Sensitive MALDI-TOF MS and 'turn-on' fluorescent genosensor for the determination of DNA damage induced by CNS acting drugs.

The genotoxic and carcinogenic adverse effects of various drugs should be considered for assessing drug benefit/risk ratio. On that account, the scope of this study is to examine the kinetics of DNA damage triggered by three CNS acting drugs; carbamazepine, quetiapine and desvenlafaxine. Two precise, simple and green approaches were proposed for probing drug induced DNA impairment; MALDI-TOF MS and terbium (Tb3+) fluorescent genosensor. The results revealed that all the studied drugs induced DNA damage manifested by the MALDI-TOF MS analysis as a significant disappearance of the DNA molecular ion peak with the appearance of other peaks at smaller m/z indicating the formation of DNA strand breaks. Moreover, significant enhancement of Tb3+ fluorescence occurred, proportional to the amount of DNA damage, upon incubation of each drug with dsDNA. Furthermore, the DNA damage mechanism is examined. The proposed Tb3+ fluorescent genosensor showed superior selectivity and sensitivity and is significantly simpler and less expensive than other methods reported for the detection of DNA damage. Moreover, the DNA damaging potency of these drugs was studied using calf thymus DNA in order to clarify the potential safety hazards associated with the studied drugs on natural DNA.

Second-derivative synchronous fluorimetry and time-programmed HPLC-fluorescence detection for simultaneous estimation of flibanserin and sitagliptin phosphate in synthetic mixtures and human plasma samples.

Diabetes Mellitus is directly related to female anaphrodesia. Female Viagra or Flibanserin (FLB), U.S. FDA approved in 2015, is specifically indicated for premenopausal Hypoactive Sexual Desire Disorder, HSDD, which is one of the primary consequences of Diabetes Mellitus. Simultaneous analysis of the concomitantly administered, FLB and oral antidiabetics, as Sitagliptin phosphate (STG), is a crucial demand to investigate mutual drug-drug interaction. The latter is responsible for uncontrolled glycaemia and higher risk of sudden hypoglycemia. Two simple, sensitive, economical and direct analytical methods, namely, Second-Derivative Synchronous Fluorimetric Spectroscopy, D2-SFS, and High Performance Liquid Chromatography with fluorimetric detection, HPLC-FD, are established for simultaneous determination of FLB and STG in their binary mixtures. First method relies on measuring D2-SFS spectra of both drugs, at Δλ = 25 nm, along linearity ranges of 0.05-1 µg/mL for both drugs. The second method is a chromatographic one with gradient elution of FLB and STG on RP-ZORBAX Eclipse C18 column (5 µm, 4.6 × 150 mm). Mobile phase; phosphate buffer: acetonitrile, pH 4.5, with a flow rate of 1 mL/min at room temperature has been used. Time programmed fluorimetric detection is optimized at λem = 305 nm for STG (0.0-5.9 min), at λem = 375 nm for FLB (6-9 min) after both excitation at λex = 257 nm, in the linear ranges of 1-40 µg/mL and 5-60 µg/mL for FLB and STG, respectively. Proposed methods have been validated according to ICH guidelines, then applied for simultaneous quantitation of FLB and STG in their laboratory-prepared mixtures and in spiked human plasma samples. Satisfactory Student's t-value and F-variance ratio have been obtained upon comparing the results of both methods.

A Validated Stability-Indicating HPTLC Assay for Determination of 10-Hydroxy-2-Decenoic Acid Content in Royal Jelly Products Using Robust Regression Methods.

A new, simple, stability-indicating high-performance thin-layer chromatography method was developed for the quantification of 10-hydroxy-2-decenoic acid (10-HDA) in some royal jelly products marketed in Egypt. The used solvent system was chloroform:acetic acid (10:1, v/v) and the bands were measured densitometrically at 210 nm. First- and second-derivative treatments of the data were performed. The present study shows a comparison between three statistical regression methods for handling data: parametric, nonparametric and weighted regression (WR) methods. The developed methods were validated as per International Conference on Harmonization guidelines. To validate the stability-indicating power of the developed analytical method, the royal jelly standard was subjected to forced degradation studies including the effect of hydrolysis, oxidation, photolysis and dry heat. It was found that derivative treatment of the chromatographic response data gives improved quantitation and sensitivity of the chromatographic signals. Weighted regression of the response data is found to be advantageous over the use of both parametric and nonparametric regression models. This was shown by a great enhancement in the accuracy and precision in the analysis of 10-HDA in royal jelly products. The % recovery in case of WR was 99.92 ± 0.16, while % recovery in case of nonparametric and parametric regressions were 99.56 ± 0.25 and 98.63 ± 0.65, respectively.

Analysis of astragalosides I, II and IV in some Egyptian Astragalus species and Astragalus dietary supplements using high-performance liquid chromatography/evaporative light scattering detector and non-parametric regression.

INTRODUCTION: GenuTs Astragalus L. is characterised by the presence of cycloartane saponins which have wide biological activities such as antioxidant, immunomodulating' hepatoprotective and anti-inflammatory activities. From these cycloartane saponins are astragalosides I, II and IV which have been regarded as the most important active constituents in Astragalus species. OBJECTIVES: This work describes the quantitative analysis of astragalosides I, II and IV in some Egyptian Astragalus species and Astragalus dietary supplements in a single run by high-performance liquid chromatography/evaporative light scattering detector (HPLC/ELSD) using gradient elution. METHODOLOGY: The method of quantitation adopted in this study is the standard addition method. First and second derivative treatment of the data was performed, and the study presents comparison between two statistical regression methods for handling data; parametric and non-parametric regression methods. RESULTS: Derivative treatment of the chromatographic response data gives improved quantitation of the chromatographic signals. Non-parametric regression of the data using Theil's method is advantageous over the usual least squares method as it assumes that errors could occur in both x- and y-directions and they might not be normally distributed. In addition, it could effectively circumvent any outlier data points. CONCLUSION: Due to the simplicity and the good accuracy and reproducibility of the suggested methods, they could be used for analysis and quality control of Astragalus species and Astragalus dietary supplements.

Enantioselective HPLC-DAD method for the determination of etodolac enantiomers in tablets, human plasma and application to comparative pharmacokinetic study of both enantiomers after a single oral dose to twelve healthy volunteers.

An enantioselective high performance liquid chromatographic method with diode array detection (HPLC-DAD) was developed and validated for the determination of etodolac enantiomers in tablets and human plasma. Enantiomeric separation was achieved on a Kromasil Cellucoat chiral column (250 mm × 4.6mm i.d., 5 µm particle size) using a mobile phase consisting of hexane: isopropanol: triflouroacetic acid (90:10:0.1 v/v/v) at a flow rate of 1.0 mL min(-1). The chromatographic system enables the separation of the two enantiomers and the internal standard within a cycle time of 8 min. The resolution between the two enantiomers was 4.25 and the resolution between each enantiomer and the internal standard was more than 2.0. Detection was carried out at 274 nm, and the purity assessment was performed using a photodiode array detector. Solid phase extraction technique using C-18 cartridge was applied to extract the analytes from the plasma samples, and the percentage recovery was more than 95% for the lower quantification limit. The method has been validated with respect to selectivity, linearity, accuracy and precision, robustness, limit of detection and limit of quantification. The validation acceptance criteria were met in all cases. The linearity range for the determination of each enantiomer in human plasma was 0.4-30.0 µg mL(-1) and the limits of quantification of R-etodolac and S-etodolac were 0.20 and 0.19 µg mL(-1), respectively. The validated method was successfully applied to the determination of etodolac enantiomers in tablets and to a comparative pharmacokinetic study of the two enantiomers after the administration of 300 mg single oral dose etodolac racemate tablets to twelve healthy volunteers.

Determination of methotrexate and indomethacin in urine using SPE-LC-DAD after derivatization.

A validated HPLC-DAD assay is presented for the simultaneous quantification of methotrexate and indomethacin in a drug combination which is used synergistically to intervene with tumoral or inflammatory tissue microenvironments. The analytes were isolated from urine via solid phase extraction. The method is based on derivatizing both analytes with a soluble carbodiimide coupler and 2-nitrophenylhydrazine directed to their commonly occurring carboxylate functions. The chromatographic separation was accomplished on an octylsilica column in less than 15 min using acetate buffer (pH 4; 10 mM)-methanol (60:40, v/v) as eluent at 1.5 ml/min and monitored at 400 nm. The selectivity of the method was demonstrated in a pre-dosed rheumatoid arthritis patient. Quality control samples were prepared and analyzed to reveal the validity of the method. Life samples collected from a healthy volunteer were monitored for both drugs and their urinary levels were determined.

Simultaneous determination of the two non-steroidal anti-inflammatory drugs ; diflunisal and naproxen in their tablets by chemometric spectrophotometry and HPLC.

Chemometric spectrophotometry and HPLC were applied to the simultaneous determination of the two non-steroidal anti-inflammatory drugs; diflunisal (I) and naproxen (II). The applied chemometric techniques are multivariate methods including classical least squares (CLS), principal component regression (PCR) and partial least squares (PLS); and the second derivative of the ratio spectra ((2)D(r)) method. To develop the multivariate methods, the UV absorption spectra of the standard solutions of the training and validation sets in methanol were recorded in the range of 242-274 nm at 2 nm intervals. The specificity of the studied multivariate methods has been tested. In the (2)D(r) method, analytical signals at 235 and 259 nm were selected for the determination of (I) and (II), respectively. The HPLC method depends on reversed-phase separation using C18 column. The mobile phase consists of a mixture of acetonitrile - acetate buffer (pH 4.2; 50 mM) (60:40, v/v). The UV detector was set at 255 nm. The developed methods were validated and successfully applied to the simultaneous determination of (I) and (II) in their tablets. The assay results obtained using the chemometric methods were statistically compared to those of the HPLC method and good agreement was observed.

Chemometric determination of rabeprazole sodium in presence of its acid induced degradation products using spectrophotometry, polarography and anodic voltammetry at a glassy carbon electrode.

Chemometric stability indicating methods are presented for the determination of rabeprazole sodium in presence of its acid induced degradation products using spectrophotometry, differential pulse polarography and differential pulse anodic voltammetry at a glassy carbon electrode. The applied chemometric techniques are multivariate ones including classical least squares (CLS), principal component regression (PCR) and partial least squares (PLS). A difference spectrophotometric (DeltaA) method has also been applied. To develop the multivariate calibrations, a training set was used, consisting of 20 mixture solutions of rabeprazole sodium and its degradation products. These mixtures show percentage degradation ranging from 0.5-65%, 0.5-95% and 0.6-75% for the spectrophotometric, polarographic and anodic voltammetric calibrations, respectively. The UV absorbances were recorded in 0.1 M NaOH within the wavelength range 220-340 nm at 2 nm intervals. The polarograms and anodic voltammograms were recorded in Britton-Robinson buffer (pH 8.0) within the potential range -500 to -1508 and 400 to 1192 mV at 6 mV intervals with a pulse amplitude of -100 and 50 mV, sweep rate of 15 and 10 mV s(-1) and pulse interval of 0.4 and 0.6 s for the polarographic and anodic voltammetric methods, respectively. All the studied methods have been validated and successfully applied to the determination of rabeprazole sodium in tablet dosage form. The results were statistically compared to those obtained using a published HPLC method. No significant difference has been found.

Polarographic chemometric determination of zinc and nickel in aqueous samples.

Polarographic chemometric methods were applied to the determination of zinc and nickel in aqueous solutions previously acidified with 0.1M acetate buffer (pH 4.2). The studied methods are multivariate methods including classical least squares (CLS), principal component regression (PCR) and partial least squares (PLS); derivative ratio methods (first, (1)D and second, (2)D derivative ratio). A comparative study was considered. The studied chemometric methods do not need the presence of any reduction potential shift reagent in spite of the great overlap between the two metals polarograms. A training set consisting of 10 binary mixture solutions in the possible combinations containing 0.13-9.30mug/ml Zn(II) and 0.20-12.25mug/ml Ni(II) was used to develop the chemometric calibrations (CLS, PCR and PLS). A validation set containing the synthetic mixtures in the range of 0.29-9.00mug/ml for Zn(II) and 0.30-11.60mug/ml for Ni(II) was used to validate the multivariate calibrations. Same mixtures were used to develop the derivative ratio methods. The polarograms were recorded and their current values were measured within the potential range -920 to -1052mV at 2mV intervals. The mean percentage recoveries obtained using CLS, PCR and PLS were found to be 99.5+/-1.5%, 100.0+/-1.1% and 100.0+/-1.0% for Zn(II) and 99.4+/-1.3%, 99.7+/-1.2% and 99.9+/-1.0% for Ni(II), respectively. The mean percentage recoveries obtained using (1)D at -950mV, (1)D at -1010mV, (1)D at -950mV-(1)D at -1010mV and (2)D at -986mV for Zn(II) were found to be 99.7+/-1.2%, 99.2+/-1.6%, 99.4+/-1.4% and 99.4+/-1.4%; and using (1)D at -1030mV and (2)D at -1010mV for Ni(II) were found to be 100.5+/-1.3% and 100.4+/-1.3%, respectively. Interferences due to the presence of Cd, Co, Pb, Fe, Mn, Ca, Mg, Cu and Al were studied. The applicability of the proposed methods was assessed through the determination of both metals in tap drinking-water. Samples were subjected if required up to a 20-fold preconcentration step by microwaving in pyrex vessels. The results were compared with those obtained using the zincon and the heptoxime colorimetric reference methods for the determination of zinc and nickel, respectively.

Spectrophotometric determination of omeprazole, lansoprazole and pantoprazole in pharmaceutical formulations.

The compensation method and other chemometric methods (derivative, orthogonal function and difference spectrophotometry) have been applied to the direct determination of omeprazole, lansoprazole and pantoprazole in their pharmaceutical preparations. The methods have been validated; the limits of detection were found to be 3.3x10(-2), 3.0x10(-2) and 3.5x10(-2) microgram ml(-1) for the three drugs, respectively. The repeatabililty of the methods were found to be 0.3-0.5%. The linearity ranges were found to be 0.5-3.5 microgram ml(-1). The proposed methods have been applied to the determination of the three drugs in their grastro-resistant formulations. The difference spectrophotometric (DeltaA) method is unaffected by the presence of acid induced degradation products; hence can be used as a stability indicating assay.