Daclatasvir Dihydrochloride Research Articles

Molecularly imprinted polymer/reduced graphene oxide‒based carbon‒paste sensor for highly sensitive determination of the anti‒HCV drug daclatasvir dihydrochloride

The recent discovery of the novel viral RNA‒dependent RNA polymerases for treatment of HCV opened the doors for the development of a large number of anti−HCV drugs which require a concurrent development of sensitive analytical methods for their quality‒control and analysis. In this work, we report the first molecularly imprinted polymer‒based electrochemical sensor for the determination of the anti−HCV drug, daclatasvir dihydrochloride (DCT). Since liver patients require continuous medical follow up during the period of their treatment, it is very necessary to develop a simple and fast method for monitoring the DCT concentration in the patient’s biological fluids. In addition, quality control laboratories are in an urgent need for various rapid analytical methods for such a novel drug. The sensor is based on the modification of the traditional carbon‒paste sensor with molecularly imprinted methacrylic acid, cross linked with ethylene glycol dimethacrylate, as a selective adsorbent for DCT, and reduced graphene oxide (RGO) as a sensitizer. The Hartree‒Fock method of calculation was used to optimize the template/functional monomer ratios so as to avoid trial‒and‒error experimental synthesis procedures and to give information about the binding of the template to the functional monomer. The sensor was fully characterized with the aid of cyclic voltammetry, FTIR analysis and scanning electron microscopy (SEM). After optimization of the analytical parameters, the sensor was found to work efficiently at pH 2.5 (Britton Robinson buffer) and a scan rate of 500 mV/s. The sensor showed a good linearity over the concentration range of 40.6 pg/mL to 812 μg/mL with a correlation coefficient of 0.9881. The detection and quantification limits were found to be 12.2 and 40.2 pg/mL, respectively. The proposed sensor was applied for the determination of DCT in bulk solution, pharmaceutical formulation and human urine and plasma samples and excellent recovery values ranging from 99.48 to 103.70% were obtained indicating the accuracy of the proposed sensor.

Xanthene based spectroscopic probe for the analysis of HCV antiviral, daclatasvir dihydrochloride, through feasible complexation reaction

Two simple and rapid analytical assay methods were developed and validated utilizing spectroscopic techniques for sensitive and selective estimation of daclatasvir dihydrochloride in pharmaceutical preparations based on anion pair complex formation with eosin Y. The maximum absorbance of the formed complex was achieved at 545 nm in 0.1 M acetate buffer at pH 3.2 and Beer's law for absorbance against daclatasvir dihydrochloride concentration obeyed in the range of 1.0–10.0 μg mL−1 for the spectrophotometric method (method I). The technicality of spectrofluorimetric method (method II) based on measuring the quenching effect of daclatasvir dihydrochloride on the native fluorescence intensity of eosin Y at the same pH and the measurements were accomplished at 543 nm after excitation at 301 nm. The rectilinear calibration graph between daclatasvir dihydrochloride concentration and the fluorescence quenching values (ΔFI) was achieved over the range of 0.1–2.0 μg mL−1. The formation constant for the binary complex was 4.4 × 104 and Gibb's free energy change was −2.7 × 104 J mol−1. Both methods were validated in agreement with ICH guidelines and a successfully applied for the analysis dosage forms with good % recovery. A suggestion for the reaction pathway was supposed.

Stability-indicating micellar enhanced spectro-fluorometric determination of Daclatasvir in its tablet and spiked human plasma.

A fast, simple and sensitive micellar enhanced spectrofluorimetric method is performed for the determination of Daclatasvir dihydrochloride (DAC) in its pharmaceutical dosage form and in spiked human plasma. The fluorescence intensity (FI) was measured at 367 nm after excitation at 300 nm. In aqueous solution, the FI of DAC was greatly enhanced by >110% in the presence of sodium dodecyl sulphate (SDS). The detection method was linear over the range of 12.93 to 161.60 ng/mL, with a limit of detection of 1.75 ng/mL. The proposed method was successfully applied to the determination of DAC in its pharmaceutical dosage form and the mean % recovery of DAC in spiked human plasma was 95.42 ± 2.52. The developed methodology was also extended to stress studies of DAC after exposure to different forced degradation conditions including acidic, alkaline, photolytic, thermal and oxidative environments.

Micellar spectrofluorimetric protocol for the innovative determination of HCV antiviral (daclatasvir) with enhanced sensitivity: Application to human plasma and stability study

Daclatasvir dihydrochloride (DAC) is a new, direct-acting antiviral drug with powerful inhibitory effect against all hepatitis C virus (HCV) genotypes. A sensitive, simple, fast and specific fluorometric method for estimation of DAC in the presence of sofosbuvir was developed and validated. The method is based on reinforcement the fluorescence intensity of DAC by 170% of its original value in an aqueous solution of hexadecyl trimethyl ammonium bromide (pH 5.5, Teorell and Stenhagen buffer). The fluorescence intensity measurements were accomplished at 387 nm with 328 nm for excitation wavelength. A linear relationship was achieved between the DAC concentration and the fluorescence intensity in a range of 50.0–2000.0 ng ml−1 with 0.9998 and 0.9999 for the determination and correlation coefficients, respectively. The detection and quantitation limits were 13.4, 40.8 ng ml−1, respectively. The excellent sensitivity and specificity of the proposed method allowed the efficient estimation of DAC in real human plasma with adequate recovery (81.78 ± 1.57), and the selective determination for DAC in its commercial dosage form without interference from tablet excipient. Moreover, the proposed method was expanded to examine the stability of DAC by determination the parent drug of DAC in the presence of its oxidative, alkaline, acidic, UV, daylight and sunlight degradations products in agreement with ICH guidelines. Furthermore, the kinetic study of acidic and oxidative degradations of DAC was inspected. In addition, the half-life times of the reaction (t1/2) and the first-order reaction rate constants were estimated. Moreover, a suggestion for the degradation pathway was supposed.

SPECTROSCOPIC STUDY OF INTERACTION OF INDOMETHACIN AND DACLATASVIR DIGIDROCHLORIDE WITH HUMAN SERUM ALBUMIN

Fluorescence spectroscopy is one of the most effective methods for studying the binding of drugs to proteins, which is important for the purposes of biochemistry and medicine. When studying the interaction in vitro, human serum albumin (HSA) is used as a model, due to the presence of tryptophan residues in its composition, which are characterized by high sensitivity to the environment. The study of binding is based on the effect of static quenching of the intrinsic fluorescence of HSA by molecules of drugs that act as quenchers. However, such a method of studying the interaction becomes unsuitable if there is overlap (partial or complete) of the fluorescence spectra of the protein and the quenching molecule. Under physiological conditions, in vitro the interaction between indomethacin (IND), daclatasvir dihydrochloride (DAC) and HSA by the by fluorescence emission spectroscopy was studied. The results of the experiment show that the intrinsic fluorescence of the IND and DAC is manifested in the same spectral region as the intrinsic fluorescence of the protein. In the case of an IND, a small site of the spectrum shifted relative to the maximum can be selected, in which quenching of the fl uorescence of the protein is observed. In the case of DAC, its emission spectrum is completely superimposed on the intrinsic fluorescence of the protein. It is established that as a result of static interaction in the HSA -IND and HSA -DAC systems, the intrinsic fluorescence of IND and DAC by protein is quenched. The possibility of determining the binding constants of HSA with molecules of drug substances by quenching their intrinsic fluorescence is shown. The constants and number of binding sites in the HSA-IND and HSADAC systems are established. It is shown that the values of the binding constants, determined by quenching the HSA and by quenching the IND, have satisfactory similarity. A value for the average distance r between IND(DAC) and HSA was derived from the fluorescence resonance energy transfer. Since, the pharmaceutical firms need standardized screens for protein binding in the first step of new drug design, this kind of study of interaction between HSA and IND (DAC) would be useful in pharmaceutical industry and clinical medicine.

Stability Indicating 1st Derivative Synchronous Spectrofluorimetric Method for the Determination of the Newly Approved Antiviral Drug Daclatasvir in Presence of Its Oxidative and Photolytic Degradation Products: Application to Tablet Dosage Form

A highly sensitive, simple and rapid first derivative synchronous spectrofluorimetric method was utilized for the determination of daclatasvir dihydrochloride (DCV) in presence of its oxidative and photolytic degradation products. Where synchronous 1st derivative spectrofluorimetric approach was utilized to quantitatively determine DCV at 373 nm in presence of its oxidative degradation product and at 388 nm in presence of its photolytic degradation product that is obtained by exposing DCV to UV light at 312 nm, these were the zero-crossing wavelengths of degradation products without interference. The synchronous fluorescence was scanned at Δ λ of 80 nm. The method was found to be linear across the concentration range of 0.5-5.0 ng/mL with lower detection limit of 0.090 and lower quantification limit of 0.275 ng/mL (at 373 nm) and 0.268 ng/mL (at 388 nm). The adopted approach was successfully applied to commercial tablet and the results exhibited that the derivative synchronous fluorescence spectroscopy is a stabilityindicating method, suitable for routine use within a short analysis time. The proposed method was carefully validated for linearity, accuracy, precision, specificity and robustness.

Reversed-Phase Liquid Chromatographic Method for Determination of Daclatasvir Dihydrochloride and Study of Its Degradation Behavior

A simple and selective reversed-phase stability-indicating liquid chromatographic method has been developed and validated for the determination of daclatasvir in drug substance and drug product. Daclatasvir was subjected to acidic, alkaline, oxidative, thermal and photo-degradation study. The LC method was based on isocratic elution of daclatasvir and its degradation products on a reversed-phase C18 Hypersil column using a mobile phase consisting of phosphate buffer (10 mM, 1 mL triethylamine L−1): acetonitrile (60:40 v/v) at a flow rate of 2 mL min−1. Quantitation was achieved with UV detection at 312 nm. Linearity, accuracy, and precision were found to be acceptable over the concentration range of 0.75–120 μg mL−1, with regression coefficient value of 0.9999, and with limit of detection and quantitation of 0.148 and 0.447 μg mL−1, respectively. Peak purity was checked for principle drug and its alkali induced degradation product, and the pathway of alkaline hydrolysis of daclatasvir was suggested by LC/MS.