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An ultrasound-assisted dispersive liquid-liquid microextraction by solidifying floating organic droplets, coupled to a form of temperature-programmed gas chromatography flame ionization detection, has been developed for the extraction and determination of thymol and carvacrol. This method utilizes undecanol as the extraction solvent, offering advantages such as facilitating phase transfer through solidification and enhancing solvent-focusing efficiency. The optimal gas chromatography conditions include a sample injection volume of 0.2 µL, a split ratio of 1:10, and a flow rate of 0.7 mL min -1 . The extraction conditions entail an extraction solvent volume of 20 µL, a disperser solvent (acetone) volume of 500 µL, pH 7.0, 7.0% NaCl (3.5 M), a sample volume of 5.0 mL, an ultrasound duration of 10 min, and a centrifuge time of 7.5 min (800 rpm). These conditions enable the achievement of a high and reasonable linear range of 3.5 to 70. 0 μg mL -1 for both thymol and carvacrol. The detection limits are found to be 0.95 and 0.89 μg mL -1 , respectively, for thymol and carvacrol. The obtained relative standard deviations, 2.7% for thymol and 2.6% for carvacrol, demonstrate acceptable precision for the purpose of quantitative analysis.

Modification of micellar liquid chromatography (MLC) with a natural deep eutectic solvent (NADES) and butanol was effectively performed for analysis of four crucial cardiovascular drugs namely aspirin, atorvastatin, metformin, and metoprolol. The multivariate software tools were employed for screening and optimizing the primary experimental parameters including sodium dodecyl sulphate concentration ([SDS]) as well as the volume percentages of NADES, butanol, and glacial acetic acid (GAC). By using the desirability function, the optimal framework of the mobile phase was obtained at volume ratio of 83:10:3.5:3.5, respectively, for SDS (0.09 mol L -1 ), butanol, NADES, and GAC. Moreover, the curvature and effect plots were applied for evaluating the robustness of the procedure. At the optimal conditions, the drugs were excellently separated within a total analysis time of 12 min when the flow rate of the mobile phase was 1 mL min -1 . To certify the performance of the developed procedure, the Food and Drug Administration guidelines for bioanalytical analysis have been implemented. The designed MLC system was successfully utilized for the quantification of the drugs in urine and plasma samples., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interest., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Modified micellar liquid chromatography (MLC) with a natural deep eutectic solvent (NADES), produced from choline chloride (ChCl) and ethylene glycol (EG), was employed for melamine (MEL) monitoring in milk matrix. This sustainable mobile phase was attained through chemometrical optimization of crucial variables including concentration of sodium dodecyl sulphate ([SDS]) along with volume percentages of both NADES and glacial acetic acid (GAC). The desirability function and central composite design were utilized as chemometrical tools. Retention time (t R - MEL ), and chromatographic peak width of MEL at 50% of its height (W 50% - MEL ) were considered for finding the best possible arrangement of the influential factors in the configuration of the mobile phase. Under the optimal experimental conditions of 0.10 mol L -1 SDS, 4% (v/v) NADES, and 4% (v/v) GAC, the results showed that both t R - MEL and W 50% - MEL drastically decreased when NADES was a part of the mobile phase composition. This indicated that ChCl-EG-based NADES had a significant impact on improving the chromatographic behaviour of an ionizable polar compound, MEL. At the optimal point, MEL was eluted in approximately 10 min without being interfered by coexisting proteins and endogenous species in milk. The practical performance of the mobile phase was established through direct injection of milk samples into the MLC system. The eligibility criteria of the United State-Food and Drug Administration (US-FDA) were considered for validation of the introduced methodology., (Copyright © 2019. Published by Elsevier B.V.)

An extracellular protease was purified from a novel moderately halophilic bacterium Salinivibrio sp. strain MS-7 by the combination of an acetone precipitation (40-80 %) step and a DEAE-cellulose anion exchange column chromatography. Kinetic parameters of the enzyme exhibited V(max) and K(m) of 130 U/mg and 1.14 mg/ml, respectively, using casein as a substrate. The biochemical properties of the enzyme revealed that the 21-kDa protease had a temperature and pH optimum of 50 °C and 8.0, respectively. The enzyme was strongly inhibited by phenylmethylsulfonyl fluoride, Pefabloc SC, chymostatin, and also EDTA, indicating that it belongs to the class of serine metalloproteases. Interestingly, Ba(2+) and Ca(2+) (2 mM) strongly enhanced the enzyme activity, while Fe(2+) and Mg(2+) activated moderately and Zn(2+), Ni(2+), and Hg(2+) decreased the enzyme activity. The effect of organic solvents with different logP on the purified protease revealed complete stability in toluene, ethyl acetate, chloroform, and n-hexane at 10 and 50 % (v/v) and moderate stability even in 50 % of DMSO and ethanol. The behavior of the MS-7 protease in three imidazolium-based ionic liquids exhibited suitable activity in these green solvent systems, especially in 1-hexyl-3-methylimidazolium hexafluorophosphate ([C(6)MIM][PF(6)]). Comparison of the purified protease with other previously reported proteases suggests that strain MS-7 secrets a novel organic solvent-tolerant protease with outstanding activity in organic solvents and imidazolium-based ionic liquids, which could be applied in low water synthetic section of industrial biotechnology.