Darunavir Ethanolate Research Articles

D-α-tocopheryl polyethylene glycol succinate-decorated dual drug-loaded lipidic nanocarriers: A strategic approach for targeting lymphatic uptake and p-gp efflux modulation to enhance oral bioavailability in HIV-1 viral reservoirs

The toxicity potential and inadequate bioavailability of antiretroviral therapy restrict their effectiveness in completely eradicating HIV-1 from viral reservoirs, as the drugs encounter difficulties in accessing these reservoirs. This study presents a combinatorial d-α-tocopheryl polyethylene glycol succinate (TPGS)-decorated nano-structured lipid carrier of Etravirine (ETR) and Darunavir Ethanolate (DRVE) that was formulated, optimized, and characterized to achieve synergistic effects against HIV-1 infection. The effectiveness of the combinatorial approach was evaluated by in silico studies, and their docking score and free binding energy showed that both drugs demonstrate synergistic effects against their respective enzymes. The cellular research of ED (ETR and DRVE) in TMZ-b1 cell lines also showed that the best ratio of ETR and DRVE (1:6.5 μg/mL) produced a combined effect. The formulation, termed ED-TPGS-NLCs, underwent optimization using central composite rotational design (CCRD) through a modified emulsification process, followed by characterization based on globule size, polydispersity index (PDI), percentage entrapment efficiency, percentage drug loading, and transmission electron microscopy (TEM) analysis. In vitro release studies demonstrated a notable improvement of drug release from the optimized lipid nanosystem, conforming to the Korsmeyer-Peppas kinetics model. Besides, the in-vitro studies, the intestinal permeation enhancement study, the intestinal depth analysis, and the pharmacokinetic assessment of the optimized formulation in Wistar rats were performed to improve permeation and increase plasma drug concentration. ED-TPGS-NLCs also stopped HIV-1 infection in TMZ-b1 cell lines by 50 %, and the lowest concentration needed to do this was about 58 times lower than purified ED suspension. These studies ensure that ED-TPGS-NLCs were taken up due to the formation of chylomicrons and inhibit the p-gp efflux system by TPGS and solutol to inhibit the first-pass hepatic metabolism, which helps to improve the intestinal permeation and enhance plasma drug concentration, leading to enhanced oral bioavailability of ED. Hence, the improved bioavailability of ED in the nanoformulation also enhanced the anti-retroviral efficacy and improved the safety margins of ED-TPGS-NLCs. These findings can also reduce the side effects of high doses of antiretroviral drugs.

Enhancing lymphatic uptake of Darunavir Ethanolate through TPGS-enriched lipid nanocarriers for oral delivery

Darunavir ethanolate (DRVE) is a promising molecule against wild-type protease inhibitors in human immunodeficiency virus (HIV) infection. It has a 37% oral bioavailability. In this study, the d-alpha-Tocopheryl Polyethylene Glycol 1000 Succinate (TPGS)-decorated DRVE-loaded lipid nanocarriers (DRVE-TPGS-LNCs) were optimized via central composite rotatable design using a modified solvent emulsification method. Various characteristic features of DRVE-TPGS-LNCs, such as globule size, polydispersity index, zeta potential, % entrapment efficiency, and % drug loading, were determined and found to be 116.5 ± 2.52 nm, 0.269 ± 0.0221, −10.8 ± 0.265 mV, 94.31% ± 10.75%, and 8.79% ± 0.937%, respectively. The spherical shape was evaluated by cryo-transmission electron microscopy. In-vitro study demonstrated % cumulative drug release of 81.70% ± 7.35% at pH 1.2 and 73.03% ± 7.17% at pH 6.8 after 12 h of drug release study. Moreover, the intestinal permeation and confocal microscope studies revealed approximately two-fold increased permeation in optimized nanoformulation across the gut sac compared to a drug suspension, and the inhibition of p-glycoprotein efflux was also confirmed by obtaining a minimum efflux ratio (0.253) of the optimized formulation compared to a drug suspension. The pharmacokinetic study demonstrated that the relative bioavailability of TPGS-DRVE-LNCs and drug suspension were found to be 19.85 ± 1.75 and 8.72 ± 0.753 μg.h/mL, respectively, which is approximately 2.25-fold more than the drug suspension which means approximately threefold increase in oral bioavailability compared to the drug suspension. The increased bioavailability in optimized DRVE-TPGS-LNCs was due to improved lymphatic uptake and p-glycoprotein efflux inhibition. Thus, the improved bioavailability of DRVE can meet the need to reduce the high dose of antiretroviral drugs to minimize the peripheral adverse reactions associated with the dose-related burden.

POLYMORPHIC STABILITY EVALUATION OF DARUNAVIR AND RITONAVIR IN FIXED DOSE DRUG COMBINATION PRODUCT BY X-RAY DIFFRACTION AND DIFFERENTIAL SCANNING CALORIMETRY TECHNIQUES

Evaluation of polymorphism for active ingredients and its stability during manufacturing and storage is a critical task for pharmaceutical firms. For solubility enhancement of ritonavir, hot melt extrusion process was selected through conversion of commercially available crystalline form to amorphous. XRD method (with normal and slow scan) and DSC method was considered for polymorphic evaluation in tablet formulation comprising darunavir 800 mg and ritonavir 100 mg. Specificity and precision of XRD method was justified by no interference of placebo and 5.6% RSD, respectively. The observed correlation coefficient of 0.9991 and average recovery of 98.7% (range 91.5 to 105.1) depicts linearity and recovery power of developed method. Studied DSC method shows exotherm for ritonavir crystalline form II at 127.4 °C and darunavir ethanolate at 105.7 °C, which were specific and sensitive to assess polymorphic form of available drug substances in ritonavir blend as well as in tablet formulation during release and stability period.

Drug Likeness Screening and Evaluation of Physicochemical Properties of Selected Medicinal Agents by Computer Aided Drug Design Tools

The current drug research techniques, combinatorial synthesis and high throughput screening, help in obtaining and pre-evaluating thousands of compounds in a short duration of time. Among the selected compounds, the best hit molecule becomes the lead, and the drug likeness property helps to optimize the selection. Lipinski's Rule of five is the most widely used filter in pre-evaluation of compounds, according to Lipinski's Rule of five the molecule with poor oral absorption and low permeation has the molecular weight >500, ClogP >5, hydrogen bond acceptor more than 10 and hydrogen bond donor more than 5. To evaluate the Rule of Five, the top selling drugs of 2018-19 were selected. Among the selected 56 drugs, 12 drugs (tenofovir disoproxil, sofosbuvir and ledipasvir, sofosbuvir/velpatasvir, trastuzumab, glecaprevir, pegtilgrastion, Lumacaftor/ ivacaftor, fluticasone propionate and salmetarol, elbasvir, darunavir ethanolate, Linagliptin, metformin, fluticasone furoate and vilanterol) did not fulfill the rule. The rule is very useful in selecting the better compound in chemolibraries, but should be used carefully and with certain criterion due to possible exclusion of promising compounds.

Comparative Study of Methods for the Pharmaceutical Preparation and Effectiveness of Darunavir Ethanolate Compositions with Mesoporous Carriers and Polymer Solid Dispersions

Comparative Study of Methods for the Pharmaceutical Preparation and Effectiveness of Darunavir Ethanolate Compositions with Mesoporous Carriers and Polymer Solid Dispersions

Influence of Water-Soluble Pharmaceutically Acceptable Polymers on the Solubility of Darunavir and Darunavir Ethanolate

The solubility of darunavir (D) and darunavir ethanolate (DE) at physiological pH values and temperatures in solutions of pharmaceutically acceptable polymeric excipients was studied to substantiate dosage form formulations. Both substances were determined to dissolve better in HCl at pH 1.2 and 37°C. Use of PVP 90 in peroral dosage form formulations was shown to be unadvisable because of a negative effect on the solubility of the substances. The effect of Soluplus® on the solubility of D and DE was complicated by multifactor and heteropolar effects. PVP 12 – 30, HPMC 606, and Kollidon® VA64 HPC had positive effects on the solubility of D and DE substances. The best results were obtained for Eudragit® E PO solutions.

The Effect of Solvents and Drying Temperature on the Physicochemical Properties of Darunavir and Darunavir Ethanolate Substances

Introduction. Usage of aggressive conditions (solvents, high temperature, etc.) in a dosage form manufacturing can lead to a change in the properties of a pharmaceutical substance. Darunavir (D) amorphous and darunavir ethanolate (DE) crystalline both have poor solubility, ability to pseudopolymorphism and are sensitive to high temperatures.Aim. Study the effect of solvents and drying temperature on the physicochemical properties of D and DE substancesMaterials and methods. D (Mylan Laboratories Limited), DE (Mylan Laboratories Limited), D (reference standard) 99,9 % (MSN Pharmachem Private Limited). D and DE weighed quantity was suspended in one of the solvents via magnetic stirrer and was dried via universal oven. Powder X-ray diffraction of dried samples was carried out via automatic powder diffractometer. Using DSC thermal properties of the samples were studied. Crystalline samples were examined using IR spectroscopy. Identification of D and DE was performed by HPLC method.Results and discussion. This article summarizes study results of the investigation of various solvents and drying temperature influence on the physicochemical properties of D and DE are presented. The impact of solvent type and drying temperatures in physicochemical properties of the APIs was studied by X-ray powder diffraction, differential scanning calorimetry, IR spectroscopy and HPLC methods. It was shown, that solvent type and drying temperatures can result in the presence of crystalline D solvates or amorphous D.Conclusion. To obtain the final drug containing as an API amorphous D, which perform better dissolution, one of the enlisted solvents can be used: dichloromethane, chloroform and heptane. In such case the intermediate product drying should be performed at not exceeding the solvent boiling point temperatures. In case of ethanol, methanol, acetone and tetrahydrofuran drying phase can be performed at temperatures, that are higher than melting points of obtained pseudopolymorphs. For the utilization of DE as an API only ethanol usage is efficient and drying temperature should not exceed 73.4 °C.

Chemo-metric assisted UV-spectrophotometric methods for simultaneous estimation of Darunavir ethanolate and Cobicistat in binary mixture and their tablet formulation

Darunavir ethanolate (DRV) and Cobicistat (CBS) is a combination of antiretroviral drugs used for the treatment of human immunodeficiency virus (HIV) infections. Two Chemo-metric assisted UV-spectrophotometric methods were developed for simultaneous estimation of DRV and CBS in tablet dosage form, namely; partial least square (PLS) and Classical least square method (CLS). The proposed methods were successfully applied for simultaneous determination of DRV and CBS in a laboratory mixture and their tablet formulation to achieve maximum sensitivity and lowest error. The applied methods were validated as per ICH guidelines and found to be linear in the concentration range of 5-30μg/mL for DRV and 5-30μg/mL for CBS. The developed methods were statistically comared with reported UPLC method where no significant difference was found relating to both accuracy and precision. Thus, the proposed methods can be effectively utilized for the routine quality control assessment of these drugs in commercial tablet dosage form.

A stress degradation kinetic study of darunavir ethanolate bulk and tablet dosage form by uv spectrophotometry

This work was aimed to develop and validate simple, rapid and economical kinetic degradation method for the estimation of Darunavir ethanolate in bulk and tablet dosage form. The spectrum was determined at 267.40nm by using methanol as a solvent. The developed method was validated as per ICH guideline. Accuracy was obtained between 97.18 - 101.12%. LOD and LOQ was found to be 0.52μg/ml and 1.60μg/ml respectively. Darunavir ethanolate was subjected to acid, base, oxidation, photolytic and thermal degradation and the amount of degradation is in the range 10 - 40%. Results confirmed that this method can be employed for the estimation of Darunavir ethanolate in presence of its degradation product in tablet dosage form.

An improved and robust scale-up process aided with identification and control of critical process impurities in darunavir ethanolate

A robust and safe industrial process, including five isolations and drying steps for widely prescribed anti-HIV (protease inhibitor) drug darunavir ethanolate 2, has been developed. A salient feature of this process is the development of procedures enabling the efficient synthesis of multi-kilogram quantity of darunavir ethanolate, and process demonstrations through plant scale preparation are offered where darunavir molecule has been prepared with overall > 70% chemical yield and > 99.8% purity without involving any purification procedure(s), with all possible process impurities below than the desired limit (not more than 0.08%) were isolated, synthesized and characterized. The developed process is entirely robust, very efficient and demonstrated up to kilograms scale.

ANALYTICAL METHOD DEVELOPMENT FOR THE ESTIMATION OF DARUNAVIR BY DIAZOTIZATION AND COUPLING BY VISIBLE SPECTROPHOTOMETRY

Objectives: This proposed work describes simple and extraction free visible methods for the estimation of darunavir ethanolate (DNV) in bulk and tablet formulations. The methodology involves diazotization of DNV with nitrous acid followed by coupling with chromotropic acid (CA) (Method A)/ Bratton Marshall Reagent (Method B)/α-naphthol (Method C) to form colored products.Methods: All the methods were developed using a PerkinElmer (LAMBDA 25) UV–Visible spectrophotometer interfaced with UV Win lab software and 1 cm quartz cells. Results: Spectrophotometrically, DNV is estimated at 520 nm, 544 nm, and 464 nm for the reddish-pink color produced by CA, dark violet color with NED, and dark-greenish yellow with α-naphthol, respectively. The linear relationship was observed between absorbance and the corresponding concentration of drug in the range of 100–350 μg/mL, 10–100 μg/mL, and 10–60 μg/mL for Methods A, B, and C, respectively.Conclusion: The colorimetric methods were extensively validated as per the ICH guidelines. The developed methods were proven to be more accurate and precise by the statistical analysis.

Single crystal fragmentation: Visualizing breakage model performance for pharmaceutical processes

The development of accurate crystal breakage kinetics is paramount to extending the capability of optimization and modelling methods within the pharmaceutical drug-product sector. However, there exists little experimental data on the performance on typical breakage models when applied to small-scale pharmaceutical crystals. Therefore, experimental investigations of crystal impacts on a steel target were carried out across a range of crystal sizes and impact velocities in order to determine critical breakage parameters for a given model. Direct observation of each impact was captured via a high-speed Shadowgraphy imaging technique, capable of 1μm crystal size resolution and up to a 125,000 frames-per-second capture rate. Crystals of Darunavir Ethanolate between a nominal 100→500μm size range were subject to impact velocities of 1→10m/s normal to the target surface. Furthermore, upon collision crystals were classified into one of three breakage modes: intact (Mode I), chipped (Mode II), or split/disintegrated (Mode III). The resulting data was then combined with a post-processing methodology utilizing a bi-variate histogram function and Kernel Density Estimation (KDE). This in turn enabled the extraction of experimentally observed crystal failure velocities, breakage modes, and selection functions detailing how crystal breakage varies with both crystal size and impact velocity. This data was then used to evaluate the performance of current literature in describing crystal breakage within pharmaceutical applications. Furthermore, the methodology implemented in this work can be applied to a range of particulate systems, yielding a flexible tool for pharmaceutical modelling capabilities.

Investigating the Physicochemical Stability of Highly Purified Darunavir Ethanolate Extracted from PREZISTA® Tablets.

Understanding physicochemical stability of darunavir ethanolate is expected to be of critical importance for the development and manufacturing of high-quality darunavir-related pharmaceutical products. However, there are no enabling monographs for darunavir to illustrate its solid-state chemistry, impurity profile, and assay methods. In addition, the US Pharmacopeia reference standard of darunavir is still not commercially available. It has been also challenging to find reliable vendors to obtain highly purified darunavir ethanolate crystals to conduct the physicochemical stability testing. In the present research, we developed a straightforward and cost-effective approach to extract and purify darunavir ethanolate from PREZISTA® tablets using reverse-engineering and crystallization. Using these highly purified crystals, we thoroughly evaluated the potential risks of degradation and form conversions of darunavir ethanolate at stressed conditions to define the manufacturing and packaging specifications for darunavir-related products. Amorphization was observed under thermal storage caused by desolvation of darunavir ethanolate. The ethanolate-to-hydrate conversion of darunavir was observed at high relative humidity conditions. Moreover, acid/base-induced degradations of darunavir have been investigated herein to determine the possible drug-excipient compatibility issues in formulations. Furthermore, it is of particular interests to allow the production of high-quality darunavir-ritonavir fixed dose combinations for marketing in Africa. Thus, a validated HPLC method was developed according to ICH guideline to simultaneously quantify assays of darunavir and ritonavir in a single injection. In summary, the findings of this study provide important information for pharmaceutical scientists to design and develop reliable formulations and processings for darunavir-related products with improved stability.

Analytical Control of Darunavir Tablet Dosage Forms. Part I.

Purposeful development of special methods to be included into pharmacopoeial monographs on the quality of darunavir (DRV) preparations is shown to allow the use of expensive imported standard samples (SSs) for drug analysis to be avoided. Part I of the present work describes these methods and their validation. UV spectrophotometry is proposed in the “Authenticity” section for detecting DRV by the coincidence of extrema in the spectrum of an aqueous MeOH extract (pH 9) with well-known peaks in the spectrum of deprotonated DRV at 230 nm (minimum) and 267 nm (maximum). This section also requires GC analysis for solvated EtOH, the presence of which indicates that DRV ethanolate was used as the active pharmaceutical ingredient (API) and the absence of which, that non-solvated amorphous DRV was used. The significant increase in the accuracy of the quantitative determination and the sharp reduction in its duration allow the use of SSs to be avoided. For this reason, the “Dissolution” section proposes determining the DRV concentration in the dissolution medium (pH 3) by spectrophotometry using a method that requires experimental determination of the specific absorption coefficient $$ \left({A}_{1\mathrm{cm}}^{1\%}\right) $$ of DRV solutions in this medium at the maximum (267 nm). The established value $$ \left({A}_{1\mathrm{cm}}^{1\%}\right. $$ = (393.4 ± 2) cm–1 was a physicochemical constant with a relative standard deviation RSD < 1% at confidence level α = 0.05.

HPLC Estimation, Ex vivo Everted Sac Permeability and In Vivo Pharmacokinetic Studies of Darunavir.

Darunavir ethanolate (DRV) is an efficient protease inhibitor (PI) used in the treatment of human immunodeficiency virus (HIV) type-1 patients. An isocratic reversed-phase HPLC method was developed to monitor concentration of darunavir in in vitro intestinal fluid samples in everted sac absorption model in the presence of bioenhancers, viz., piperine, quercetin, naringenin. The method was validated and successfully applied to everted sac and pharmacokinetic studies in rats. The absorption profiles of DRV and apparent permeability coefficients were determined. The proposed method was found to be simple, rapid, robust and selective and was applied for continuous ex vivo monitoring of DRV in everted sac absorption studies. Of the three bioenhancers screened at different concentrations, piperine caused highest and significant 1.5-fold increase in apparent permeability of DRV across everted sac tissue. Further, co-administration of piperine significantly increased the maximum plasma concentration of DRV by 1.18-fold confirming the enhancement in its absorption.

A Novel RP- HPLC Method with Core Shell Tecnology Column for Assay Analysis of Darunavir Ethanolate in Tablet Dosage Form

A Novel RP- HPLC Method with Core Shell Tecnology Column for Assay Analysis of Darunavir Ethanolate in Tablet Dosage Form

Методы аналитического контроля таблетированных лекарственных форм дарунавира. Сообщение I

Показано, что целенаправленная разработка специальных методик для включения в монографии, регламентирующие качество препаратов дарунавира (ДРВ), позволяет избежать использования в процессе анализа дорогостоящих импортных стандартных образцов (СО). В сообщении I представлены такие методики и результаты их валидации. В разделе «Подлинность» предложено использовать спектрофотометрическое подтверждение наличия ДРВ в препарате по факту совпадения абсцисс экстремумов УФ-спектра полученной из него водно-метанольной вытяжки (pH 9) с известными для растворов непротонированного ДРВ значениями: 230 нм (минимум), 267 нм (максимум). В разделе предусмотрено также обязательное определение в препарате сольватного этанола методом ГЖХ. Его присутствие указывает на то, что в качестве активной фармацевтической субстанции (АФС) использовался ДРВ этанолят, а отсутствие указывает на то, что в качестве АФС использовался несольватированный аморфный ДРВ. Заметное повышение точности количественного определения и резкое сокращение его продолжительности обеспечивает отказ от применения СО. По этой причине в разделе «Растворение» концентрацию ДРВ в среде растворения при pH 3 предложено определять спектрофотометрически по методике, применение которой потребовало экспериментального определения величины удельного показателя поглощения () растворов ДРВ в этой среде в максимуме при 267 нм. Найденное значение = (393,4 ± 2) см–1 является физико-химической константой, его относительная ошибка (RSD) &lt; 1 % при степени надёжности α = 0,05.

Issues Related to Patent Protecton of Darunavir and its Analogs

It is shown that prolongation of patent protection for the antiviral drug darunavir by patenting it again as darunavir ethanolate is illegal. The claims of the successor patent “Pseudopolymorphic forms of a HIV protease inhibitor,” WO 2003106461 A2, are analyzed and shown to be invalid from both scientific and ethical viewpoints. It is established that darunavir was initially used as its ethanolate. Darunavir is one of the best protease inhibitors of both types of HIV. Currently, not only darunavir ethanolate but also amorphous darunavir (unsolvated) are used as active pharmaceutical ingredients with INN darunavir. This was confirmed by x-ray diffraction studies. Justification for the orderly invalidation of this patent in the RF was given.

Characterization of Darunavir: Β-Cyclodextrin complex and Comparison with the Forms of Darunavir Ethanolate and Hydrate

Characterization of Darunavir: Β-Cyclodextrin complex and Comparison with the Forms of Darunavir Ethanolate and Hydrate Darunavir, a protease inhibitor used in the treatment of HIV infection, presents solubility problems and so it was complexed with β-cyclodextrin. Complexation increased the solubility of the drug in more than 28 times compared to the free drug. Then, it is important to characterize this recent advance of pharmaceutical technology as well as its comparison with the forms of free darunavir, hydrate and ethanolate. Spectroscopy in the infrared (IR), thermal techniques of thermogravimetry (TG) and differential scanning calorimetry with heat flux (DSC) and X-ray powder diffraction were elected for the characterization of darunavir:β-cyclodextrin complex and comparison with darunavir ethanolate and darunavir hydrate.

Powder X-ray diffraction of darunavir ethanolate, C27H37N3O7S(C2H5OH)

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