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Quality assessment of fluconazole capsules and oral suspensions compounded by pharmacies located in the United States OBJECTIVE To evaluate pharmaceutical characteristics (strength or concentration, accuracy, and precision), physical properties, and bacterial contamination of fluconazole compounded products.

Carine M. Laporte vmd Crisanta Cruz-Espindola bs Kamoltip Thungrat dvm, phd Anthea E. Schick dvm

SAMPLE Fluconazole compounded products (30- and 240-mg capsules; 30- and 100mg/mL oral suspensions) from 4 US veterinary compounding pharmacies.

Thomas P. Lewis II dvm Dawn M. Boothe dvm, phd Received January 15, 2016. Accepted July 14, 2016. From Dermatology for Animals, 86 W Juniper St, Gilbert, AZ 85233 (Laporte, Schick, Lewis); the Department of Anatomy, Physiology, and Pharmacology, College of Veterinary Medicine, Auburn University, Auburn, AL 36849 (Cruz-Espindola, Boothe); and the Department of Pharmacology, Faculty of Veterinary Science, Chulalongkorn University, Bangkok 10330, Thailand (Thungrat). Address correspondence to Dr. Laporte (laporte. [email protected]).

PROCEDURES Fluconazole compounded products were ordered 3 times from each of 4 pharmacies at 7- or 10-day intervals. Generic fluconazole products (50- and 200-mg tablets; 10- and 40-mg/mL oral suspensions) served as references. Compounded products were evaluated at the time of receipt; suspensions also were evaluated 3 months later and at beyond-use dates. Evaluations included assessments of strength (concentration), accuracy, precision, physical properties, and bacterial contamination. Acceptable accuracy was defined as within ± 10% of the labeled strength (concentration) and acceptable precision as within ± 10%. Fluconazole was quantified by use of highperformance liquid chromatography. RESULTS Physical characteristics of compounded products differed among pharmacies. Aerobic bacterial cultures yielded negative results. Capsules (30 and 240 mg) had acceptable accuracy (median, 96.3%; range, 87.3% to 135.2%) and precision (mean ± SD, 7.4 ± 6.0%). Suspensions (30 and 100 mg/mL) had poor accuracy (median, 73.8%; range, 53.9% to 95.2%) and precision (mean ± SD, 15.0 ± 6.9%). Accuracy and precision were significantly better for capsules than for suspensions. CONCLUSIONS AND CLINICAL RELEVANCE Fluconazole compounded products, particularly suspensions, differed in pharmaceutical and physical qualities. Studies to evaluate the impact of inconsistent quality on bioavailability or clinical efficacy of compounded fluconazole products are indicated, and each study should include data on the quality of the compounded product evaluated. (Am J Vet Res 2017;78:421– 432)

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ompounding, as defined by the National Association of Boards of Pharmacy, is the preparation, mixing, assembling, packaging, or labeling of a drug or device as the result of a practitioner’s order or initiative within the practitioner-patient-pharmacist relationship.1 The AMDUCA of 1994 describes the legitimate use of compounded veterinary medications as one in which no approved animal or human drug will, in the available dosage form and concentration, appropriately treat the diagnosed condition when used as labeled.2 Compounding of drug products has historically played an important role in ABBREVIATIONS BUD Beyond-use date HPLC High-performance liquid chromatography RSD Relative SD USP US Pharmacopeia

the treatment of veterinary patients.3 Advantages of compounded products include the ability to optimize veterinary treatment protocols by improving drug palatability or by adjusting drug vehicle or inactive components for patients that have had adverse effects when treated with FDA-approved products. Importantly, compounded products can be used to enhance dosage accuracy and safety by tailoring drug strengths or concentrations for administration to extremely large or small animals. However, the reformulation of any drug or bulk substance into a compounded product can alter its pharmacological properties and biological effects, which potentially places animals at risk from toxic events or from inappropriate or ineffective treatment. The FDA-approved products include pioneer (brand name) and generic drugs. Approval by the

AJVR • Vol 78 • No. 4 • April 2017

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FDA of any drug product verifies certain pharmaceutical characteristics, including strength or concentration (amount of active ingredient per unit of drug weight), accuracy (difference between actual and predicted strengths or concentrations), and precision (variability around the mean and coefficient of variation). In addition, FDA approval verifies certain physical characteristics of a drug, which differ according to drug formulation. Examples of regulated physical characteristics include size of particles, dispersion of particles, and ease of redispersion of particles in suspensions; size, shape, and weight of solid oral administration formulations4; pH; specific gravity; and microbiological quality. Physical characteristics of a drug may impact a number of patient factors for drug administration (eg, drug absorption, dissolution, and bioavailability) as well as accuracy and precision of the drug product. In contrast to approved drug formulations, compounded products are not regulated by the FDA, although they are considered by the FDA to be new (unapproved) drugs. Instead, the act of compounding (but not the compounded products themselves) is regulated at the state level through state Boards of Pharmacy, with regulations differing between states. Among the guidelines to which many state Boards of Pharmacy require that compounding pharmacists adhere are those generated by the USP, particularly those delineated in 3 chapters of the USP National Formulary.5–7 Compounded products are not subjected to the same rigorous predispensing quality assessments as are FDA-approved products; therefore, the drug dosage, composition, physical appearance, and amount of bacterial contamination cannot be assured in any compounded product. This has resulted in substantial pharmacological or treatment (or both) discrepancies between compounded products and FDA-approved products in veterinary medicine.8–13 Fluconazole is available as FDA-approved products (pioneer and generic) and as compounded products. Used in veterinary medicine for the treatment of numerous cutaneous and noncutaneous fungal infections, fluconazole may have several advantages against susceptible infections, including in vitro and in vivo efficacies against many fungi, high oral bioavailability (> 90%), low protein binding (approx 10%), intestinal absorption unaffected by food or pH, good tissue penetration, and fewer adverse effects than for ketoconazole or itraconazole.14–27 Fluconazole also increases serum cyclosporine concentrations, which potentially can reduce the cost of immunosuppressive treatments.28,29 However, there currently is no approved veterinary formulation of fluconazole. Difficulties with administration, palatability, or accurate dosing of veterinary patients by use of formulations approved for use in humans have led some veterinarians to seek pharmaceutical compounded products as an alternative. A recent study30 revealed variable quality of fluconazole compounded capsules intended for human use in Brazil. However, 422

studies of fluconazole compounded products, fluconazole compounded products produced by US pharmacies, and other fluconazole compounded preparations for veterinary use are lacking. Therefore, there is a need for in-depth analyses of fluconazole compounded products to ensure quality control of prescribed veterinary treatments. The purpose of the study reported here was to evaluate the quality of fluconazole compounded products (capsules and oral suspensions) obtained from veterinary pharmacies in the United States. Specific objectives were to evaluate physical characteristics of the fluconazole compounded products; determine the strength or concentration, accuracy, and precision of the compounded products, including comparisons across time; and quantify bacterial contamination of the compounded oral suspensions across time. We hypothesized there would be significant variability in the quality of the fluconazole compounded products.

Materials and Methods Samples

An internet-based search was conducted, which identified 4 pharmacies (pharmacy 1,a 2,b 3,c and 4d) in the United States that marketed fluconazole compounded products with nationwide distribution. Replicate orders for fluconazole compounded products were placed with each of the 4 pharmacies at 7- to 10-day intervals (3 orders/pharmacy; 12 orders total). Orders were designated as 1, 2, and 3, which corresponded with the sequential dates on which the orders were received. For example, for each pharmacy, order 1 was received on day 0, order 2 was received on day 7 to 10, and order 3 was received on day 14 to 20. Each order included 20 each of 30- and 240mg compounded capsules and 45 mL each of 30- and 100-mg/mL compounded oral suspensions. Strengths or concentrations were ordered from the pharmacies to simulate common clinical use (ie, for large and small animals) and differed from those of available approved products. Pharmacies were not made aware of the study to avoid bias in their compounding practices and procedures.

Experimental design

Each pharmacy was contacted via telephone. Before an order was placed, each pharmacy was asked to indicate verbally the BUD for each compounded product. All compounded oral suspensions in this study had a 6-month BUD, whereas the BUD for compounded capsules differed (range, 1 to 2 years). The BUD of a compounded product was confirmed at the time of receipt of the product, provided the BUD was listed on the package label. Samples were stored in accordance with label instructions until the BUD or end of the study, whichever came first. If no instructions for storage were provided, samples were stored on a shelf at controlled room temperature (22° to 23°C) away from direct sunlight and excessive heat and cold.31


All compounded products were subjected to pharmaceutical and physical analyses. Physical analyses of samples were performed at Dermatology for Animals, and pharmaceutical analyses of samples were performed at the Department of Anatomy, Physiology, and Pharmacology of the College of Veterinary Medicine at Auburn University. The investigators who performed the analyses (CML, CCE, and DMB) were not aware of the pharmacy origin of each compounded product. For compounded suspensions, analysis was performed at 3 time points: the initial time point (defined as within 1 week after receipt of the order), 3 months (± 1 week) after receipt, and at the BUD (± 1 week). Additionally, aerobic bacterial cultures were performed on suspensions at the initial and BUD analyses. For compounded capsules, analysis was limited only to the initial time point for 2 reasons. First, according to the USP, there is a greater risk of loss of stability for suspensions containing water (including all suspensions used in the present study).6 Second, the BUD for all compounded product capsules extended beyond the study period, which prohibited BUD analyses. Thus, the authors elected to focus primarily on analysis of the compounded suspensions. Two approaches were used for control samples. Approved generic fluconazole products (tablets, 50e and 200 mgf; suspensions, 10g and 40 mg/mLg) were evaluated. Furthermore, vehicles (a colloidal agents–blended vehicleh and a food starch–based vehiclei) used by 2 of the 4 pharmacies were obtained, and pure fluconazole powderj was added to create positive control solutions of known concentrations (30 and 100 mg/mL). For analysis of the generic tablets, it was not possible to blind the investigators because of differences in physical characteristics of the 2 doses of the generic tablets and the fact that compounded products were provided in capsules rather than tablets. Generic fluconazole samples and positive control solutions were evaluated with the compounded products.

Sample analysis

Assessment of physical characteristics—All product labels and packages were assessed and compared with FDA guidelines and recommendations.32,33 Capsule physical characteristics were evaluated, including capsule and content appearance, capsule and content weight, and presence of defects.4,34 Ten compounded capsules from each order were weighed in their entirety (capsule plus contents), and mean ± SD weight of the 10 capsules was reported (designated as capsule weight).5 Capsules were then emptied and the contents weighed, which also was reported as mean ± SD values (designated as content weight). Capsules were excluded from further physical analysis if they could not be completely emptied of contents. Physical characteristics of oral suspensions were recorded for each concentration from each pharmacy. Suspension characteristics evaluated were color, clarity, particle size, globule size, specific gravity, pH, and extent of sedimentation after 24 hours without

agitation.5,35 Particle and globule size of compounded suspensions were assessed by use of scales of 1 to 5 and 1 to 8, respectively, as described elsewhere.13 The pH was measured with a test strip,k and specific gravity was measured by use of a pycnometer.l Extent of sedimentation in a 2-mL volume of suspension was measured with a ruler and expressed as a percentage of the total volume.35 When large particulate matter in compounded suspensions obstructed the pipette tip during sampling, the matter was dislodged and additional samples were obtained from the suspensions until the desired volume was aspirated. Mean ± SD values were reported for specific gravity, sedimentation, and globule size of each suspension preparation (2 concentrations) for all orders. Evaluation of product strength or concentration, accuracy, and precision—Analyses (2 capsules and 2 aliquots of suspension [100 µL/aliquot]) were performed for each order from each pharmacy. When physical defects were detected in the capsules of a particular order, analyses were performed with 1 capsule without defects and 1 capsule with defects. Similar to the procedures for physical characteristics, capsules were excluded from measurement if they could not be completely emptied of contents. Fluconazole was quantitated in capsules and oral suspension samples by use of HPLC with UV detection. The method was a modification of a protocol described elsewhere.36,37 The HPLC system consisted of a separation modulem and a dual λ absorbance detector.n Separation was achieved with a 5-µm, 4.6 X 150-mm chromatographic columno protected by a 5-µm, 4.6 X 20-mm guard cartridge.p The mobile phase consisted of 0.02M monobasic potassium phosphate buffer (pH, 2.5) and acetonitrileq (80:20 [vol/ vol]). Flow rate was 1.5 mL/min.36,37 The standard curve for capsules and oral suspensions was generated (range of concentrations, 25 to 200 µg/mL) by fortifying diluent (methanol and distilled water [50:50 {vol/vol}]) with known amounts of fluconazolej reference standard.36,37 Samples were monitored at 260 nm.36,37 For capsules, the powder of a previously weighed capsule was transferred to a volumetric flask (100 µL), and 50 µL of diluent was added. The solution was mixed in a vortex device and then sonicated for 15 minutes. The solution was allowed to equilibrate to room temperature, and the volumetric flask was filled to the mark with the diluent. A portion of the solution was centrifuged at 1,900 X g for 10 minutes at room temperature. A portion of the centrifuged solution was diluted to obtain a final solution with a concentration of 100 µg of fluconazole/mL.36 An aliquot (50 µL) of the final solution was injected into the HPLC system. Solutions were assayed in duplicate, and the mean of the duplicates was calculated. For oral suspensions, the suspension was manually shaken for 1 minute. An aliquot (100 µL) was then transferred to a glass tube (6 mL [30-mg/mL suspensions] or 10 mL [100-mg/mL suspensions]).


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Diluent (2,900 mL [30-mg/mL suspensions] or 9,900 mL [100-mg/mL suspensions]) was added. The solution was mixed in a vortex device and then sonicated for 20 minutes with intermittent shaking.37 The solution was centrifuged at 1,900 X g for 10 minutes at room temperature. A portion of the centrifuged solution was diluted to obtain a final solution with a concentration of 100 µg of fluconazole/mL.37 An aliquot (50 µL) of the final solution was injected into the HPLC system. Products were assayed in duplicate, and the mean of the duplicates was recorded as the concentration for that formulation. The HPLC assay was validated by use of the solution used to dilute the preparations. Fluconazole was quantitated by comparison of signals from unknowns (capsules and suspensions) with signals in a calibration curve generated by use of fluconazole in the diluent. The calibration curve ranged from 25 to 200 µg/mL, and precision and accuracy were based on results for 3 control samples (25, 100, and 200 µg/mL) that spanned the breadth of the calibration curve. The linear correlation coefficient for the fluconazole calibration curve was 0.999. Upper and lower limits of quantification of the curve, which were based on ± 10% for accuracy and precision, were 200 and 25 µg/ mL, respectively. The limit of detection was 10 µg/ mL. For the fluconazole assay, accuracy and precision of the fluconazole content in the methanol–distilled water diluent were a mean ± SD of 99.81 ± 0.5% and mean of 0.34%, respectively. Strength or concentration, precision, and accuracy were determined for each fluconazole compounded product formulation (capsule or oral suspension; 2 preparations/formulation). Strength or concentration was a measure of the amount of active pharmaceutical ingredient (fluconazole) and was expressed on the basis of the amount in each unit (ie, per capsule or per milliliter of suspension). Precision was a measure of repeatability (ie, degree of agreement in preparation strength or concentration among units [capsules or milliliters of suspension]). Precision was expressed as the percentage RSD (ie, coefficient of variation) by use of the following equation: – X 100 Percentage RSD = σ⁄ x – is the arithmetic mean. Acwhere σ is the SD and x ceptable precision was defined as within ± 10%. Accuracy (a measure of the closeness of the actual preparation strength or concentration, compared with the label strength or concentration) was expressed as percentage recovery by use of the following equation: percentage recovery = (measured amount/label amount) X 100. Acceptable accuracy was defined as within ± 10% of the label concentration. Bacterial culture of suspensions—According to the USP guidelines for fluconazole oral suspensions,37 total aerobic microbial count for fluconazole suspensions should not exceed 102 CFUs/g, and samples should be free of Escherichia coli. Each oral suspension was mixed for 30 seconds and visually assessed; a 1-mL aliquot then was placed onto each of 2 tryp424

tic soy agar plates.q A negative control plate as well as a positive control plate with an E coli strain were included in each incubation. Plates were incubated at 37°C in 5% CO2; they were visually inspected for growth at 24 and 48 hours.7,38

Statistical analysis

Distributions for strength or concentration, accuracy, and precision data were tested for normality by use of the Shapiro-Wilk test. Strength or concentration and accuracy data had a right-skewed distribution (P = 0.01) and unequal variances among pharmacies (Levene test; P = 0.01). Therefore, accuracy and strength or concentration were reported as median and range (ie, minimum to maximum), and precision was reported as mean ± SD and 95% confidence interval. Strength or concentration and accuracy were reported for each label strength or concentration, each order, and each pharmacy. Concentration was also reported for each suspension (mean of all orders) for each pharmacy at initial, 3-month, and BUD analyses. Strength or concentration, accuracy, and precision then were reported by formulation (mean of all orders) for all pharmacies, and overall accuracy and precision were determined for capsules and suspensions. For inferential statistics, nonparametric procedures were used to detect significant differences for strength or concentration and accuracy data. The Wilcoxon 2-sample test was used to compare overall accuracy between capsules and suspensions for the analysis. The Kruskal-Wallis test was used to compare accuracy for each formulation among pharmacies and each pharmacy among orders. These analyses were limited to the initial time point with all orders combined. Strength or concentration of each formulation for each pharmacy was compared among times (initial, 3-month, and BUD analyses) by use of the Kruskal-Wallis test. A multiple comparisons post hoc test for a Kruskal-Wallis analysis was used to identify significant differences among > 2 groups.39 A Student 2-tailed t test was used to compare precision between capsules and suspensions at the initial time point for all pharmacies combined. All analyses were conducted by use of a commercial software program r; values were considered significant at P < 0.05.

Results Labels

All labels of fluconazole compounded products contained information about the active ingredient, dosage strength or concentration, quantity or volume dispensed, prescriber and pharmacy, pharmacist initials, date the drug was compounded, prescription number, special storage and handling instructions, and directions for use. The dosage formulation was listed on all labels. Labels of 3 pharmacies indicated it as capsule, and 1 pharmacy indicated it as cap. Similarly, 3 pharmacies indicated it as suspension, and 1 pharmacy indicated it as liquid. However, contrary to


USP or FDA recommendations, several items were not provided on the labels; these items included lot or batch number (n = 2 pharmacies), the statement “not for resale” (3 pharmacies), the statement “for use only in [species or disease condition]” (4 pharmacies), and a statement about reporting of adverse events (4 pharmacies). All labels stated that the drug was compounded, but only 1 pharmacy stated this clearly on the label. For the remaining pharmacies, such information was included on a second label affixed out of the field of vision (a user had to turn the container to read the second label; 1 pharmacy), in a smaller font (1 pharmacy), or as an abbreviation (ie, cmpd; 1 pharmacy). Two pharmacies used the term “beyond use date” on their labels; the other 2 pharmacies referred to this information by use of the terms “expiry” or “discard after.” Compared with generic fluconazole, which must have distinct label or packaging identifiers, it was difficult to discriminate the strengths or concentrations of the various fluconazole compounded products on the basis of packaging or labeling. Reasons for this included look-alike labels and containers for differing dosages from a specific pharmacy, distribution of key label information over multiple fields of vision (ie, a user must turn the container to read the entire label), and labels that were cluttered and difficult to read because of font, text size, or space-occupying pharmacy logos.

Physical characteristics Control samples—The 50-mg generic tablets were trapezoidal (0.7 X 0.5 X 0.2 cm) and dark pink, whereas the 200-mg generic tablets were oval (length, 1.3 cm; diameter, 1.0 cm) and light pink. Mean ± SD

weight of the approved generic tablets (n = 10) was 111.2 ± 0.8 mg for the 50-mg tablets and 445.3 ± 4.6 mg for the 200-mg tablets. No physical defects of the generic tablets were visually observed. Fluconazole generic suspensions (10 and 40 mg/ mL) were provided as white uniform powders with separate vials that contained a standard 24 mL of filtered distilled water for reconstitution, which yielded a total volume of 35 mL. Suspensions were reconstituted in accordance with package directions, and suspensions of both concentrations were white, opaque, and homogenous. No sedimentation was observed at 24 hours, with both suspensions remaining white, opaque, and homogenous. Globule size was 4.0, pH was 4.0, and specific gravity was 1.226 g/mL for both concentrations at all time points. The labels indicated that both concentrations had orange flavor, and both had a citrus odor. Compounded capsules—Physical characteristics of the compounded capsules were summarized (Table 1). All fluconazole compounded capsules had hard shells. Products differed among pharmacies for a particular dose with regard to capsule color and size. Within a given pharmacy for a given dose, the same capsule type was used for all 3 orders. Each pharmacy used color or size, but never both, to differentiate the 30- and 240-mg doses. None of the capsules had an odor. Importantly, physical defects were detected in capsules from all pharmacies in at least 1 order (Figure 1). These defects included desiccated capsules that cracked when opened (9/24 [37.5%] orders; capsules from 3 pharmacies), an undefined white or light blue powder on the outside of the capsules (7/24 [29%] orders; capsules from 3 pharmacies), crushing of 1 end or both ends of the capsules (4/24 [17%] orders; capsules from 1 pharmacy), and loose

Table 1—Physical characteristics of compounded fluconazole capsules obtained for each of 3 orders from each of 4 pharmacies. Capsule

Length X Pharmacy diameter (cm)

Capsule color

Physical defects

30 mg 1 1.5 X 0.5 Clear 2 1.5 X 0.5 White 3 1.5 X 0.5 Clear with yellow hue 4 1.5 X 0.5 Half navy, half yellow

Capsule contents

Capsule weight Content (mg)* weight (mg)†

Powder on several capsules; Fine white powder 150.8 ± 31.5 130.3 ± 13.4 several loose capsules Several desiccated and Fine white powder 251.5 ± 17.8 203.9 ± 16.7 easily cracked capsules Crushing of several capsules; Fine white powder 227.6 ± 11.4 186.1 ± 8.4 several desiccated and easily cracked capsules White powder on capsules Granular white powder 258.9 ± 42.3 206.3 ± 36.0

240 mg 1 1.5 X 0.5 Cerulean White powder on capsules Fine white powder 303.2 ± 7.6 252.4 ± 13.8 2 2.1 X 0.7 White Several desiccated Fine white powder‡ 517.5 ± 13.9 431.7 ± 12.6 and easily cracked capsules 3 1.5 X 0.5 Half red, half turquoise Light blue or white powder Fine white powder 311.2 ± 6.0 253.9 ± 9.0 on several capsules; crushing of 1 or both ends of several capsules; several desiccated and easily cracked capsules 4 2.1 X 0.7 Half navy, half yellow White powder on capsules; Granular white powder 520.8 ± 64.9 426.6 ± 60.5 several desiccated and easily cracked capsules Orders were placed at each pharmacy at intervals of 7 to 10 days; thus, order 1 was received on day 0, order 2 was received on day 7 to 10, and order 3 was received on day 14 to 20. Compounded products were evaluated during the initial analysis, which was performed within 1 week after receipt of each order from the pharmacies. *Value reported is mean ± SD weight of 10 capsules and their contents for all 3 orders from each pharmacy. †Value reported is mean ± SD weight of only the contents of 10 capsules for all 3 orders from each pharmacy. ‡One capsule contained several dark-brown flecks.


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capsules that opened easily when manipulated (2/24 [8%] orders; capsules from 1 pharmacy). Contents of capsules were white powders, which ranged from a fine texture to a more granular texture. Several dark-brown flecks were intermixed in the contents of 1 capsule from pharmacy 2. For several pharmacies, variable amounts of the capsule contents were packed within the ends of the capsules and could not be removed, particularly when the capsules were damaged. Weights of capsule contents differed among pharmacies and also differed among orders within a given pharmacy. Overall, mean ± SD content weight of 30-mg capsules was 226.7 ± 42.4 mg, and mean content weight of 240-mg capsules was 413.2 ± 114.3 mg. Compounded suspensions—Physical characteristics of the compounded oral suspensions were summarized (Table 2). Compounded 100-mg/mL suspensions for orders 1 and 2 from pharmacy 4 were uniform with no visible particulate matter. In contrast, the 100-mg/mL suspension for order 3 from pharmacy 4, as well as all of the compounded suspensions for all 3 orders from the other 3 pharmacies, had visible, variably sized, and frequently large particulate matter. Particulate matter from 2 pharmacies (all

Figure 1—Photograph depicting examples of defects in compounded fluconazole capsules that were evident on receipt of the products. Notice the white powder on the external surface of the blue capsule and crushing (ie, indentation) of the end of the red capsule.

3 orders from pharmacy 1 and 1 order from pharmacy 2) obstructed the sampling pipette tips, with the 100-mg/mL suspensions causing more frequent and severe obstructions than did the 30-mg/mL suspensions. Manual agitation of the compounded products for 1 minute prior to sampling did not alter suspension particle size for any of the pharmacies. The pH was 4.5 for all compounded suspensions from 3 pharmacies. However, the pH of compounded suspensions from the other pharmacy was consistently more basic (pH, 6.5). Interestingly, testing of the 100-mg/ mL suspension in order 3 from pharmacy 4 revealed 2 pH values; the pH for the liquid component of the suspension (pH, 4.5) differed from that of a particle on the test strip (pH, 7.0). This phenomenon was not observed when the particulate matter contacted the pH test strip with any other compounded suspension. All suspensions of both concentrations had globule sizes between 4 and 5 on the globule scale.13 Overall mean ± SD specific gravity for the 30- and 100-mg/ mL compounded suspensions was 1.126 ± 0.115 g/mL and 1.149 ± 0.114 g/mL, respectively. Overall mean percentage sedimentation was 26 ± 40% and 15 ± 23% for the 30- and 100-mg/mL suspensions, respectively. Color, clarity, and pH of compounded suspensions were unchanged at the 3-month and BUD analyses. Particle size, specific gravity, and percentage sedimentation were variable at all time points, and no consistent pattern could be determined. For the 30- and 100-mg/mL suspensions, globule size (mean ± SD calculated for all orders and pharmacies) at the initial, 3-month, and BUD analyses were 4 ± 0.5, 5 ± 0.7, and 5 ± 0.8, respectively. When globule size was assessed by order or by pharmacy over time, globule size increased by 1 between the initial and 3-month analysis and between the 3-month and BUD analyses in 7 of 12 (30-mg/mL) and 8 of 12 (100-mg/mL) compounded suspensions.

Bacterial contamination

Corresponding with USP guidelines,37,38 neither the control samples nor any compounded sus-

Table 2—Physical characteristics of compounded fluconazole suspensions obtained for each of 3 orders from each of 4 pharmacies. Suspension Pharmacy

Color and clarity

Particle size*

30 mg/mL 1 White and opaque 1–5 2 White and opaque 1–4 3 White and opaque 1–3 4 White and opaque 1–3 100 mg/mL 1 White and opaque 1–5 2 White and opaque 1–3 3 White and opaque 1–3 4 White and opaque Homogenous for orders 1 and 2; 1–3 for order 3

Globule size† 4.0 4.3 ± 0.6 4.7 ± 0.6 4.3 ± 0.6 4.0 4.7 ± 0.6 4.0 4.7 ± 0.6

Specific gravity (g/mL)† pH† 1.044 ± 0.007 1.140 ± 0.005 1.299 ± 0.011 1.025 ± 0.005 1.060 ± 0.018 1.171 ± 0.016 1.316 ± 0.045 1.054 ± 0.031

4.5 4.5 6.5 4.5 4.5 4.5 6.5 4.5‡

Sedimentation (%)† 0 33.0 ± 57.2 58.7 ± 50.8 30.7 ± 49.7 0 11.0 ± 19.1 47.7 ± 21.2 22.0 ± 31.4

Compounded products were evaluated during the initial analysis, which was performed within 1 week after receipt of each order from the pharmacies. Particle size and globule size were assessed on a scale of 1 to 5 and 1 to 8, respectively, as described elsewhere.13 Sedimentation in a 2-mL volume of suspension was measured with a ruler and expressed as a percentage of the total volume. *Represents range of particle sizes visually observed in samples for all 3 orders from each pharmacy. †Represents mean ± SD for all 3 orders from each pharmacy. ‡The pH for the liquid component of the suspension for order 3 was 4.5, but a particle in that suspension had a pH of 7.0. See Table 1 for remainder of key.

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Figure 2—Actual (measured) strength of compounded fluconazole capsules (30 mg [A] and 240 mg [B]) obtained for each of 3 orders from each of 4 pharmacies (pharmacy 1 [circles], pharmacy 2 [squares], pharmacy 3 [triangles], and pharmacy 4 [inverted triangles], respectively); strength was measured during the initial analysis, which was performed within 1 week after receipt from the pharmacies. Orders were placed at each pharmacy at intervals of 7 to 10 days; thus, order 1 was received on day 0, order 2 was received on day 7 to 10, and order 3 was received on day 14 to 20. For both strengths (30 and 240 mg), 2 capsules in each order from each pharmacy were assayed, and the mean for the 2 capsules was recorded as the concentration. Generic fluconazole tablets (50 mg [A] and 200 mg [B]) served as control samples (diamonds). Assays were performed in duplicate, and each data point on the graph represents results for 1 assay. The horizontal dotted line represents the target concentration for the compounded capsules, and the dashed-and-dotted lines represent ± 10% of the target.

pensions had bacterial growth for initial or BUD analyses.

Strength or concentration, accuracy, and precision

Control samples—Fluconazole generic and positive control samples were evaluated along with the

Figure 3—Actual (measured) concentration of compounded fluconazole suspensions (30 mg/mL [A] and 100 mg/mL [B]) obtained for each of 3 orders from each of 4 pharmacies; concentration was measured during the initial analysis, which was performed within 1 week after receipt from the pharmacies. For both concentrations (30 and 100 mg/mL), 2 aliquots (100 µL/aliquot) for each order from each pharmacy were assayed, and the mean of the 2 aliquots was recorded as the concentration. Fluconazole powder in a food starch–based vehicle (30 mg/mL [A] and 100 mg/mL [B]) served as positive control samples (diamonds). Assays were performed in duplicate, and each data point represents results for 1 assay. See Figure 2 for remainder of key.

compounded products. Median strength for the fluconazole generic 50- and 200-mg tablets was 48 mg (range, 47.5 to 50.4 mg) and 195 mg (range, 192.6 to 197.5 mg), respectively. Accuracy and precision for each tablet size were within acceptable limits. Median concentration for fluconazole generic 10- and 40-mg/mL suspensions was 8.3 mg/mL (range, 8.0 to 8.6 mg/mL) and 31.2 mg/mL (range, 30.5 to 33.4 mg/ mL), respectively. Precision for each suspension was within acceptable limits. For the positive control sus-


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Table 3—Strength or concentration, accuracy, and precision for compounded fluconazole capsules and suspensions (2 concentrations of each) obtained for each of 3 orders from each of 4 pharmacies. Compounded product Pharmacy

Strength or concentration* Label Measured† Accuracy (%)†‡

Precision (%RSD)§

Capsule

1 2 3 4

30 30 30 30

37.0 (26.2–40.6) 29.2 (27.7–29.4) 28.3 (27.6–32.1) 27.1 (26.8–27.7)

123.2 (87.3–135.2) 97.2 (92.2–97.9) 94.3 (91.9–106.9) 90.2 (89.4–92.4)

22 4 9 4

Capsule

1 2 3 4

240 240 240 240

225.1 (212.0–243.0) 238.2 (231.7–242.9) 228.6 (217.1–245.6) 237.4 (227.3–237.4)

93.8 (88.3–101.3) 99.2 (96.6–101.2) 95.3 (90.4–102.3) 97.9 (94.7–98.9)

7 4 7 4

Suspension

1 2 3 4

30 30 30 30

21.4 (20.0–23.9) 23.5 (23.3–26.0) 16.7 (16.2–22.4) 27.2 (26.9–28.5)

71.5 (66.8–79.7) 78.3 (77.5–88.6) 55.7 (53.9–74.7) 90.8 (89.8–95.2)

15 3 24 17

Suspension

1 2 3 4

100 100 100 100

64.2 (62.1–67.9) 74.1 (71.3–77.9) 67.1 (65.8–71.0) 88.9 (86.9–90.6)

64.2 (62.1–67.9) 74.1 (71.3–77.9) 67.1 (65.8–71.0) 88.9 (86.9–90.6)

12 11 24 14

For each compounded product and label strength or concentration, 2 capsules or two 100-µL aliquots of suspension for each order from each pharmacy were assayed during the initial analysis to determine strength or concentration. Mean of the 2 capsules or aliquots was recorded as the strength or concentration for that assay; each assay was performed in duplicate. Thus, reported values represent results for 6 assays for all 3 orders from each pharmacy. *Value reported is milligrams for capsules and milligrams per milliliter for suspensions. †Value reported is median (range). ‡Acceptable accuracy was defined as 90% to 110% of label strength or concentration. §Acceptable precision was defined as within a range of 20% (ie, ± 10% of label strength or concentration). See Table 1 for remainder of key.

pensions created with the food starch–based vehicle,i accuracy was acceptable for both the 30-mg/mL suspensions (median, 98.3%; range, 97.8% to 98.8%) and the 100-mg/mL suspensions (median, 93.3%; range, 93.2% to 93.4%). Precision for both suspension concentrations was also acceptable (0.7% and 0.1% for the 30- and 100-mg/mL suspensions, respectively). For the same suspensions created by use of the colloidal agents–blended vehicle,h accuracy was poor for the 30-mg/mL suspensions (median, 88.1%; range, 86.2% to 90.0%) and the 100-mg/mL suspensions (median, 84.0%; range, 81.4% to 86.9%). Precision was acceptable (3.1% and 4.8% for the 30- and 100-mg/mL suspensions, respectively). Compounded capsules and suspensions—Actual (measured) strength or concentration was determined for each compounded product (capsule and suspension), label strength or concentration, and pharmacy by each order (Figures 2 and 3). Strength or concentration, accuracy, and precision of the compounded products were delineated for each pharmacy (all orders) on initial analysis (Table 3). Overall accuracy for capsules was acceptable. Although accuracy for the compounded capsules did not differ significantly among the pharmacies, accuracy for the 30-mg capsules from pharmacy 1 was outside of the acceptable limit. Precision (22%) for the 30-mg capsules from pharmacy 1 was also outside the acceptable limit. All compounded suspensions, except for 428

the 30-mg/mL suspension from pharmacy 4, were outside of acceptable limits for accuracy and precision. Accuracy for the suspensions did not differ among pharmacies. Concentration of suspensions did not differ significantly among the 3 time points (initial, 3-month, and BUD analyses; Figure 4). Regarding accuracy across orders, no differences could be detected for compounded capsules from any pharmacy, but there were differences across orders for suspensions (30 and 100 mg/mL) from 3 of the 4 pharmacies (accuracy increased significantly [P = 0.048] between orders 1 and 3 from pharmacy 1, accuracy increased significantly [P = 0.007] between orders 2 and 3 from pharmacy 3, and accuracy decreased significantly [P = 0.002] between orders 2 and 3 from pharmacy 4). Accuracy of compounded capsules consistently was significantly (P < 0.001) higher, compared with accuracy of compounded suspensions from each pharmacy. Similarly, precision (as indicated by the percentage RSD) for compounded capsules (mean ± SD, 7.4 ± 6.0%; 95% confidence interval, 2.4% to 12.5%) was significantly (P = 0.03) better, compared with the precision for compounded suspensions (mean ± SD, 15.0 ± 6.9%; 95% confidence interval, 9.2% to 20.7%).

Discussion Quality of compounded fluconazole capsules and oral suspensions obtained from 4 veterinary


Figure 4—Actual (measured) concentration and accuracy (percentage recovery) over time for compounded fluconazole suspensions (30 mg/mL [A] and 100 mg/mL [B]) obtained for each of 3 orders from each of 4 pharmacies. Concentration was measured during the initial analysis, 3 months after receipt from the pharmacies, and at the BUD. For each suspension (30 and 100 mg/mL), 2 aliquots (100 µL/aliquot) for each order from each pharmacy were assayed at each time point. Assays were performed in duplicate, and each data point represents the median (indicated by the symbol) and range of all assays for all orders from each pharmacy at a particular time point. See Figures 2 and 3 for remainder of key.

compounding pharmacies in the United States was evaluated in the study reported here. Results indicated marked variability in the pharmacological and physical qualities of these fluconazole compounded products. To the authors’ knowledge, this was the first study in which compounded fluconazole capsules and oral suspensions intended for veterinary patients were evaluated. Results of the present study are similar to those for a recent study30 on compounded fluconazole products for use in human medicine and suggest that caution is warranted in the use of compounded fluconazole products. Strength or concentration, accuracy, and precision of the compounded fluconazole products were compared among pharmacies and between formulations (capsule vs oral suspension). Accuracy and pre

cision of generic fluconazole tablets, generic 10-mg/ mL oral suspension, and positive control fluconazole suspensions were within acceptable limits, which verified that the detected deficiencies in accuracy or precision of the compounded products were not the result of analytic methods. Similar to results of a previous study,13 precision and accuracy for the study reported here were better for compounded capsules than for compounded suspensions. Overall, accuracy and precision of both the 30- and 240-mg compounded capsules were acceptable. However, it is important to mention that accuracy of 30-mg capsules from 1 pharmacy was outside acceptable limits, which indicated that quality of compounded capsules differed among pharmacies. Overall, accuracy and precision of compounded suspensions were poor and were outside acceptable limits for most of the suspensions of both concentrations (30 and 100 mg/mL). For all the compounded suspensions, concentration was less than the labeled concentration. Additionally, concentration of suspensions from 3 of the 4 pharmacies differed significantly among orders, which indicated variable quality, even in products from the same pharmacy. Taken together, these discrepancies would have the potential to result in drug dosage disparities, which may clinically translate into diminished or altered therapeutic efficacy, toxic events, or an overdose. Physical characteristics were variable among the compounded products. The most pronounced discrepancies in physical characteristics were for particle size and pH of the compounded suspensions and for physical integrity of the capsule shells and contents. Physical characteristics of drug products have the potential to influence a number of patient factors during oral administration of drugs, including transit of the drug through the gastrointestinal tract, dose uniformity, rates of dissolution and absorption, and bioavailability.40 Heterogeneous physical characteristics may further reflect a lack of standardization of manufacturing processes or a lack of stability. Generic fluconazole suspensions were consistently homogenous and uniform. In comparison, only one compounding pharmacy produced uniform suspensions; all other pharmacies produced suspensions with variably sized particulate matter. Two pharmacies produced suspensions with particulate matter that clogged the sampling pipette tips. This may have reflected a combination of factors, including particle size, particle shape, and interparticle interactions. Solid phases that cannot be redispersed in a suspension may be an indication of instability, and the presence of large particles may be reflective of excessive crystal growth within the suspension.31 Variability of particle size (ie, nonuniformity) within suspensions may also preclude reliable and accurate dosing in clinical settings. For example, a variable amount of drug may be contained within the particulate matter, which results in inaccurate and nonuniform concentrations of drug per volume of product administered;


429

particulate matter may obstruct the tip of the dosing syringe, which results in inconsistent volumes of suspension aspirated into the syringe; and particle size may impact solubility in the gastrointestinal tract and bioavailability of certain drugs. Variability in pH was detected among pharmacies and, for 1 sample, between the particulate and liquid components of the suspension. Compounded suspensions from pharmacy 3 were consistently more basic (pH, 6.5) than those of the other 3 pharmacies (pH, 4.5). One suspension from pharmacy 4 had a more basic pH of the particulate matter (pH, 7) than for the liquid component (pH, 4.5). The pH is an important pharmacological characteristic that impacts solubility and gastrointestinal tract absorption of orally administered drugs as well as drug stability.5,40 Despite substantial differences in gastrointestinal physiology between humans and various veterinary species, the criteria currently used for classifying solubility of veterinary drugs are based on human gastrointestinal physiology.40 According to USP guidelines, the pH of fluconazole oral suspensions intended for humans should be 3.0 to 5.0.37 Variation in physical characteristics may have contributed to the overall poor accuracy and precision of the compounded suspensions in the present study. Further studies are warranted to evaluate the effect of particle size and pH on the bioavailability and therapeutic efficacy of compounded fluconazole suspensions as well as the clinical impact of particle size on dosing accuracy of those compounded products. Numerous compounded capsules in the present study were damaged, and their contents were frequently irretrievably packed into the damaged shells. This was in contrast to the loose and powdery contents of most of the undamaged capsules. In a clinical setting, physical defects in drug capsule preparations may influence drug dosage, solubility, absorption, and bioavailability.40 It is possible that damage to the compounded capsules from this pharmacy resulted in a smaller capsule volume that could contain the drug during the compounding process or loss of drug after compounding. Despite this, the overall acceptable accuracy and precision for all capsules suggested that the physical defects did not substantially alter the active fluconazole content in those capsules. Package and label designs of the compounded products were also evaluated in the present study. Product labeling, packaging, and drug design are fundamental for conveying drug information to owners.32 An owner’s ability to find and interpret drug information is a critical factor in the prevention of medical errors in veterinary medicine. Medical errors are a major cause of human deaths in the United States and have resulted in a reported 44,000 to 98,000 deaths annually.41 Key components for drug labeling include information, nomenclature, and terminology.32 Labels of the compounded products in the present study did not provide all information as recommended by the FDA,32,33 and dosage strengths 430

from a given pharmacy were difficult to determine in part because of label design. Further, abbreviations or alternative terms were used on labels from the various pharmacies. For example, cmpd was used for compounded, cap was used for capsule, or liquid was used for suspension. Rather than use of the term BUD, 2 pharmacies referred to expiry or discard-after dates. A BUD indicates the time by which a pharmacist deems a compounded product must be used before it becomes at risk for microbial contamination, permeation of the packaging, or chemical degradation. Because compounded preparations are intended for immediate or short-term administration, BUDs are assigned on the basis of different criteria than those applied to determining expiry dates of manufactured drug products. Expiry dates are based on results of rigorous analytic and performance testing by a product manufacturer for specific factors, such as choice of drug container and exposure to illumination and temperature. In contrast, BUDs are assigned by individual pharmacies and are often based on professional experience and literature research rather than rigorous, preparation-specific chemical assays.7 Ultimately, the use of alternative and sometimes inaccurate terms could contribute to owner confusion and inappropriate drug administration. Packaging and drug designs are also important factors in an owner’s ability to safely administer medication to a pet.32,40 Similarities in the appearances of packaging for different drug products or similarities in the appearance of different solid forms for oral dosages have led to the dispensing and administration of the wrong strength of a drug product.32 In the study reported here, each pharmacy used look-alike containers, regardless of drug dosage. Additionally, whereas generic tablet sizes were easily recognizable because of 4 distinct physical characteristics (color, shape, size, and imprinting), strengths of compounded fluconazole capsules were distinguishable on the basis of size or color characteristics, but not both. It is possible that use of individual identifying capsule characteristics, rather than a combination of characteristics, may be insufficient in certain cases to prevent medication errors, which could include an accidental overdose. Taken together, difficulties in reading and interpreting labels, differences in label terminology, and use of look-alike packaging and drug design characteristics may increase the risk of inappropriate dispensing or administration of fluconazole compounded products. The USP and FDA provide strict guidelines for the allowable amount and type of bacterial contamination in compounded drug products.42,43 Microbial contamination of orally administered drugs can be detrimental to a patient in several ways, including induction of infection, alteration of a drug’s physical and pharmacological properties (which may result in diminished or eliminated therapeutic efficacy), conversion of drug ingredients into toxic products, or inclusion of bacterial metabolites or toxins that can


be harmful to patients. In the present study, neither compounded nor generic suspensions yielded any microbial growth after culture for 48 hours. It is noteworthy that the compounded suspensions in the present study had 6-month BUDs. All compounded suspensions in this study used waterbased vehicles. In the absence of stability information applicable to the specific preparation, the USP recommendation for BUDs of nonsterilized, water-containing oral formulations is ≤ 14 days when products are stored at controlled cold (2° to 8°C) temperatures.6 Primary concerns are increased risk of microbial contamination, permeation of packaging, and drug degradation after 14 days. A limitation of the study reported here was the small number of pharmacies, which were restricted geographically to the United States. In addition, we did not evaluate bioavailability of the compounded products or in vivo efficacy. The source of fluconazole used by the various pharmacies could not be ascertained without revealing the identity of the pharmacies; however, the source of fluconazole may have contributed to variations in physical and pharmacological characteristics, such as uniformity of the suspension, purity of the active ingredient, and potential impurities. In turn, this would have the potential to impact the safety or efficacy profiles of fluconazole compounded products. With regard to microbial contamination, the study design did not reflect common clinical use, whereby a compounded suspension generally would be opened and doses removed much more frequently. Thus, there is the potential that exogenous microbial contamination would be greater in a clinical setting. Future studies to evaluate bioavailability and clinical efficacy of fluconazole compounded products are warranted, which ideally would involve obtaining samples from a greater number of pharmacies. Compounded products are not subject to the same premarket requirements for safety, efficacy, and manufacturing quality as are FDA-approved products. As a result, compounded products have the potential to place animals at undue risk from unsafe or ineffective treatment. The present study revealed marked variability in physical characteristics, strength or concentration, accuracy, and precision of fluconazole compounded products from 4 pharmacies in the United States. Quality of suspensions was more variable than that of capsules in this study. Because such variability has the potential to affect absorption, bioavailability, and dosing of the drug, fluconazole compounded products should be prescribed with caution. Clinicians should be cognizant that generic and compounded fluconazole formulations are not interchangeable, and treatment failures, overdoses, or toxicoses may result.

Acknowledgments Supported in part by the American College of Veterinary Dermatology. Presented in abstract form at the 8th World Congress of Veterinary Dermatology, Bordeaux, France, June 2016.

The authors thank Dr. Marike Visser and Elizabeth Zepeda for technical assistance.

Footnotes a. b. c. d. e. f. g. h. i. j. k. l. m. n. o. p. q. r.

Veterinary Pharmacies of America, Houston, Tex. Diamondback Drugs, Scottsdale, Ariz. BCP Veterinary Pharmacy, Houston, Tex. Roadrunner Pharmacy, Phoenix, Ariz. Glenmark, Mahwah, NJ. Citron, East Brunswick, NJ. Greenstone, Peapack, NJ. Orablend, Fagron Inc, Scottsdale, Ariz. Syrspend, Fagron Inc, Scottsdale, Ariz. US Pharmacopeial Convention, Rockville, Md. British Drug House Ltd, Poole, Dorset, England. VWR International, Radnor, Pa. Alliance Waters 2695, Waters Corp, Milford, Mass. Waters 2489 dual λ absorbance detector, Waters Corp, Milford, Mass. Gemini C18, Waters Corp, Milford, Mass. Sunfire C18, Waters Corp, Milford, Mass. BD, Franklin Lakes, NJ. SAS, version 9.2, SAS Institute Inc, Cary, NC.

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