Bioequivalence Evaluation of Topical Metronidazole Products Using Dermal Microdialysis in New Zealand Rabbits.

Comparative assessment of cutaneous pharmacokinetics (cPK) by dermal microdialysis (dMD) appears to be suitable to evaluate the bioequivalence (BE) of topical dermatological drug products applied to the skin (TDDPs). Although dMD studies in the literature have reported inconclusive BE assessments, we have addressed several methodological deficiencies to improve dMD's capability to assess BE between reference (R) and approved generic (referred to as test (T)) gel and cream products of metronidazole (MTZ). The 90% confidence interval (CI) of the geometric mean ratios for the Ln(AUC0-24) and Ln(Cmax) endpoints was centered within the BE limits of 80-125%. The CIs extended outside this range as the proof-of-principle study was not statistically powered to demonstrate BE (N = 7 rabbits). A power analysis suggests that, with the variability observed in this study, 21 rabbits for the cream and 11 rabbits for the gel would be sufficient to support an evaluation of BE with the 2 probe replicates we used, and only 10 and 5 rabbits would be sufficient to power the study for the cream and gel, respectively, if 4 probe replicates are used for each treatment per rabbit. These results indicate that dMD when properly controlling variables can be used to support BE assessments for TDDPs.

Development and in-vivo evaluation of ondansetron gels for transdermal delivery.

Nausea and vomiting are some of the major side effects caused by certain drug therapies, e.g. chemotherapy, radiotherapy and general anesthesia. Because of the nature of the symptoms, oral delivery is inappropriate, while intravenous administration may be unpractical. The aim of the present study was to develop a transdermal gel (2% Klucel®) for ondansetron, a first line 5-HT3-receptor-antagonist antiemetic. The effects of the penetration enhancer camphor and isopropyl-myristate (IPM) were first investigated in-vitro using modified Franz diffusion-cells and then tested in-vivo in a rabbit model by measuring skin and plasma concentrations. Since a disadvantage of transdermal delivery is a prolonged lag-time, the effect of skin treatment with a micro-needle roller was tested. The in-vitro permeation studies through excised porcine ear skin showed that the presence of 2.5% camphor or IPM increased steady state flux by 1.2- and 2.5-fold, respectively, compared to the control gel. Ondansetron was not detectable in either skin or plasma following in-vivo application of the base-gel, whereas the camphor gel and IPM gel delivered 20 and 81 µg/cm(2) of ondansetron, respectively. Microporation led to an increase in plasma Cmax and AUC by 10.47 ± 1.68-fold and 9.31 ± 4.91-fold, respectively, for the camphor gel, and by 2.31 ± 0.53-fold and 1.59 ± 0.38-fold, respectively for the IPM gel. In conclusion, the 2.5% IPM gel demonstrated optimal in-vivo transdermal flux. Skin pretreatment with a micro-needle roller slightly improved the delivery of the IPM gel, whereas dramatically increased the transdermal delivery of the camphor gel.

Effect of microporation on passive and iontophoretic delivery of diclofenac sodium.

Skin pretreatment with a microneedle roller (microporation (MP)) appears a simple and inexpensive technique to increase transdermal delivery of topically applied drug products. This study investigates the effect of MP on the passive and iontophoretic delivery of diclofenac (DCF) by quantifying dermis and plasma levels of DCF in a rabbit model. New Zealand albino female rabbits received either: (i) a topical application of 4 g of Voltaren® 1% gel with or without pretreatment with a microroller (0.5 mm needle length; density 23 microneedles per cm(2) area) or (ii) a DCF solution (40 mg/2.5 mL) via iontophoresis (IOMED transQ(E) medium size patch), with or without microroller pretreatment. A 300 µA/cm(2) cathodic current was applied for 20 min for a total of 80 mA. DCF concentrations were monitored in dermis with microdialysis sampling every 20 min for 5 h. Plasma samples were collected over the same period. In the passive delivery studies, microroller pretreatment increased Cmax by 1.5- and 2.0-fold in skin and plasma, respectively, and AUC by 1.5- and 2.4-fold in skin and plasma, respectively. In the iontophoresis delivery studies, microporation increased Cmax by 2.0-fold both in skin and in plasma, and AUC by 1.1- and 1.8-fold in skin and plasma, respectively. In conclusion, microneedle pretreatment increased significantly the systemic exposure of DCF from either passive or iontophoretic delivery, whereas the effect in skin was less pronounced.

Iontophoresis of amoxicillin and cefuroxime: rapid therapeutic concentrations in skin.

CONTEXT: Amoxicillin (AMX) and cefuroxime (CFX) are antibiotics used often to treat skin bacterial infections. Typically, high oral doses are required to achieve minimum inhibitory concentration (MIC) at the site of infection that may affect only a very small area of skin. OBJECTIVES: To lower side effects and increase therapeutic effectiveness, the percutaneous absorption and retention of AMX and CFX administered by iontophoresis was investigated in a rabbit model by measuring dermis concentrations via microdialysis. METHODS: Iontophoresis was performed using a stainless steel electrode and a non-woven polypropylene pad. The cartridge pad was soaked with a solution of AMX in glycerin or of CFX in glycerin/water (60:40). Constant current density of 0, 100, 200 or 300 µA/cm(2) was applied for 60 min. RESULTS: For AMX, therapeutically effective skin concentrations were detected immediately after the application of electrical current for any of the current density tested and remained above it for at least 2 h from the end of iontophoresis. For CFX, skin concentrations rose above MIC only at the higher current densities and fell below the MIC by the end of the experiment. CONCLUSION: Iontophoresis is a promising method to obtain a fast and sustained concentration of AMX and CFX in skin.

The dose-duration effect on cutaneous pharmacokinetics of metronidazole from topical dermatological formulations in Yucatan mini-pigs.

Dermal microdialysis (dMD) permits the investigation of cutaneous pharmacokinetics (cPK) for topical dermatological drug products (TDDP). dMD involves probe implantation into the dermis and a sample collection system that restricts subjects' movements for the experimental duration. A truncated dose-duration, by TDDP removal at predetermined time-points, may help to adequately characterize the cPK in a relatively short time. The goals of this study were to: assess and compare the dose-duration effect on the dermal exposure of metronidazole (MTZ) containing TDDPs; and characterize MTZ dermal elimination following TDDP application and direct dermal delivery of MTZ utilizing a retrodialysis/microdialysis approach that we termed "dermal infusion." MTZ cream and gel were applied on three Yucatan mini-pigs for dose-durations of 6-hr, 12-hr, or 48-hr. The gel's dermal exposure was similar among the three dose-durations. Conversely, at the 6-hr dose-duration, the cream's dermal exposure was significantly lower than other cream dose-durations while also comparable to the gel. In comparison, the 12-hr and 48-hr cream exposures were not significantly different. Terminal-phase half-live differences between the MTZ TDDP's and dermal-infusion indicate flip/flop cPK. Truncating topical dose-duration may provide a valuable strategy to reduce experimental duration; however, dose-duration must be carefully selected if the goal is to discriminate between formulations.

Quantification of dermal and transdermal delivery of meloxicam gels in rabbits.

BACKGROUND: This study was designed to quantify the effects of penetration enhancers on systemic bioavailability of 0.3% meloxicam (MLX) hydroxypropylcellulose gels. Cutaneous microdialysis was also performed to assess dermis availability and to better understand the penetration process. The gels tested were a 1% oleic acid gel, a 5% menthol gel, and a control gel without penetration enhancers. METHODS: To assess systemic bioavailability, three female rabbits received according to a crossover design 0.135 g/cm(2) of gel applied to a 7.5 × 7.5 cm area of their shaved back and a short (5 min) infusion of 1 mg. In each experiment, blood samples were collected serially for 36 h and analyzed by a validated HPLC method. For skin bioavailability studies, 0.135 g/cm(2) of the same gels were applied to a 1 × 2 cm area on top of a microdialysis probe previously inserted in the dermis. Dialysate samples were collected for 6 h every 30 min. RESULTS: Systemically, the 5% menthol gel delivered 3.93 ± 0.85 mg of MLX versus the 1.41 ± 0.24 mg of the oleic acid gel. Only traces of MLX were detectable from the control gel. In dermis, substantial concentrations of MLX were detected only after the application of the menthol gel, whereas skin concentration from the control gel and the 1% oleic acid gel were always below the lowest limit of quantification (LLOQ). CONCLUSIONS: The 5% menthol gel can possibly deliver therapeutically relevant amount of MLX in vivo. Dermis concentrations can be predictive of systemic plasma levels.

Evaluation of local bioavailability of metronidazole from topical formulations using dermal microdialysis: Preliminary study in a Yucatan mini-pig model.

Dermal microdialysis (dMD) can measure the rate and extent to which a topically administered active pharmaceutical ingredient (API) becomes available in the dermis. Using multiple test-sites on the same subject, and replicate probes at each test-site, it is feasible to compare the cutaneous pharmacokinetics of an API from different topical dermatological drug products in parallel on the same subject with this technique. This study design would help to reduce variability. However, there are technical considerations related to the dMD experimental methods that must be characterized and optimized to ensure that an in vivo dMD study is selective, sensitive, discriminating, and reproducible. The goals of this study were to assess: the minimum distance required between test-sites to prevent cross-talk between probes due to potential lateral-diffusion; the sensitivity of the dMD method to detect differences in the local concentration of metronidazole (MTZ) among single escalating doses; the ability to discriminate between the two different formulations; and the stability of the dMD-probes over 48 h. Results indicate that lateral-diffusion and systemic redistribution of the API following topical application of the drug product were negligible, thus MTZ measured by dMD can be selectively attributed to the dermal bioavailability of the API from the applied topical dose. The dMD methodology was able to detect differences in the bioavailability of MTZ from the cream compared to the gel when applied at the same dose, as well as among different doses of the same formulation over a 48-hour sampling duration; therefore, the method is sensitive. The percentage loss of D3-MTZ from the probe compared to its original concentration in the perfusate indicates that the probe performance was stable over the 48 h.

Pharmacokinetics of amitriptyline in rabbit skin and plasma following iontophoretic administrations.

BACKGROUND: Amitriptyline (AMT) is a tricyclic antidepressant with demonstrated local analgesic effects in human skin. AIM: We investigated the feasibility and mechanisms of iontophoretic delivery of AMT to rabbit dermis and plasma. METHOD: Two microdialysis probes were inserted into the upper dorsal shaved skin of tranquilized rabbits. After 1 hour, an iontophoresis cartridge was placed on top of one probe. The cartridge consisted of a stainless steel electrode covered with a pad that was filled with a 4.3 % AMT glycerin/water (50:50). Iontophoresis was performed at 100, 200, or 300 microA/cm(2) constant-current density for 60 minutes. Dialysate samples were collected every 8 minutes for 3 hours and analyzed for AMT via a validated high-performance liquid chromatographic assay. Retrodialysis was performed at the other site. Blood samples were collected serially for 4 hours. RESULTS: In vivo retrodialysis recovery was 89 +/- 2%. AMT skin exposure increased non-proportionally with current density: AUCs were 19 +/- 7, 119 +/- 56, and 615 +/- 302 mg/L/min for the 100, 200, and 300 muA/cm(2), respectively, and C(max) were 107 +/- 15, 1070 +/- 537, and 5870 +/- 1289 microg/L. In vivo plasma concentrations were always below LLOQ (0.1 microg/mL). CONCLUSION: AMT can be administered by iontophoresis and produces significant skin concentrations and negligible plasma levels.

Quantification and prediction of skin pharmacokinetics of amoxicillin and cefuroxime.

The purpose of this project was to develop and validate a pharmacokinetic model and to quantify the rate and extent of distribution between plasma and skin of two beta-lactam antibiotics, amoxicillin (AMX) and cefuroxime (CFX), which are frequently administered systemically to treat skin and skin structure infections. Dosing regimens are usually based on plasma concentration, however, concentrations at the target site are better correlated with the effect. For each antibiotic, three different i.v. bolus doses were administered to three female rabbits according to a randomized cross-over design and plasma samples were collected serially. Skin concentrations were obtained by continuous microdialysis. Skin and unbound plasma concentrations were fitted simultaneously using a semi-physiological model and the transfer constants plasma/skin (K(in)) and skin/plasma (K(out)) were estimated. K(in) and K(out) were then used to predict skin concentrations from the plasma levels obtained from an oral administration of AMX or from an i.v. bolus of CFX. The predicted skin profiles were similar to those measured by microdialysis during the actual experiments. In conclusion, this study shows that it is possible to generate a reasonable prediction of skin pharmacokinetics from any plasma level once a careful characterization of the transfer process between plasma and skin has been made.

Pharmacokinetic applications of cutaneous microdialysis: Continuous+intermittent vs continuous-only sampling.

INTRODUCTION: Microdialysis is a technique that allows interstitial-fluid sampling with minimal tissue-damage. In a microdialysis study, samples are collected serially (continuous microdialysis, CMD) and participant's movements are reduced for the entire study. Intermittent Cutaneous Microdialysis (IMD) is a modified version of CMD, which allows for unrestrained periods in between samples. However, in separate experiments, pharmacokinetic parameters estimated with IMD showed higher variability than with CMD. The purpose of this study is to simultaneously assess and compare the skin pharmacokinetic profiles obtained with a combination of CMD and IMD with those obtained with traditional CMD sampling only, in the same experiment. METHODS: Two linear microdialysis (MD) probes were inserted into the shaved dorsal skin of three rabbits. Following the oral administration of three different doses (20, 40 and 80mg/kg) of ciprofloxacin (CPLX), for the first 2h, samples were collected from both probes according to traditional CMD in order to assess intrinsic differences between the two sites. After 2h, one of the probes was switched to IMD schedule. Skin-exposure parameters were estimated with non-compartmental analysis. RESULTS: Two of the nine experiments showed a difference larger than 30% between the concentrations measured from the two probes when both were on the CMD schedule. Otherwise, the skin concentration profiles were almost superimposable. Pharmacokinetic parameters were not statistically different. CONCLUSION: The results of this study show that skin pharmacokinetic parameters measured via a combination of CMD and IMD were not statistically different from those estimated via traditional CMD sampling alone.

Cutaneous Penetration-Enhancing Effect of Menthol: Calcium Involvement.

Menthol is a naturally occurring terpene used as a penetration enhancer in topical and transdermal formulations. Literature shows a growing interest in menthol's interactions with the transient receptor potential melastatin 8. A decrease in extracellular Ca2+ due to the activation of the transient receptor potential melastatin 8 receptor produces inhibition of E-cadherin expression that is responsible for cell-cell adhesion. Because calcium is present in the entire epidermis, the purpose of this study is to evaluate whether the aforementioned properties of menthol are also related to its penetration-enhancing effects. We formulated 16 gels: (i) drug-alone (diphenhydramine or lidocaine), (ii) drug with menthol, (iii) drug, menthol, and calcium channel blocker (CCB; verapamil or diltiazem), and (iv) drug and CCB. In vitro studies showed no effect of the CCB on the release of the drugs either with or without menthol. In vivo experiments were performed for each drug/menthol/CCB combination gel by applying 4 formulations on a shaved rabbit's dorsum on the same day. Dermis concentration profiles were assessed with microdialysis. The gels containing menthol showed higher penetration of drugs than those without whereas the addition of the CCB consistently inhibited the penetration-enhancing effects of menthol. In summary, these findings strongly support the involvement of calcium in the penetration-enhancing effect of menthol.

Fingolimod hydrochloride gel shows promising therapeutic effects in a mouse model of atopic dermatitis.

OBJECTIVES: To assess the efficacy of topically applied 2% hydroxypropyl cellulose gels containing 0.5% fingolimod hydrochloride (FNGL) with or without 6% colloidal oatmeal in an in vivo model of atopic dermatitis (AD). METHODS: AD-like lesions were induced in SKH1/Hr hairless mice and were treated with FNGL gels, non-medicated base gels and Elidel(®) cream for 6 weeks. The severity/improvement of the lesions was assessed regularly using the Eczema Area and Severity Index (EASI), pH of the skin, transepidermal water loss, g/m(2) /h (TEWL), humidity and temperature. At the end of the experiments, the plasma levels of cytokines, FNGL and white blood cells were determined. KEY FINDINGS: The EASI score was almost unchanged for the vehicle-only groups compared to before the treatments, whereas the medicated groups showed a significant decrease in the overall EASI score (P < 0.01), although there was non-significant differences among them (P > 0.081). Both the FNGL groups also showed a significant (P ˂ 0.05) reduction in blood WBC. CONCLUSION: This study shows that the gels containing 0.50% FNGL and FNGL 0.50% plus 6% colloidal oatmeal have potential for the treatment of AD. The presence of colloidal oatmeal may provide additional benefits.

Transdermal Delivery of Etoposide Phosphate II: In Vitro In Vivo Correlations (IVIVC).

A dependable in vitro in vivo correlation (IVIVC) is a vital tool to optimize drug formulation and expedite product development time. Although many IVIVC examples are available for oral delivery systems, IVIVC for transdermal delivery is far less common, especially for electrical-assisted delivery. The objective of this study was to develop an IVIVC for the iontophoretic delivery of the anticancer drug etoposide. Iontophoresis was performed at 4 current densities (100, 200, 300, and 400 µA/cm(2)) both in vitro using a standard Franz-cell apparatus with excised porcine skin as membrane, and in vivo in a rabbit model. There was strong correlation between the in vitro % permeated across porcine skin and in vivo absorption (AUC, Cmax) in the range 100-300 µA/cm(2). The correlation between in vitro flux and in vivo input rate (R0) permitted to predict the R0 from a different set of in vitro data (external validation). Convolution of such input rate accurately predicted in vivo plasma profiles (PE% <15) in the absorption phase, whereas the elimination phase was slightly under-predicted (PE% >20). In vivo absorption profiles obtained with deconvolution did not overlap directly with the in vitro profiles; however, correction for the lag time and the application of a scaling factor estimated from Levy' s plots resulted in excellent correlation.

Transdermal Delivery of Etoposide Phosphate I: In Vitro and In Vivo Evaluation.

Cancer chemotherapy frequently requires long periods of multiple intravenous infusions that often results in patients opting out of treatment. The main purpose of this study was to investigate the feasibility of delivering one of these anticancer agents: etoposide phosphate (ETP) transdermally using iontophoresis and a combination of iontophoresis/microporation. The iontophoresis conditions for ETP were first optimized in vitro then tested in vivo in a rabbit model. Both ETP and its active form etoposide (VP) were quantified in dermis (via microdialysis sampling) and in plasma, with a specially developed high-performance liquid chromatography method. In vitro, the amount of total etoposide permeated and the steady state flux increased (p < 0.05) with increase in iontophoretic current densities (100-400 µA/cm(2)). At 300 µA/cm(2), microporation/iontophoresis further improved both parameters by 2- and 2.8-fold, respectively. In vivo, exposure increased proportionally to current density in plasma, whereas dermal concentration dropped significantly at the highest current density. Microporation led to a 50% increase in Cmax and AUClast values in both skin and plasma. In conclusion, a mild current density (300 µA/cm(2)) and a small surface area (10.1 cm(2)) achieved and maintained the minimum effective concentration for the entire duration of electrical current delivery; microporation further increased the plasma concentrations at the same current density.

Permeation study of five formulations of alpha-tocopherol acetate through human cadaver skin.

Alpha-tocopherol (AT) is the vitamin E homologue with the highest in vivo biological activity. AT protects against the carcinogenic and mutagenic activity of ionizing radiation and chemical agents, and possibly against UV-induced cutaneous damage. For stability consideration, alpha-tocopherol is usually used as its prodrug ester, alpha-tocopherol acetate (ATA), which once absorbed into the skin is hydrolyzed to alpha-tocopherol, the active form. The objective of this research was to characterize in vitro the permeation properties of ATA from various solutions and gel formulations. Permeation studies were conducted using modified Franz diffusion cells and human cadaver skin as the membrane. Specifically, 5% (w/w) alpha-tocopherol acetate was formulated in the following vehicles: ethanol, isopropyl myristate, light mineral oil, 1% Klucel gel in ethanol, and 3% Klucel gel in ethanol (w/w). The receiver temperature was 37 degrees C. Samples from the receiver were collected at 2, 4, 6, 8, 12, 24, 30, 36, and 48 hours and analyzed by HPLC for concentrations of alpha-tocopherol acetate and alpha-tocopherol. The permeabilities of ATA through human cadaver skin were 1.0x10(-4), 1.1x10(-2), 1.4x10(-4), 2.1x10(-4), and 4.7x10(-4) cm/h for the ethanol solution, isopropyl myristate solution, light mineral oil solution, 1% Klucel gel, and 3% Klucel gel, respectively. The results show that the formulation had relatively minor effects on the permeability coefficients of ATA through cadaver skin in all cases except for the isopropyl myristate solution.

Pharmacokinetics of methotrexate in rabbit skin and plasma after iv-bolus and iontophoretic administrations.

The pharmacokinetics of methotrexate (MTX) in rabbit's skin and plasma after iv-bolus and iontophoretic delivery at different current densities was studied. Linear microdialysis probes were introduced into the upper dorsal shaved skin of tranquilized rabbits. Commercially available patches were used to deliver MTX for 1 h at different current densities (100, 200, and 300 microA/cm2) on different occasions. Iv-boluses (10 mg/kg) of MTX were also administered. Retrodialysis was performed at the end of the experiments to estimate probe recovery. Plasma and microdialysis samples were analyzed using a validated HPLC assay. Following iv-bolus, MTX showed a bi-exponential decay both in plasma and in skin. Cmax in skin occurred with a delay of 22 min compared with plasma. No quantifiable concentration of MTX was detected in the skin on passive drug delivery. Systemic exposure to MTX (AUC) and Cmax increased linearly with current density. Nevertheless, exposure to MTX in the skin did not increase linearly with current density, whereas Cmax did. In conclusion, iontophoresis remarkably improved the dermal delivery of MTX over passive diffusion. However, total exposure did not increase with current density in the skin, suggesting that for local applications lower current densities may achieve the same effects with minimal systemic exposure.

Pharmacokinetics of acyclovir in rabbit skin after i.v.-bolus, ointment, and iontophoretic administrations.

The aim of this study was to characterize and compare the pharmacokinetics of acyclovir (ACV) in skin and plasma after iontophoresis, i.v.-bolus, and ointment administrations in rabbit. On five occasions, each separated by at least 1-week washout, rabbits received a 10 mg/kg dose of ACV as i.v.-bolus, ACV iontophoresis for 1 h at different current densities (100, 200, 300 microA/cm2) or a commercially available ointment for two hours. Blood samples were collected serially up to 6 h. Skin ACV concentrations were monitored via microdialysis using linear microdialysis probes (1 cm window). Cathodic iontophoresis was performed using commercially available patches (10 cm2 contact area). Following i.v.-bolus, C(max) in skin occurred with a delay of 38 +/- 4 min compared with plasma. No quantifiable concentration of ACV was detected in the skin on passive drug delivery. Following iontophoresis, skin exposure to ACV was 40, 22, and 11% of that following i.v.-bolus. Conversely, systemic exposure to ACV was negligible and plasma concentrations were below the limit of quantification at any time-point. In skin dialysate, C(max), AUC, and half-life increased with current density. During ointment application, ACV in dermis was detectable only for the first 30 min thereafter ACV skin concentrations were below the LOQ (30 ng/ml).

Iontophoresis successfully delivers dexamethasone sodium phosphate to dermis as measured by microdialysis.

Despite its widespread and long term use, the effectiveness of iontophoresis to increase the delivery of dexamethasone sodium phosphate (DSP) remains controversial. The goal of this study was to quantitatively compare the DSP concentrations in dermis' dialysates in two delivery scenarios: with and without iontophoresis. Interstitial fluid concentrations were measured by cutaneous microdialysis. Passive and active iontophoresis were applied simultaneously on the skin of the forearm in eight healthy adult participants using each participant as his/her own control. The iontophoresis apparatus and procedures were identical to those used in common clinical practice. Iontophoresis electrodes were loaded with 2 mL of 4.4 mg/mL of preservative-free DSP solution. Electric current (4 mA) was applied for 20 min. Dialysate samples were collected for 2 h and analyzed for DSP and its active metabolite dexamethasone (DXM). Seven out of eight iontophoresis sites contained quantifiable levels of DSP and DXM, whereas none of the samples collected at the passive site contained either form of the drug. In conclusion, this study demonstrates that iontophoresis significantly (p < 0.0001) increases delivery of DSP to the dermis compared with passive delivery of the same, and that microdialysis can be used to monitor DSP delivery and DXM formation in skin.

Skin microdialysis-based estimation of systemic bioavailability fraction.

Systemic bioavailability is usually determined from plasma data. However, when plasma is difficult to access, as in young children, alternative methods would be particularly beneficial. The present study investigates the possibility of calculating systemic bioavailability fraction (F) from skin concentrations measured by two microdialysis (MD) sampling methods: continuous microdialysis and intermittent microdialysis. When the drug concentration in skin is a linear and time-invariant function of plasma concentration, the area under the drug concentration curve in skin is directly proportional to the drug absorbed systemically. To verify this theory, we compared the F estimated from MD concentrations in the skin with that obtained from the plasma data in the same experiment. Two model drugs were selected for the study: amoxicillin and ketoprofen. Drugs were administered to rabbits as intravenous infusion or oral suspension according to a randomized crossover design. F estimated by either MD method was not significantly different from that obtained from the plasma for both drugs tested. However, the skin data exhibited a larger variability. These results confirm that skin MD could be an alternative way to obtain data for the calculation of systemic fraction of drug absorbed.

Optimization of perfusate pH to improve microdialysis recovery of lipophilic compounds.

INTRODUCTION: Microdialysis (MD) allows sampling of compounds in-vivo from tissues' interstitial fluid. However, molecules insoluble at physiological pH have usually extremely low recovery. The addition of albumin to the perfusate or the use of isotonic lipoemulsion improves recovery of these molecules although it requires a cleaning step before HPLC analysis. This study investigates the possibility of improving the MD recovery of compounds insoluble at physiological pH but soluble at a different pH. The probe is perfused with an isotonic solution adjusted to pH values at which the compound has maximum solubility. Ketoconazole (KTC), clotrimazole (CLT) and tretinoin (TTN) were selected as model drugs because they are almost insoluble at pH 7.4 but soluble at pH 4 for KTC and CTL; and at pH 9 for TTN. METHODS: Linear microdialysis probes were used to collect KTC, CLT or TTN from a standard solution of the compounds. Probes were perfused with 0.01 M pH 7.4 isotonic buffer solution (1) without or (2) with 5% Bovine Serum Albumin (BSA); or (3) with 20% isotonic lipoemulsion; or (4) with 0.01 M pH 4 isotonic buffer solution for KTC and CLT or 0.01 M pH 9 isotonic buffer solution for TTN. The method was then tested in-vivo, in rabbit skin, to assess the skin tolerance to the non-physiological perfusates and to monitor KTC and TTN delivery from commercial cream products. RESULTS: In-vitro, the optimized-pH perfusate increased MD recovery significantly (P<0.001): 6.9 (KTC), 8.3 (CLT), and 2.0 (TTN) times compared to the physiological pH and 1.4 and 1.2 compared to the BSA and lipoemulsion respectively. No evidence of irritation or edema was observed in-vivo. However, KTC and TTN were not detected in-vivo with any of the modified perfusate tested. DISCUSSION: These findings show that the optimized-pH perfusate effectively increases the in-vitro microdialysis recovery of KTC, CLT and TTN and that it is well tolerated in-vivo. However, the compounds tested (KTC and TTN) could not be detected in-vivo.