these changes in composition and manufacturing tech-
`nique that unintended changes in bioavailability and bioequiva-
`lence may occur. Revalidation of bioequivalence may be needed
`for major changes in the manufacturing process, whereas small
`changes may not raise significant bioavailability concerns.
`In situations involving minor changes in the manufacturing
`process, comparative dissolution testing of the original and
`reformulated product provides adequate documentation of con-
`tinued product quality, if the resulting dissolution profiles are
`similar. These considerations apply to all drug manufacturers,
`both innovator and generic companies. A description of the for-
`mulation of dosage forms and the factors that must be consid-
`ered is given in Chapter 9.
`
`BIOEQUIVALENCE TESTING
`The awareness of
`the potential for clinical differences
`between otherwise chemically equivalent drug products has
`been brought about by a multiplicity of factors that include,
`among others:
`
`• better methods for clinical efficacy evaluation
`• development of techniques to measure microgram or
`nanogram quantities of drugs in biological fluids
`
`351
`BIOAVAILABILITY AND BIOEOUIVALENCE TESTING
`• improvements in the technology of dosage form formula-
`tion and physical testing
`• awareness of reported clinical differences from the litera-
`ture in otherwise similar products
`• increased cost of classical clinical evaluation
`• objective and quantitative nature of bioavailability tests
`• an increase in the number of chemically equivalent
`products on the market because of patent expirations
`and the Drug Price Competition and Patent Term
`Restoration Act of 1984 (1-Iatch-Waxman Act), which
`established the generic drug approval procedures that
`are in place today.
`
`The increase in the number of drugs that are available from
`multiple sources frequently has placed people involved in the
`delivery of healthcare in the position of having to select one
`from among several marketed products. As with any decision,
`the more data available, the more comfortable one is in arriv-
`ing at the final decision. The need to make these choices, in
`light of the potential failure to demonstrate in vivo equivalence
`between products or different batches of the same product, has
`increased the demand for quantitative data. Bioequivalence
`testing represents a bridging alternative to clinical testing for
`efficacy and safety in such cases and is the means by which ge-
`neric drugs are approved for marketing. In addition, this is also
`the means by which the quality of all drug products is main-
`tained in situations involving major changes in formulation or
`manufacturing process.
`Requirements for bioequivalence data on drug products
`should be applied reasonably. The reason for bioequivalence
`testing should not be overlooked (i.e., it is used as a surrogate,
`in certain situations, for the clinical evaluation of drug prod-
`ucts). In some instances, bioequivalence data cannot reliably
`be obtained if the bioanalytical methodology is not available.
`However, in such cases pharmacodynamic data may provide
`a more sensitive, objective evaluation of a product's thera-
`peutic equivalence and may be explored as an alternative
`evaluation method in the absence of relevant bioanalytical
`methodology.
`Basic pharmacokinetic evaluation of bioavailability data is
`not necessary to show bioequivalence of two drug products.
`Pharmacokinetics has its major utility in the prediction or pro-
`jection of dosage regimens and/or in providing a better under-
`standing of observed drug reactions or interactions that result
`from the accumulation of drug in some specific site, tissue, or
`compartment of the body. The basis for the conclusion that two
`drug products are bioequivalent must be that the drug concen-
`trations measured in a biological matrix, or alternatively the
`pharmacological response, for one drug product are essentially
`the same for the second drug product. The more straightfor-
`ward decisions in the evaluation of bioequivalence between two
`drug products are those in which the two products are exactly
`superimposable (definitely bioequivalent). Those in which the
`two products differ in their bioequivalence parameters by a
`large amount, such as SO% or more, are most definitely not bio-
`equivalent. Statistical evaluation of the data is necessary for all
`situations, particularly for data that exist between these two
`extremes.
`
`METHODS FOR DETERMINING BIOEQUIVALENCE
`Bioequivalence usually involves human testing but sometimes
`may be demonstrated using an in vitro bioequivalence stan-
`dard, especially when such an in vitro test has been correlated
`with human in vivo bioavailability data. In other situations,
`bioequivalence may be demonstrated through comparative
`clinical trials or pharmacodynamic studies.
`The FDA has categorized (21CFR320.24) various in vivo and
`in vitro approaches that may be utilized to establish bioequiva-
`lence. These are, in descending order of accuracy, sensitivity
`and reproducibility,
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`PHARMACEUTICAL ANALYSIS AND QUALITY CONTROL
`1. An in 'Vivo test in humans in which the active drug sub-
`stance, as well as active metabolites when appropriate, is
`measured in plasma.
`2. An in vitro test that has been correlated with human
`in vivo bioavailability data. This approach is most likely
`for oral modified release products and is described in
`detail in FDA Guidance.
`3. An in vivo test in animals that has been correlated with
`human bioavailability data.
`4. An in vivo test in humans, where urinary excretion of
`the active drug substance, as well as active metabolites
`when appropriate, is measured.
`5. An in vioo test in humans in which an appropriate acute
`pharmacological effect is measured.
`6. Well-controlled clinical trials in humans that establish
`the safety and efficacy of the drug product, for estab-
`lishing bioavailability. For bioequivalence, comparative
`clinical trials may be considered. This approach is the
`least accurate, sensitive, and reproducible approach and
`should be considered only if other approaches are not
`feasible.
`7. A currently available in 'Vitro test, acceptable to FDA,
`that ensures bioavailability. This approach is intended
`only when in 'Vitro testing is deemed adequate, but no
`in vitro-in v ivo correlation (IVIVC) has been estab-
`lished. It also can relate to considerations involving the
`Biopharmaceutics Classification System (BCS).
`
`Most bioequivalence studies involve the direct measurement
`of the parent drug, as described in item 1 above. Bioequiva-
`lence testing in animals is not a recommended approach due to
`possible differences in metabolism, gastrointestinal physiology,
`weight, and diet.
`
`MINIMIZING THE NEED FOR BIOEQUIVALENCE
`STUDIES
`If a drug product has been adequ ately tested and approved for
`marketing, and if no changes in the manufacturing of the prod-
`uct are made, it is reasonable to assume that all subsequent
`batches of the product would be expected to be bioequivalent
`to the original product. If subsequently manufactured batches
`meet all tests of quality, including the dissolution test, no fur-
`ther human bioequivalence testing is needed.
`Depending on the degree of change, bioequivalence may
`sometimes need to be reconfirmed. Although it is somewhat
`difficult to categorize such major changes, this issue has been
`described in a series of FDA guidance documents related to
`Scale-Up and Post-Approval Changes (SUPAC).
`Drug characteristics related to solubility and permeability
`may allow a reasonable expectation that the drug is unlikely
`to be subject to significant bioavailability problems. For such
`drugs, in 'Vitro dissolution testing may be adequate, in lieu of
`in 'Vivo testing. These concepts are described in the Biophar-
`maceutics Classification System (BCS). This classification sys-
`tem provides a scientific framework for classifying drugs based
`on aqueous solubility and intestinal permeability. In addition,
`criteria for rapid dissolution are described (not less than 85%
`dissolved in 30 minutes, using mild agitation and physiologi-
`cal media). The BCS permits waivers of in 'V'ivo bioequivalence
`testing for high solubility, high permeability drugs (Class I),
`which are formulated into immediate release dosage forms hav-
`ing rapid dissolution. The basic ten et behind the BCS is that
`solutions of drugs are thought to have few bioavailability or bio-
`equivalence issues. Dosage forms containing drugs that are of
`high solubility and exhibit rapid dissolution behave similarly
`to a solution. Particularly for such drugs that are, in addition,
`highly permeable (well absorbed), the likelihood of bioavailabil-
`ity problems is quite small, and consequently, bioequivalence
`testing for such drugs is thought to be unnecessary. Similarly
`for oral solutions, bioequivalence testing is not necessary.
`
`EVALUATION OF BIOEQUIVALENCE DATA
`The following sections highlight some considerations when
`evaluating data from bioequivalence studies. The topics dis-
`cussed are directed specifically toward plasma level evaluations.
`With minor modifications, the approaches outlined can be used
`for urinary excretion measurements or for suitable, quantita-
`tive, pharmacological response measurements.
`Bioequivalence studies are usually conducted in healthy
`adults under standardized conditions. Most often, single doses
`of the test and reference product will be evaluated. However,
`in selected cases, multiple-dose regimens may be used (e.g. ,
`when patients are used and they cann ot be discontinued from
`a medication). The goal of the study is to evaluate the in 'V'i'VO
`performance, as measured by rate and extent of absorption, of
`the dosage forms under standardized conditions to minimize
`patient-related and other variability.
`The protocol should define the acceptable age and weight
`range for the subjects to be included in the study, as well as the
`clinical parameters that will be used to characterize a healthy
`adult (e.g., physical examination observations , clinical chemis-
`try, and hematological evaluations). The subjects should have
`been drug-free for at least two weeks prior to testing to elimi-
`nate possible drug-induced influences on liver enzyme systems.
`Normally, the subjects will fast overnight for at least ten hours
`prior to dosing and will not eat until a standard meal is provided
`four hours post-dosing. The dosage forms should be given to
`subjects in a randomized manner, using a suitable crossover
`design, so that possible daily variations are distributed equally
`between the dosage forms tested. The protocol should define
`sample collection times and techniques to collect the biologi-
`cal fluid. The method of sample storage should also be defined.
`
`BIOEQUIVALENCE ASSESSMENT
`AND DATA EVALUATION
`Several parameters are used to provide a general evaluation of
`the overall rate and extent of absorption of a drug. An analy-
`sis of all characteristics is required before one can determine
`bioequivalence or lack of bioequivalence. It is implicit that the
`analytical methodology used for analysis of drug in the samples
`is specific, sensitive, and precise.
`In assessing the bioequivalence of drug products, one must
`quantitate the rate an d extent of absorption, which can be
`determined by evaluating parameters derived from the blood-
`level concentration-time profile. Three parameters describing
`a blood-level curve are considered important in evaluating the
`bioequivalence of two or more formulations of the same drug.
`These are the peak-height concentration (Cmax), the time of the
`peak concentration (Tm:uJ, and the area under the blood (se-
`rum or plasma) concentration-time curve (AUC).
`
`PEAK-HEIGHT CONCENTRATION (Cmax)
`The peak of the blood-level-time curve represents the highest
`drug concentration achieved after oral administration. It is re-
`ported as an amount per volume measurement (e.g., microgram/
`milliliter (pgtmL) , unit/mL, or gram/100 mL). The importance
`of this parameter is illustrated in Figure 18-5, wh ere tl1e blood
`concentration-time curves of two different formulations of a
`drug are represented. A line has been drawn across the curve
`at 4 pg.'mL. Suppose that the drug is an analgesic, and 4 pg.'mL
`is the minimum effective concentration (!vlEC) of the drug in
`blood. If the blood concentration curves in Figure 18-5 rep-
`resent the blood levels obtained after administration of equal
`doses of two formulations of the drug and it is known that an-
`algesia would not be produced unless the MEC was achieved
`or exceeded, it becomes clear that formulation A would be ex-
`pected to provide pain relief, while formulation B, even though
`it is well absorbed regarding extent of absorption, might be inef-
`fective in producing analgesia.
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`/Peak
`
`..
`
`::::;
`-®,
`.=;,6.0
`c:
`0
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`~ 13 4.0
`0 c:
`0
`0
`u g 2.0
`:0
`Q)
`Ol
`~
`Q)
`
`~ 0
`
`8
`6
`16
`14
`12
`10
`2 3 4
`Time after drug administraion (hours)
`Figure 18-5. Blood concentration-time curves obtained for two dif-
`ferent formulations of the same drug, demonstrating relationship of
`the profiles to the minimum effective concentration (MEC).
`
`20
`
`-
`
`-
`
`AREA
`(0-20 hours)
`.
`Jlg
`A= 34.4 ml x hours
`Jlg
`A-·-t:> A = 34.2 ml x hours
`Formulation A
`·- ·- ·- ·-·-·-·-·-·-·-·- ·- ·- ·- ·- ·- ·MTC
`-----.:.
`........ --~:>
`-,-Formulation B
`:" ·':c...._
`--- -- ~~-- - - · - · -· -· - ·- ·----MEC
`
`·-6.--
`
`·-e... _ ___ _
`.
`
`::::;
`.E
`E:6.0
`c: g
`~ 13 4.0
`0 c:
`0
`0
`u g 2.0
`:0
`Q)
`Ol
`~
`Q)
`~
`
`0 1 2 3 4
`8
`6
`16
`14
`12
`10
`Time after drug administraion (hours)
`Figure 18-6. Blood concentration-time curves obtained for two dif-
`ferent formulations of the same drug, demonstrating relationship of
`the profiles to the minimum toxic concentration (MTC) and the mini-
`mum effective concentration (MEC).
`
`20
`
`On the other hand, if the two curves represent blood con-
`centrations following equal doses of two different formulations
`of the same cardiac glycoside, and 4 j.ig/mL now represents the
`minimum toxic concentration (MTC) and 2 jlg/mL represents
`the MEC (Figure 18-6), formulation A, although effective, may
`also present safety concerns, while formulation B produces con-
`centrations well above the MEC but never reaches toxic levels.
`Time of Peak Concentration (T max)
`The second parameter of importance is the measurement of the
`length of time necessary to achieve the maximum concentration
`
`353
`BIOAVAILABILITY AND BIOEOUIVALENCE TESTING
`after drug administration. This parameter is called the time of
`peak blood concentration (T013J. In Figure 18-S, for formula-
`tion A the time necessary to achieve peak blood concentration
`is 1 h. For formulation B, Tmax is 4 h. This parameter is related
`closely to the rate of absorption of the drug from a formulation
`and may be used as a simple measure of rate of absorption bu t
`is normally not evaluated statistically.
`To illustrate the importance of Tmax> suppose that the two
`curves in Figure 18-6 now represent two formulations of an an-
`algesic and that in this case the MEC is 2.0gtmL. Formulation
`A will achieve the MEC in 30 minutes; formulation B does not
`achieve that concentration until 2 h. Formulation A would pro-
`duce analgesia much more rapidly than formulation B and would
`probably be preferable as an analgesic agent. On the other hand,
`if one were more interested in the duration of the analgesic ef-
`fect than on the tin1e of onset, formulation B would present more
`prolonged activity, maintaining serum concentrations above the
`MEC for a longer time (8 h) than formulation A (5.5 h).
`AREA UNDER THE CONCENTRATION-TIME
`CURVE (AUC)
`The third, and sometimes the most important parameter for
`evaluation, is the area under the serum, blood, or plasma con-
`centration-time curve (AUC). This area is reported in amount/
`volume multiplied by time (e.g., jlg/mL x h or g/100 mL x h)
`and can be considered representative of the amount of drug
`absorbed following administration of a single dose of the drug.
`Although several methods exist for calculating the AUC, the
`trapezoidal rule method is most often used. This method as-
`sumes a linear function, y = bt + a, and its accuracy increases
`as the number of appropriate sampling intervals are increased.
`Table 18-1 and Figu re 18-7 describe the process for calculating
`the AUC using the trapezoidal rule.
`Returning to Figure 18-6, the curves, although much differ-
`ent in shape, have approximately the same areas (A = 34.4 jlg/
`mL x h ; B = 34.2 j.ig/mL x h), and both formulations can be
`considered to deliver the same amount of drug to the systemic
`circulation. Thus, one can see that AUC should not represent
`the only criterion on which bioequivalence is judged. All there-
`sults, as a composite, must be considered in reaching a decision
`about bioequivalence since no single parameter is adequate to
`serve this purpose.
`The plasma concentration- time curve is the focal point of
`bioequivalence assessment and is obtained when serial blood
`samples are analyzed for drug concentration. The concentra-
`tions are plotted on the ordinate (y-axis), and the times after
`drug administration that the samples were obtained are plotted
`on the abscissa (x-axis).
`A drug product is administered orally at time zero, and the
`plasma drug concentration at this time clearly should be zero.
`As a product passes through the gastrointestinal (GI) tract, it
`must u ndergo a sequence of events depicted in Figure 18-1. As
`
`AUC(o-E;) is used for bioequivalence analyses when the AUC(O·I) makes up ~ 80% of the AUC(o- £;)· AUC(O-I) is used when the AUC(O·I)
`makes up < 80% of the AUC(o-E;)· When drugs with long half-lives are evaluated, AUC(o-r) may sometimes be used with a truncated
`time point.
`Area under the concentration-time curve from time zero to time t (AUCo.t)
`1. Plot the concentration-time data for each subject
`2. Divide the curve into trapezoids by drawing vertical lines from each datum point to the x -axis. Calculate the area of the trapezoids
`using the following formula:
`3. AUC(t2-t1l = ((C2 + C1)(t2 - t1)] I 2
`AUC(o-t) is then calculated by summing the individual area.s to the time of the last concentration:
`4. AUC(o-t) = AUC(t2·l1) + AUC(t3-t2) + ... + AUC(In-(ln-1))
`Area under the concentration-time curve from time zero to infinity (AUCo-E)
`5. To calculate AU~(O-E;)• the tail region of the c urve must be added to AUC(O-I): AUC(o-E;) = AUC(O·Il + AUC "tail"
`6. AUC "tail" = C1!"-z_, where: C1 is the last detectable c oncentration, and "-._ is the terminal elimination rate const ant (see Figure 18-9).
`
`AMN1067
`Amneal Pharmaceuticals LLC v. Alkermes Pharma Ireland Limited
`IPR2018-00943
`
`

`

`354
`
`PHARMACEUTICAL ANALYSIS AND QUALITY CONTROL
`AUC0
`
`_
`
`c
`.Q
`~ <:
`~ c
`
`0
`(.)
`
`Sampling time
`Figure 18-7. Graphical depiction of use of the trapezoidal rule for
`calculating the area under the concentration-time curve.
`
`the drug is absorbed, increasing concentrations of the drug are
`observed in successive samples until the maximum concentra-
`tion is achieved. This point of maximum concentration (C,wx)
`is the peak of the concentration-time curve. If a simple model
`describes the pharmacokinetics of the drug tested, the peak
`concentration represents approximately the point in time when
`absorption and elimination of the drug have equalized.
`The section of the curve to the left of the peak represents
`the absorption phase (or absorption and distribution), during
`which absorption predominates over elimination. The section
`of the curve to the right of the peak is called the elimination
`phase, during which el