A new institutional clinical trial assessed the improvement of sleep disorders in 40 children with autism treated by immediate-release melatonin formulation in different regimens (0.5 mg, 2 mg, and 6 mg daily) for one month. The objectives of present study were to (i) prepare low-dose melatonin hard capsules for pediatric use controlled by two complementary methods and (ii) carry out a stability study in order to determine a use-by-date. Validation of preparation process was claimed as ascertained by mass uniformity of hard capsules. Multicomponent analysis by attenuated total reflectance Fourier transformed infrared (ATR-FTIR) of melatonin/microcrystalline cellulose mixture allowed to identify and quantify relative content of active pharmaceutical ingredients and excipients. Absolute melatonin content analysis by high performance liquid chromatography in 0.5 mg and 6 mg melatonin capsules was 93.6% ± 4.1% and 98.7% ± 6.9% of theoretical value, respectively. Forced degradation study showed a good separation of melatonin and its degradation products. The capability of the method was 15, confirming a risk of false negative < 0.01%. Stability test and dissolution test were compliant over 18 months of storage with European Pharmacopoeia. Preparation of melatonin hard capsules was completed manually and melatonin in hard capsules was stable for 18 months, in spite of low doses of active ingredient. ATR-FTIR offers a real alternative to HPLC for quality control of high-dose melatonin hard capsules before the release of clinical batches.

Phyllanthus species plants are a rich source of phenolics and widely used due to their medicinal properties. A liquid chromatography–tandem mass spectrometry (LC–MS/MS) method was developed using high-pressure liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry (HPLC-ESI-QTOF-MS/MS) for the identification and characterization of quercetin, kaempferol, ellagic acid and their derivatives in ethanolic extracts of Phyllanthus species. The chromatographic separation was carried out on Thermo Betasil C8 column (250 mm×4.5 mm, 5 μm) using 0.1% formic acid in water and 0.1% formic acid in methanol as the mobile phase. The identification of diagnostic fragment ions and optimization of collision energies were carried out using 21 reference standards. Totally 51 compounds were identified which include 21 compounds identified and characterized unambiguously by comparison with their authentic standards and the remaining 30 were tentatively identified and characterized in ethanolic extracts of P. emblica, P. fraternus, P. amarus and P. niruri.

Sofosbuvir is one of the new direct-acting antiviral drugs against hepatitis C virus (HCV) infection. This drug has recently been launched into the market, and generic versions of the medication are expected to be produced by local drug producers in some countries. Therefore, new methods are required to control sofosbuvir in pharmaceuticals. In the present study, a new method based on reversed phase (RP)-ultra-high performance liquid chromatography (UHPLC) coupled to diode array detection (DAD) and mass spectrometry (MS) was developed to facilitate the qualitative and quantitative analysis of sofosbuvir in film coated tablets. A wavelength of 260 nm was selected to perform a cost-effective quantification and the method showed adequate linearity, with an R2 value of 0.9998, and acceptable values of accuracy (75%–102%) and precision (residual standard deviation < 5%). The detection and quantification limits were 0.07 μg/mL and 0.36 μg/mL, respectively. Furthermore, the use of high-resolution MS enabled us to ensure the specificity, check impurities and better sensitivity. Therefore, this methodology promises to be suitable not only for the routine analysis of sofosbuvir in pharmaceutical dosage forms, but also for potential degradants.

Electromembrane extraction (EME) is an analytical microextraction technique, where charged analytes (such as drug substances) are extracted from an aqueous sample (such as a biological fluid), through a supported liquid membrane (SLM) comprising a water immiscible organic solvent, and into an aqueous acceptor solution. The driving force for the extraction is an electrical potential (dc) applied across the SLM. In this paper, EME is reviewed. First, the principle for EME is explained with focus on extraction of cationic and anionic analytes, and typical performance data are presented. Second, papers published in 2016 are reviewed and discussed with focus on (a) new SLMs, (b) new support materials for the SLM, (c) new sample additives improving extraction,(d) new technical configurations, (e) improved theoretical understanding, and (f) pharmaceutical new applications. Finally, important future research objectives and directions are defined for further development of EME, with the aim of establishing EME in the toolbox of future analytical laboratories.

Macrothelypteris torresiana is a fern species belonging to the family Thelypteridaceae. The present study was conducted to evaluate hepatoprotective potential of ethanol extract from M. torresiana aerial parts (EEMTAP) and detect the polyphenolic compounds present in the extract using high performance thin layerchromatography (HPTLC). Hepatoprotective potential of EEMTAP were tested at doses of 300 and 600 mg/kg, per os (p.o.), on Wistar albino rats. The extract and silymarin treated animal groups showed significant decrease in activities ofdifferent biochemical parameters like serum glutamic oxaloacetic transaminase(SGOT), serum glutamate-pyruvate transaminase (SGPT), and alkaline phosphatase (ALP), which were elevated by carbon tetrachloride (CCl4) intoxication. The levels of total bilirubin and total protein along with the liver weight were also restoredto normalcy by EEMTAP and silymarin treatment. After CCl4 administration, the levels of hepatic antioxidant enzymes such as glutathione (GSH) and catalase (CAT) were decreased whereas the level of hepatic lipid peroxidation (LPO) was elevated. The levels of these hepatic antioxidant enzymes were also brought to normalcy by EEMTAP and silymarin treatment. Histological studies supported the biochemical findings, and treatment with EEMTAP at doses of 300 and 600 mg/kg, p.o. was found to be effective in restoring CCl4-induced hepatotoxicity in rats. A simple HPTLC analysis was conducted for the detection of polyphenolic compounds in EEMTAP, and the result revealed the presence of caffeic acid as phenolic acid and quercetin as flavonoid. The proposed HPTLC method is simple and concise and provides a good resolution of caffeic acid and quercetin from other constituents present in EEMTAP.

In this study, the molecular interactions between valrubicin, an anticancer drug, and fish sperm DNA have been studied in phosphate buffer solution (pH 7.4) using UV–Vis spectrophotometry and cyclic voltammetry techniques. Valrubicin intercalated into double stranded DNA under a weak displacement reaction with methylene blue (MB) molecule in a competitive reaction. The binding constant (kb) of valrubicin-DNA was determined as 1.75×103 L/mol by spectrophotometric titration. The value of non-electrostatic binding constant (kt0) was almost constant at different ionic strengths while the ratio of kt0/kb increased from 4.51% to 23.77%. These results indicate that valrubicin binds to ds-DNA via electrostatic and intercalation modes. Thermodynamic parameters including ΔH0, ΔS0 and ΔG0 for valrubicin-DNA interaction were determined as ?25.21×103 kJ/mol, 1.55×102 kJ/mol K and ?22.03 kJ/mol, respectively. Cyclic voltammetry study shows a pair of redox peaks for valrubicin at 0.45 V and 0.36 V (vs. Ag/AgCl). The peak currents decreased and peak positions shifted to positive direction in the presence of DNA, showing intercalation mechanism due to the variation in formal potential.

In the pharmaceutical industry, the analysis of atropisomers is of considerable interest from both scientific and regulatory perspectives. The compound of interest contains two stereogenicaxes due to the hindered rotation around the single bonds connecting the aryl groups, which results in fourpotential configurational isomers (atropisomers). The separation of the four atropisomers was achieved on aderivatized β-cyclodextrin bonded stationary phase. Further investigation showed that low temperatureconditions, including sample preparation (?70 °C), sample storage (?70 °C), and chromatographic separation (6 °C),were critical to preventing interconversion. LC-UV-laser polarimetric analysis identified peaks 1 and 2as a pair of enantiomers and peaks 3 and 4 as another. Thermodynamic analysis of the retention data indicatedthat the separation of the pairs of enantiomers is primarily enthalpy controlled as indicated by the positiveslope of the van't Huff plot. The difference in absolute Δ (Δ H), ranged from 2.20 kJ/mol to 2.42 kJ/mol.

Our previous study demonstrated that human KIAA0100 gene is a novel acute monocytic leukemia-associated antigen (MLAA) gene. But the functional characterization of human KIAA0100 gene has remained unknown to date. Here, firstly, bioinformatic prediction of human KIAA0100 gene was carried out using online software;Secondly, human KIAA0100 gene expression was downregulated by the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) 9 system in U937 cells. Cell proliferation and apoptosis were next evaluated in KIAA0100-knockdown U937 cells. The bioinformatic prediction showed that human KIAA0100 gene was located on 17q11.2, and human KIAA0100 protein was located in the secretory pathway. Besides, human KIAA0100 protein contained a signal peptide, a transmembrane region, three types of secondary structures (alpha helix, extended strand, and random coil) , and four domains from mitochondrial protein 27 (FMP27). The observation on functional characterization of human KIAA0100 gene revealed that its downregulation inhibited cell proliferation, and promoted cell apoptosis in U937 cells. To summarize, these results suggest human KIAA0100 gene possibly comes within mitochondrial genome; moreover, it is a novel anti-apoptotic factor related to carcinogenesis or progression in acute monocytic leukemia, and may be a potential target for immunotherapy against acute monocytic leukemia.

Two simple and sensitive high performance liquid chromatography–tandem mass spectrometry (HPLC–MS/MS) methods were developed and validated for the determination of fenticonazole in human plasma after percutaneous and intravaginal administration. Mifepristone was used as an internal standard (IS), and simple protein precipitation by acetonitrile containing 2%acetic acid was utilized for extracting the analytes from the plasma samples. Chromatographic separation was performed on a Kinetex XB-C18 column. The quantitation was performed by a mass spectrometer equipped with an electrospray ionization source in multiple reactions monitoring (MRM) positive ion mode using precursor-to-product ion transitions of m/z 455.2–199.1 for fenticonazole and m/z 430.2–372.3 for mifepristone. The validated linear ranges of fenticonazole were 5–1000 pg/mL and 0.1–20 ng/mL in plasma for the methods A and B, respectively. For the two methods, the accuracy data ranged from 85% to 115%, the intra- and inter-batch precision data were less than 15%, the recovery data were more than 90%, and no matrix interference was observed. The methods A and B were successfully validated and applied to the pharmacokinetic studies of fenticonazole gel in Chinese healthy volunteers after percutaneous and intravaginal administration, respectively.

Larotaxel, a new taxane compound prepared by partial synthesis from 10-deacetyl baccatin Ⅲ, is active against tumors. In this research, a selective LC–MS method was developed and validated for the study of degradation kinetics of larotaxel, which was carried out in aqueous solutions with different pH (1.5, 3.0, 5.0, 6.5, 7.4, 9.0, 10 and 11.0) and temperature (0, 25, 37 and 45 °C). The linear range was 0.5–25μg/mL, the intra-and inter-day precisions were less than 7.0%, and accuracy ranged from 97.4–104.5% for each analyte. The observed rate obtained by measuring the remaining intact larotaxel was shown to follow first-order kinetics. The activation energies for degradation were 126.7 and 87.01 kJ/mol at pH 1.5 and 11, respectively. Although larotaxel was stable in pH 5, 6.5 and 7.4 buffers at 37 °C for 24 h during our study, increasing or decreasing the pH of the solutions would decrease its stabilities. Moreover, three main degradation products in alkaline condition were separated by HPLC and identified by Q–TOF–MS. The three degradation products were confirmed as 10-deacetyl larotaxel, 7, 8-cyclopropyl baccatin Ⅲ and 10-deacetyl-7, 8-cyclopropyl baccatin Ⅲ.