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Mariana Sato de Souza de Bustamante Monteiro,1 Rafael Antonio Ozzetti,2 André Luiz Vergnanini,2 Lycia de Brito-Gitirana,3 Nadia Maria Volpato,4 Zaida Maria Faria de Freitas,1 Eduardo Ricci-Júnior,1 Elisabete Pereira dos Santos11Medicines Department, Federal University of Rio de Janeiro, Rio de Janeiro, 2University of Campinas, São Paulo, 3Laboratory of Animal and Comparative Histology, Institute of Biomedical Science, Federal University of Rio de Janeiro, Rio de Janeiro, 4Department of Production and Medicine Control, Federal University of Rio Grande do Sul, Rio Grande do Sul, BrazilPurpose: Awareness of the harmful effects of ultraviolet radiation has led to the increasing use of sunscreens, thus, the development of safe and effective antisolar preparations is important. The inclusion of sunscreen molecules in different release systems, like liposomes (lipo) and cyclodextrins (CD) is therefore required.Methods: The in vivo sun protection factor (SPF), water resistance, and in vitro transdermal penetration test of octyl p-methoxycinnamate (OMC) in different dispersions, such as OMC encapsulated in liposomes (lipo/OMC), OMC encapsulated in β-cyclodextrins (β-CD/OMC), OMC encapsulated in both release systems (lipo/OMC and β-CD/OMC), and an OMC-free formulation were determined.Results: Although the formulation containing only the lipo/OMC system revealed high value of in vivo SPF (11.0 ± 1.3) and water resistance (SPF = 10.3 ± 2.2), the formulation containing both release systems (lipo/OMC + β-cyclodextrin/OMC) showed the best result in the in vivo SPF test (11.6 ± 1.6). In the penetration test, the formulation containing the lipo/OMC system had better performance, since a high amount of OMC in the epidermis (18.04 ± 1.17 µg) and a low amount of OMC in the dermis (9.4 ± 2.36 µg) were observed. These results suggest that liposomes interact with the cells of the stratum corneum, promoting retention of OMC in this layer.Conclusion: According to our study, the lipo/OMC system is the most advantageous release system, due to its ability to both increase the amount of OMC in the epidermis and decrease the risk of percutaneous absorption.Keywords: octyl p-methoxycinnamate, sunscreen, liposomes, β-cyclodextrin, penetration

The absence of oral liquid pharmaceutical forms appropriate for use in pediatric and adult patients with difficulty swallowing is a public health problem, especially in the hospital context. Baclofen is a muscle relaxant of choice for treating spasticity, generally marketed only in tablet form, highlighting the need for liquid formulations to facilitate dose adjustment, administration, and swallowing. The present study aimed to develop oral liquid formulations containing baclofen, optimize them through the quality by design approach, and evaluate their physicochemical and microbiological stability. Preformulation and preliminary stability studies were carried out for the development of formulations. Experimental screening and optimization designs resulted in eleven experiments for each step that were evaluated for 28 days. A stability-indicating method by high-performance liquid chromatography presented linearity, low limits of detection and quantification, precision, accuracy, and robustness. The experimental design led to two optimized formulations containing baclofen, glycerin, potassium sorbate, citric acid, ultrapure water, flavor, and sucrose syrup or sodium carboxymethylcellulose solution as a vehicle, the last one with sucralose as a sweetener. The formulations were placed in amber glass flasks and subjected to a physicochemical and microbiological stability study. Both formulations showed physicochemical and microbiological stability when stored at room temperature and refrigerated for 84 days. The results of this study may serve as a reference in the preparation of liquid oral formulations containing baclofen in the hospital routine and collaborate with the safety and adherence to the treatment of adult and pediatric patients., (© 2022. The Author(s), under exclusive licence to American Association of Pharmaceutical Scientists.)

Analysis of impurities is an important step in the quality control of pharmaceutical ingredients and final products. From drug synthesis or excipients, even in small concentrations, impurities may affect efficacy and safety. The method was developed following Quality by Design (QbD) for the analysis of the antidiabetic empagliflozin. The concept of QbD is used as a tool for the development of methods and formulations. Through predefined objectives and risk analysis, robust methodologies and reduced solvent consumption are developed. A simple HPLC method was developed and validated for the quantitative determination of empagliflozin and its organic impurities from the synthesis process. The method was carried out in a Shim-pack phenyl column with a mobile phase consisting of an acetonitrile/water mixture (72 : 28), with isocratic elution and the detector wavelength was 230 nm. The validation process, in accordance with international guidelines, shows that the method was linear, precise and accurate for empagliflozin, impurity 1 and impurity 2. Limits of detection (0.01, 0.02 and 0.01 μg mL -1 ) and quantification (0.10, 0.10 and 0.05 μg mL -1 ) were determined for EMPA, IMP1 and IMP2, respectively. The HPLC method for impurity determination in empagliflozin was linear, precise, accurate and robust. It can be successfully applied in the quality control of empagliflozin and the synthesis of impurities, being adequate for routine analysis., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

Background: The presence of impurities in some drugs may compromise the safety and efficacy of the patient's treatment. Therefore, establishing of the biological safety of the impurities is essential. Diabetic patients are predisposed to tissue damage due to an increased oxidative stress process; and drug impurities may contribute to these toxic effects. In this context, the aim of this work was to study the toxicity, in 3 T3 cells, of the antidiabetic agents sitagliptin, vildagliptin, and their two main impurities of synthesis (S1 and S2; V1 and V2, respectively)., Methods: MTT reduction and neutral red uptake assays were performed in cytotoxicity tests. In addition, DNA damage (measured by comet assay), intracellular free radicals (by DCF), NO production, and mitochondrial membrane potential (ΔψM) were evaluated., Results: Cytotoxicity was observed for impurity V2. Free radicals generation was found at 1000 μM of sitagliptin and 10 μM of both vildagliptin impurities (V1 and V2). A decrease in NO production was observed for all vildagliptin concentrations. No alterations were observed in ΔψM or DNA damage at the tested concentrations., Conclusions: This study demonstrated that the presence of impurities might increase the cytotoxicity and oxidative stress of the pharmaceutical formulations at the concentrations studied.

Objective: This study aimed to develop and validate an in vitro dissolution method based on in silico-in vivo data to determine whether an in vitro-in vivo relationship could be established for rivaroxaban in immediate-release tablets., Significance: Oral drugs with high permeability but poorly soluble in aqueous media, such as the anticoagulant rivaroxaban, have a major potential to reach a high level of in vitro-in vivo relationship. Currently, there is no study on scientific literature approaching the development of RIV dissolution profile based on its in vivo performance., Methods and Results: Drug plasma concentration values were modeled using computer simulation with adjustment of pharmacokinetic properties. Those values were converted into drug fractions absorbed by the Wagner-Nelson deconvolution approach. Gradual and continuous dissolution of RIV tablets was obtained with a 30 rpm basket on 50 mM sodium acetate +0.2% SDS, pH 6.5 medium. Dissolution was conducted for up to 180 min. The fraction absorbed was plotted against the drug fraction dissolved, and a linear point-to-point regression (R 2  = 0.9961) obtained., Conclusion: The in vitro dissolution method designed promoted a more convenient dissolution profile of RIV tablets, whereas it suggests a better relationship with in vivo performance.

A simple, fast and sensitive analytical method by high-performance liquid chromatography (HPLC) was developed and validated for the simultaneous determination of ticagrelor and two synthesis impurities. The HPLC method was established using an Agilent 1200 Series equipment coupled to photodiode array detector (PDA) at 270nm with a Zorbax Plus C 8 column (150×4.6mm, 5.0μm), injection volume of 20μL, and a constant temperature of 25°C. The mobile phase consisted of acetonitrile: ammonium acetate 50mM (57:43, v/v) and pH adjusted to 8.2 with ammonium hydroxide 6M, at a flow rate of 0.7mL/min. No interference peaks from excipients and diluent system indicated the specificity of the method. The calibration curves showed determination coefficients (r 2 )>0.99, calculated by linear regression. The limit of quantitation (LOQ) for impurities 1 and 2 were 2.0 and 0.2μg/mL, respectively. Intra and interday relative standard deviations (RSDs) were <2% for ticagrelor and <6% for the impurities, proving the precision of the method. Besides, two mayor degradation products formed when sample solutions of ticagrelor were exposed to UVC radiation were elucidated and the mechanisms involved in the photolytic degradation of ticagrelor were proposed., (Copyright © 2016 Elsevier B.V. All rights reserved.)

The present study describes the development and validation of a dissolution method for delapril (DEL) and manidipine (MAN) combination tablets, using a simulated absorption profile based on in vivo data for MAN. The suitable in vitro dissolution profile for this formulation was obtained using 900 mL of citrate buffer pH 3.2 at 37 °C±0.5 °C as dissolution medium and USP apparatus 2 (paddle) at 75 rpm. All samples were analyzed by a liquid chromatography (LC) method. Under these conditions, a significant linear relationship between the absorbed (calculated by deconvolution approach) and dissolved fractions of MAN was obtained ( R =0.997) and an in vivo-in vitro (IVIV) correlation for this particular formulation containing MAN can be established. Validation parameters for dissolution methodology such as the specificity, linearity, accuracy and precision were also evaluated according to the international guidelines, giving results within the acceptable range. Therefore, the proposed dissolution conditions can be applied for the simultaneous release analysis of DEL and MAN from the solid dosage form, contributing to the improvement of the quality control of pharmaceutics and minimizing the number of bioavailability studies.

Gemifloxacin mesylate (GFM) is a synthetic, broad-spectrum, fluoroquinolone antibacterial agent. It is different from other class members because it achieves adequate plasma concentrations to inhibit both topoisomerase IV and gyrase. The aim of this study was to develop and validate a dissolution test for GFM in coated tablets, using a simulated absorption profile based on in vivo data obtained from the literature. The fraction and percentage of the dose absorbed were calculated using model-dependent Loo-Riegelman approach for two compartments. The best in vitro dissolution profile was obtained using 900 mL of pH 6.0 phosphate buffer as a dissolution medium at 37 °C ± 0.5 °C and paddles at 50 rpm. The in vitro dissolution samples were analyzed using a liquid chromatography method, and the validation was performed according to USP 34 (2011). The method showed specificity, precision, accuracy, robustness and linearity. Under these conditions, a level-A in vitro-in vivo correlation was suggested (r = 0.9926). The prediction errors were calculated to determine the validity and accuracy of the suggested correlation. The dissolution test can be used to evaluate the dissolution profile of GFM-coated tablets and minimize the number of bioavailability studies as part of new formulation development.

A stability-indicating capillary zone electrophoresis (CZE) method was validated for the determination of vildagliptin (VLG) in pharmaceutical dosage forms using ranitidine hydrochloride (RH) as internal standard. The CZE method was carried out in a fused silica capillary (64.5 cm total length and 56.0 cm effective length, 50 microm i.d.) by applying a potential of 25 kV (positive polarity), hydrodynamic injection by 50 mbar for 5 s and the temperature of the capillary cartridge was 25 degreesC. The selected background electrolyte (BGE) consisted of 25 mM potassium phosphate (pH 8.0) with UV/PDA detection at 207 nm. The electrophoretic separation was obtained within 6 min and was linear in the range of 50-200 microg/mL (r= 0.9994). The specificity and the stability-indicating capability were demonstrated through degradation studies, which also showed that there was no interference of the formulation excipients. The method was validated in accordance to ICH guidelines acceptance criteria for specificity, linearity, precision, accuracy, robustness and system suitability. The proposed method was compared with HPLC method previously validated for this drug, and statistical analysis showed no significant difference between the methods.

A dissolution method to analyze atorvastatin tablets using in vivo data for RP and test pilot (PB) was developed and validated. The appropriate conditions were determined after solubility tests using different media, and sink conditions were established. The conditions used were equipment paddle at 50 rpm and 900 mL of potassium phosphate buffer pH 6.0 as dissolution medium. In vivo release profiles were obtained from the bioequivalence study of RP and the generic candidate PB. The fraction of dose absorbed was calculated using the Loo-Riegelman method. It was necessary to use a scale factor of time similar to 6.0, to associate the values of absorbed fraction and dissolved fraction, obtaining an in vivo-in vitro correlation level A. The dissolution method to quantify the amount of drug dissolved was validated using high-performance liquid chromatography and ultraviolet spectrophotometry, and validated according to the USP protocol. The discriminative power of dissolution conditions was assessed using two different pilot batches of atorvastatin tablets (PA and PB) and RP. The dissolution test was validated and may be used as a discriminating method in quality control and in the development of the new formulations.

A stability-indicating micellar electrokinetic chromatography (MEKC) method was developed and validated for simultaneous analysis of delapril (DEL) and manidipine (MAN) using salicylic acid as an internal standard. The MEKC method was performed using a fused-silica capillary (effective length of 72 cm) with 50 mM of borate buffer and 5 mM of anionic surfactant sodium dodecylsulfate at pH 9.0 as the background electrolyte. The separation was achieved at 25 kV applied voltage and 35 degrees C. The injection was performed at 50 mbar for 5 s, with detection at 208 nm. The method was linear in the range of 15-150 microg/mL (r2 = 0.9966) for DEL and 5-50 microg/mL (r2 = 0.9985) for MAN with adequate results for the precision (< or = 1.87%) and accuracy (98.94% for DEL and 100.65% for MAN). The specificity of the method and its stability-indicating capability was demonstrated through forced degradation studies, which showed that there was no interference from the excipients. The Plackett-Burman experimental design was used for robustness evaluation, giving results within the acceptable range. The method was successfully applied for analysis of the drugs, and the results were compared to an LC method, resulting in nonsignificant differences (P = 0.78 and 0.84 for DEL and MAN, respectively).

This study evaluated the use of ultrasound in combination with the commercial anti-inflammatory drugs ketoprofen and sodium diclofenac, according to the parameters used in physiotherapy. Ketoprofen and sodium diclofenac were used in the Franz diffusion cell model adapted to an ultrasound transducer in three conditions: no ultrasound, one application of ultrasound and two applications of ultrasound. High-performance liquid chromatography was used to quantify the total amount of drug permeating skin per unit area, as well as flux and latency. The results showed that for ketoprofen, the amount of drug permeating skin and flux increased with two ultrasound applications. Permeation of sodium diclofenac decreased in the presence of ultrasound. Ultrasound parameters and drug properties must be considered in the use of phonophoresis., (Copyright © 2013 World Federation for Ultrasound in Medicine & Biology. Published by Elsevier Inc. All rights reserved.)

A first-order derivative spectrophotometric (1D-UV) method was developed and validated for simultaneous determination of delapril (DEL) and manidipine (MAN) in tablets. The 1D-UV spectra were obtained using change lambda = 4.0 nm and wavelength set at 228 nm for DEL and 246 nm for MAN. The method was validated in accordance with the ICH requirements, involving the specificity, linearity, precision, accuracy, robustness and limits of detection and quantitation. The method showed high specificity in the presence of two drugs and formulation excipients and was linear over the concentration range of 18-54 microg mL(-1) (r2 = 0.9994) for DEL and 6-18 microg mL(-1) (r2 = 0.9981) for MAN with adequate results for the precision (< or = 1.47%) and accuracy (98.98% for DEL and 100.50% for MAN). Moreover, the method proved to be robust by a Plackett-Burman experimental design evaluation. The proposed 'D-UV method was successfully applied for simultaneous analysis of DEL and MAN in tablets and can be used as alternative green method to separation techniques. The results were compared with the validated liquid chromatography, capillary electrophoresis and liquid chromatography-tandem mass spectrometry methods, showing non-significant difference.

Objective: The present study aimed to evaluate the influence of the relative composition of trace elements and vitamins in physicochemical stability of neonatal parenteral nutrition., Material and Methods: Three formulations for neonatal administration were selected; the main variable was the presence of trace elements and vitamins. The analyses where carried out immediately after preparation and at 24 h, 48 h, 72 h and 7 days after preparation. Three methods were selected to determine globule size: light obscuration, dynamic light scattering and optical microscopy. Complementary evaluation including visual inspection, determination of pH and osmolarity, peroxide levels and measurements of zeta potential were also performed., Results: There was an observable alteration in color and phase separation in the PN stored at 25°C and 40°C. Neither globule size pattern, nor any other physicochemical characteristic evaluated appeared to be considerably altered in any of the analyzed formulations even after 7 days of storage at 5°C. Globule size in all the PN studied was consistent with the established limit, below 500 nm by DLS measurement, and PFAT5 was below 0.05% under all storage temperatures., Conclusion: Concomitant presence of trace elements and vitamins in the same neonatal formulation did not alter the evaluated aspects of stability.

A new method for the quantification of butenafine hydrochloride (BTF) present in the main skin layers was validated and a study conducted with the aim of analyzing the penetration and/or the permeation of the drug. The quantification was performed by liquid chromatography. To evaluate the specificity of the method, the influence of the components of the skin was analyzed, as well as the skin in contact with the excipient ingredients. Linearity was assessed with concentrations in the range of 0.1-10 μg/mL (r(2) = 0.9999) and ANOVA showed non-significant linear deviation (p > 0.05). Adequate results were obtained for repeatability, intra-day precision and accuracy. The obtained values for the limit of quantification and the limit of detection were 68.4 and 17.7 ng/mL, respectively. Also, a comparative study of BTF cutaneous penetration through porcine skin was performed applying two different formulations: Tefin, present in the Brazilian market, and Lotrimin Ultra(®) , available in the American market. No statistical difference was found in the skin (epidermis plus dermis) and in the epidermis (p > 0.05), although in the dermis the difference was significant (p < 0.05). During the experimental period (8 h), no drug permeation from either formulation was detected in the receptor fluid., (Copyright © 2011 John Wiley & Sons, Ltd.)

A stability-indicating liquid chromatography method for the determination of the antifungal agent butenafine hydrochloride (BTF) in a cream was developed and validated using the Plackett-Burman experimental design for robustness evaluation. Also, the drug photodegradation kinetics was determined. The analytical column was operated with acetonitrile, methanol and a solution of triethylamine 0.3% adjusted to pH 4.0 (6:3:1) at a flow rate of 1 mL/min and detection at 283 nm. BTF extraction from the cream was done with n-butyl alcohol and methanol in ultrasonic bath. The performed degradation conditions were: acid and basic media with HCl 1M and NaOH 1M, respectively, oxidation with H(2)O(2) 10%, and the exposure to UV-C light. No interference in the BTF elution was verified. Linearity was assessed (r(2) = 0.9999) and ANOVA showed non-significative linearity deviation (p > 0.05). Adequate results were obtained for repeatability, intra-day precision, and accuracy. Critical factors were selected to examine the method robustness with the two-level Plackett-Burman experimental design and no significant factors were detected (p > 0.05). The BTF photodegradation kinetics was determined for the standard and for the cream, both in methanolic solution, under UV light at 254 nm. The degradation process can be described by first-order kinetics in both cases.

Background: The objective of this work was to study the vitamins B1, B2, B6 and C stability in a pediatric formulation containing high amounts of calcium in the presence of organic phosphate, amino acids, glucose, sodium chloride, magnesium sulfate, pediatric vitamins and trace elements under different conditions using developed and validated analytical methods., Methods: The study was carried out during 72 h with formulations packaged in recommended storage temperature (4°C) and 25°C, with and without photoprotection., Results: The results showed that the methodologies used for assessing the chemical stability of vitamins B1, B2, B6 and C in the formulation were selective, linear, precise and accurate. The vitamins could be considered stable in the formulation during the three days of study if stored at 4°C. When stored at 25°C vitamin C presented instability after 48 h., Conclusion: The pediatric formulation containing high amount of calcium in the presence of organic phosphate, amino acids, glucose, sodium chloride, magnesium sulphate, pediatric vitamins and trace elements packaged in bag-type trilaminate presented a shelf life of the 72 h, when maintained under refrigeration, between 2°C and 8°C. This shelf life was measured considering the vitamins studied. Further studies are needed including all the vitamins present in this formulation.

A simple, specific, fast and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the simultaneous analysis of delapril (DEL) and manidipine (MAN) from their combination formulation was developed and validated using fesoterodine as the internal standard (IS). The LC-MS/MS method was carried out on a Luna C8 column (50 × 3.0 mm i.d., 3 µm) with a mobile phase consisting of methanol and 10 mmol L(-1) ammonium acetate (90 : 0, v/v), run at a flow rate of 0.25 mL min(-1). The mass spectrometry method was performed employing positive electrospray ionization operating in multiple reaction monitoring mode, monitoring the transitions of m/z 453.1 → 234.1 for DEL, m/z 611.1 → 167.0 for MAN and m/z 412.2 → 223.0 for IS. The total analysis time was 3 min and the method was linear in the concentration range of 6-1080 ng mL(-1) and 2-360 ng mL(-1) for DEL and MAN, respectively. Parameters investigated for the method validation, such as the specificity, linearity, precision, accuracy and robustness, gave results within the acceptable range. Moreover, the proposed method was successfully applied for the simultaneous determination of DEL and MAN and the results were compared to validated liquid chromatography and capillary electrophoresis methods showing non-significant differences (P = 0.9).

Background: Preterm infants need high amounts of calcium and phosphorus for bone mineralization, which is difficult to obtain with parenteral feeding due to the low solubility of these salts. The objective of this study was to evaluate the physicochemical compatibility of high concentrations of calcium associated with organic phosphate and its influence on the stability of AIO admixtures for neonatal use., Methods: Three TPN admixture formulas were prepared in multilayered bags. The calcium content of the admixtures was adjusted to 0, 46.5 or 93 mg/100 ml in the presence of a fixed organic phosphate concentration as well as lipids, amino acids, inorganic salts, glucose, vitamins and oligoelements at pH 5.5. Each admixture was stored at 4 degrees C, 25 degrees C or 37 degrees C and evaluated over a period of 7 days. The physicochemical stability parameters evaluated were visual aspect, pH, sterility, osmolality, peroxide formation, precipitation, and the size of lipid globules., Results: Color alterations occurred from the first day on, and reversible lipid film formation from the third day of study for the admixtures stored at 25 degrees C and 37 degrees C. According to the parameters evaluated, the admixtures were stable at 4 degrees C; and none of them presented precipitated particles due to calcium/phosphate incompatibility or lipid globules larger than 5 mum, which is the main parameter currently used to evaluate lipid emulsion stability. The admixtures maintained low peroxide levels and osmolarity was appropriate for parenteral administration., Conclusion: The total calcium and calcium/phosphorus ratios studied appeared not to influence the physicochemical compatibility and stability of AIO admixtures.

The dissolution test for oral dosage forms has recently widened to a variety of special dosage forms such as suspensions. For class II drugs, such as nimesulide (NMS), this study is very important because formulation problems may compromise drug bioavailability. In the present work, tests with four brands of commercially available NMS (RA, TS, TB, and TC) have been performed in order to study their dissolution at different conditions. The suspensions have been characterized relatively to particle size, pH, and density besides NMS assay and the amount of drug in solution in the suspension vehicles. The dissolution study was conducted using the following media: simulated intestinal fluid, pH 6.8, containing polysorbate 80 (P80) or sodium lauryl sulfate (SLS); phosphate buffer, pH 7.4, with P80 and aqueous solution of SLS. Concerning the quantitative analysis, the UV-VIS spectrophotometry could have been used in substitution to high-performance liquid chromatography since the methodology had been adequately validated. The influence of the drug particle size distribution was significant on the dissolution profiles of NMS formulations, confirming to be a factor that should be strictly controlled in the development of oral suspensions.

Microencapsulation is essential to preserve biological activity of ascorbic acid (AA) and pea protein has not been used as a carrier in such processes. This work aimed to produce microparticles by a spray-drying process using pea protein (PPC) as wall material of AA and evaluate the retention of the core by HPLC, overall morphology SEM, size distribution by light scattering and release kinetics. Carboxymethylcellulose (CMC) and blends with maltodextrin (M) were produced for comparative analyses. The yields were compatible with the applied technology and the retention was above 84% for all materials. The PPC microparticles presented irregular and rough surfaces, CMC produced a regular and smooth surface and agglomeration was more intense in microparticles with M. Mean particle diameters were all below 8 microm. The microparticle release rates were lower than those with free AA, being best correlated to the Higuchi kinetic model. These results support the utilization of PPC for microencapsulation of AA.

A validated method for on-line solid-phase extraction coupled with high-performance liquid chromatography tandem mass spectrometry (SPE-HPLC-MS-MS) is described for the quantification of bromazepam in human plasma. The method involves a dilution of 300 muL of plasma with 100 muL of carbamazepine (2.5 ng/mL), used as internal standard, vortex-mixing, centrifugation, and injection of 100 muL of the supernate. The analytes were ionized using positive electrospray mass spectrometry then detected by multiple reaction monitoring (MRM). The m/z transitions 316-->182 (bromazepam) and 237-->194 (carbamazepine) were used for quantification. The calibration curve was linear from 1 ng/mL (limit of quantification) to 200 ng/mL. The retention times of bromazepam and carbamazepine were 2.6 and 3.2 minutes, respectively. The intraday and interday precisions were 3.43%-15.45% and 5.2%-17%, respectively. The intraday and interday accuracy was 94.00%-103.94%. This new automated method has been successfully applied in a bioequivalence study of 2 tablet formulations of 6 mg bromazepam: Lexotan(R) from Produtos Roche Químicos e Farmacêuticos SA, Rio de Janeiro, Brazil (reference) and test formulation from Laboratórios Biosintética Ltda, São Paulo, Brazil. Because the 90% CI of geometric mean ratios between reference and test were completely included in the 80%-125% interval, the 2 formulations were considered bioequivalent. The comparison of different experimental conditions for establishing a dissolution profile in vitro along with our bioavailability data further allowed us to propose rationally based experimental conditions for a dissolution test of bromazepam tablets, actually lacking a pharmacopeial monograph.

In a previous study aimed to compare the bioavailability of two levothyroxine tablets, we found a good relation between their pharmacokinetics parameters and dissolution profiles, employing the USP dissolution conditions in use at that time (24th edition). Despite the formulations were considered bioequivalent, the test product presented values of AUC and concentrations at steady-state significantly lower (about 10%) than the reference ones. The purpose of the present study was to evaluate if the actual pharmacopeial conditions (with alterations introduced in the first supplement of USP 24) would also allow a good correlation between bioavailability and dissolution data. The partial AUCs were correlated with cumulative levothyroxine amount dissolved at three different times, for each dissolution condition. Employing the old method, test tablets had a slower dissolution rate than the reference ones, resulting in a quite good multiple level C in vitro/in vivo (IVIV) correlation. On the other hand, the very fast dissolution profiles obtained in the actual condition lead to a worse IVIV correlation. Present work indicates that the mild conditions proposed in the older US Pharmacopeia were better than the actual in order to discriminate dissolution profiles of levothyroxine tablets which present subtle, but significant, differences in their pattern of absorption.

The aim of the present study was to evaluate the bioequivalence and therapeutic equivalence of the two most commonly prescribed L-thyroxine (monsodium L-thyroxine hydrate, CAS 25416-65-3) formulations in Brazil in patients treated for hypothyroidism. Twenty-four patients received 100 micrograms L-thyroxine daily of either Puran T4 (test) or the Brazilian reference formulation (reference) during 42 days, in a two-period crossover design. Serum samples obtained over a 24-h interval were analyzed for their total T4 concentration by a chemiluminescent immunoassay. Content and uniformity of the tablets and dissolution studies were also assessed according to USP 24 monograph using an isocratic HPLC-UV system and a rotating-paddle method. The mean pharmacokinetic parameters for total T4, expressed as geometric means (CV), for the test and reference were, respectively: Cmax (microgram/dl) 9.8 (14.3%) and 10.8 (14.9%); AUC0-24 h (microgram/dl.h) 206.8 (13.9%) and 230.4 (14.9%). Median values (90% CI) for Tmax (h) were 3 (2-3) and 2 (2-4) for the test and reference, respectively. 90% CI for ratios of LogCmax and LogAUC0-24 h were 86.6-94.9 and 86.3-93.4, respectively. Although the test exhibited values of Cmax and AUC0-24 h around 10% lower than the reference, these formulations must be considered bioequivalent since the 90% CI for both Cmax and AUC0-24 h mean ratio were within the 80-125% interval as proposed by the US Food and Drug Administration and the Brazilian legislation. TSH dosages within the normal range further support therapeutic equivalence between the two formulations. Dissolution data were roughly in agreement with in vivo results since both formulations comply with the USP dissolution criteria although the test tablets had a slower dissolution rate than the reference tablets. As a conclusion, the two oral formulations of L-thyroxine are both bioequivalent and therapeutically equivalent although presenting a small difference in their extent of absorption. Noteworthy, the dissolution profiles of the tablets correlate well with their bioavailability in the present experimental conditions.

The purpose of the present work was to study the in vitro distribution of acyclovir in human skin layers after iontophoresis, applied in order to increase the amount of drug in the basal epidermis, site of Herpes simplex infections. Experiments were done with Franz diffusion cells applying, as donor, acyclovir solutions (pH values: 3.0 and 7.4) or a commercial cream. Quantification of drug at different skin depths was performed by horizontal slicing of frozen skin, and drug extraction and analysis by high-performance liquid chromatography. Seven h of transdermal iontophoresis (0.5 mA cm-2 induced an accumulation of acyclovir in epidermis and dermis ranging from 80 to 150 micrograms cm-3, characterized by homogeneous distribution of the drug in skin layers. After short current application time (30 min) however, the concentration profile of drug in skin was not significantly different from the obtained after seven h of passive diffusion, employing pH 3.0 donor solution. After 30 min of iontophoresis, the acyclovir reservoir on the skin was maintained for up to five h producing a dramatic increase of drug concentration in skin, evening out over 80 micrograms cm-3 until a depth of 300 micrograms. Acyclovir can be accumulated at target site more quickly and maintained at higher level through application of a iontophoretic pulse and by keeping the drug reservoir on skin.

This paper describes an assay procedure for acyclovir quantification in human skin after in vitro transdermal transport experiments. The procedure employs warm distilled water for acyclovir (ACV) extraction and high-performance liquid chromatography (HPLC) as analytical method. The procedure has good reproducibility, sensitivity and specificity, resulting in a reliable method for biopharmaceutical studies of ACV distribution in skin tissue. The chromatographic conditions set up, using distilled water as mobile phase, makes the analytical procedure simple and easy to perform.

Salmon calcitonin (sCt) was administered by transdermal iontophoresis in rabbits, using a new drug reservoir assembled directly on the skin, based on a dry disc containing sCt to be dissolved at the application site. The hypocalcemic effect was taken as a measure of the pharmacodynamic response. In rabbits, the results obtained show that salmon calcitonin skin penetration by iontophoresis, using pulsatile current of 0.8 mA/cm2 on a reservoir containing 100 IU/Kg of sCt, was governed by the quantity of electric charge applied, mimicking the hypocalcemic response of 10 IU/Kg intravenous administration.

Purpose: The aim of the work was to study iontophoretic transdermal administration of salmon calcitonin (sCt) in rabbits, with particular attention to drug reservoir composition. A dry sCt disc, to be dissolved on the application site, was used for preparing the reservoir for transdermal iontophoresis. As a reference drug reservoir, a pad wetted with drug solution was used., Methods: Experiments were done in rabbits depositing 100 IU of salmon calcitonin on skin and applying anodal iontophoresis. Serum calcium concentration was measured during iontophoresis, passive diffusion and after i.v. administration. Parameters such as pH value and reservoir type were examined., Results: Transdermal iontophoresis of sCt elicited a decrease in the serum calcium level, whereas, in the absence of electric current, no significant fall was measured. Using the reservoir prepared from drug solution, anodal iontophoresis at pH 4.2 was more effective than at pH 7.4, probably due to higher sCt net positive charge. Using the reservoir prepared from dry disc, similar kinetics and extent of drug effect were observed at both pH values. The reservoir prepared from solid drug deposit concentrated sCt next to the skin., Conclusions: Anodal iontophoresis for transdermal calcitonin administration shows therapeutical applicability. The type of reservoir is an important parameter affecting sCt transdermal iontophoresis.

Purpose: Iontophoresis was employed for enhancing the transdermal delivery of acyclovir through nude mouse skin in vitro, with the aim of understanding the mechanisms responsible for drug transport, in order to properly set the conditions of therapeutical application., Methods: Experiments were done in horizontal diffusion cells, using as donor a saturated solution of acyclovir at two different pH values (3.0 and 7.4). Different electrical conditions (current density and polarity) were employed., Results: At pH 3.0, acyclovir anodal transport was due to electrorepulsion, since acyclovir was 20% in the protonated form. In acyclovir anodal iontophoresis at pH 7.4 the main mechanism involved was electroosmosis, since the drug was substantially unionized and the negative charge of the skin at this pH caused the electroosmotic flow to be from anode to cathode. In the case of cathodal iontophoresis at pH 3.0, acyclovir transport was enhanced approx. seven times, due to the presence of an electroosmotic contribution caused by the reversal of the charge of the skin. At pH 7.4 during cathodal iontophoresis acyclovir transport was not enhanced because the electroosmotic flow was in the opposite direction, compared to drug electric transport, i.e. anode to cathode. The increased skin permeability caused by current application was demonstrated to be less important than electrorepulsion and electroosmosis., Conclusions: Anodal iontophoresis shows potential applicability for enhancing acyclovir transport to the skin, considering that both electric transport and electroosmosis can be used by appropriately setting the pH of the donor.