A UHPLC-Q-Exactive-Orbitrap-MS method for simultaneous determination of three flavonoids from Parasitic loranthus and their pharmacokinetics in rat plasma

Purpose: To develop and validate a chromatographic method for the simultaneous determination of plasma levels of rutin, avicularin and quercitrin using UHPLC-Q-Exactive-Orbitrap-MS. Methods: A sensitive, selective, and reliable UHPLC-Q-Exactive-Orbitrap-MS method was developed and validated for simultaneous determination of the three flavonoids, with puerarin as internal standard (IS). Plasma samples were first treated with methanol, and then acidified using hydrochloric acid (HCl) prior to liquid-liquid extraction with ethyl acetate. The flavonoids were separated on a Syncronis C18 column (100×2.1mm, 1.7 µm) using an elution gradient of acetonitrile and 0.1 % formic acid at a flow rate of 0.3 mL/min. Results: A linear correlation was obtained for the three flavonoids over the investigated concentration range, with correlation coefficients > 0.9954. The values of validated lower limit of quantification (LLOQ) were 0.68, 1.42 and 2.54 ng/mL for rutin, avicularin and quercitrin, respectively. Intra- and inter-day precision (RSD) were < 10 %, while accuracy (RE) ranged from −3.76 to 4.04 %. Conclusion: The proposed method has been successfully validated and is suitable for studying the pharmacokinetics of the three analytes in rats treated with parasitic loranthus extract (PLE).

Simultaneous Determination of 25 Ginsenosides by UPLC-HRMS via Quantitative Analysis of Multicomponents by Single Marker

A method using UPLC-HRMS has been developed for a rapid, simultaneous qualitative and quantitative analysis of twenty-five ginsenosides. Chromatographic separation was achieved on a C18 analytical column with an elution gradient comprising 0.1% aqueous formate/acetonitrile as the mobile phase. HRMS detection acquired full mass data for quantification and fullms-ddms2 (i.e., data-dependent scan mode) yielded product ion spectra for identification. Furthermore, quantitative analysis of multiginsenosides by single marker (QAMS) was developed and validated using a relative correction factor. Under optimal conditions, we could simultaneously separate eight groups of isomers of the 25 ginsenosides. Good linearity was observed over the validated concentration range for each analyte (r2 > 0.9924), showing excellent sensitivity (LODs, 0.003–0.349 ng/mL) and lower limit quantification (LOQs, 0.015–1.163 ng/mL). The LC-MS external standard method (ESM) and QAMS were compared and successfully applied to analyze the ginsenoside content from Panax ginseng roots. Overall, our UPLC-HRMS/QAMS approach provides high precision, stability, and reproducibility and can be used for high-throughput analysis of complex ginsenosides and quantitative analysis of multiple components and quality control of traditional Chinese medicines (TCM).

Efficient analysis of monosaccharides and oligosaccharides from hydrolyzed hemicellulose of Spartina anglica

Spartina anglica, a plant that controls coastal erosion, is widely distributed throughout the world and is rich in cellulose, hemicellulose, and lignin. The hemicellulose from Spartina anglica can be extracted and hydrolyzed into monosaccharides and xylooligosaccharides under acid or enzymatic digestion conditions. In this study, an effective PMP(1-phenyl-3-methyl-5-pyrazolone)-derivatized HPLC (High performance liquid chromatography) method was developed for monitoring monosaccharides and xylooligosaccharides of Spartina anglica. With phosphate buffer (0.04 M, pH 8.06) as mobile phase A, and acetonitrile as mobile phase B, in which the elution gradient was set as A:B/79:21, the monosaccharides (glucose, xylose and arabinose) and xylooligosaccharides (xylobiose, xylotriose, xylotetraose, xylopentaose, xylohexaose) could be separated completely using the C18 column. This provides an economical, rapid, and efficient method for process monitoring in the bioconversion of Spartina anglica.

MICROPROCESSOR VALVE CONTROL UNIT FOR SEMI-PREPARATIVE CHROMATOGRAPH

Software has been developed for the microprocessor unit of the valve control board of a semi-preparative chromatograph. A model for constructing an elution gradient using the solenoid valves CTV-31-32U-4 and MTV-3-1 / 4UFH-3 with two channels and three ports for connection (one common input and two output) was developed and implemented.

Determination of dyes and marker in diesel using high performance liquid chromatography

A fast, precise and accurate high performance liquid chromatography method has been developed for the determination of dyes (Solvent Red 19 and Solvent Blue 35) and a marker (Solvent Yellow 124) in diesel. Separation was carried out on a 250 × 4.60 mm Agilent Zorbax Rx-SIL column (5 µm particle size). Detection was done in a visible wavelength range. The best performance of fuel dye separation and the shortest retention times were achieved when using hexane, toluene and ethyl acetate as a mobile phase. During this research the eluent composition and the elution gradient were optimized consequently that helped to perform the analysis within 15 min. The developed method was applied for the analysis of real samples of dyed diesel fuel. Preparation of the samples for the analysis simply consisted of filtering through a 0.45 µm filter previous to direct injection of the sample into the HPLC system for analysis.

Determination of Polyphenols in White Wines by Liquid Chromatography: Application to the Characterization of Alella (Catalonia, Spain) Wines Using Chemometric Methods

The determination of polyphenols in wines is of great interest in the field of food analysis due to health and organoleptic implications. In addition, the applicability of polyphenols as food descriptors to be used for characterization, classification, and authentication purposes is gaining popularity. In this work, a simple and reliable method based on HPLC separation in reversed-phase mode with UV-Vis detection was developed and applied to determine polyphenolic compounds in white wines. The chromatographic separation was performed using aC18 column under a methanol elution gradient and assessed by an experimental design approach. Analytical parameters were established under the optimal experimental conditions. LOD values were between 3 and 220 µg/L, and repeatability values were better than 1% for most of the analyzed polyphenols. Compositional data were further exploited to characterize white wines based on principal component analysis to discriminate among mono- and polyvarietal compositions.

New RP-HPLC Method for Separation of Naja haje haje Venom and Studies of its Bactericidal Effect

Background: Snake venom is a complex mixture of biologically active substances. Some peptides and proteins from snake have already demonstrated their therapeutically potential. The venom of Naja haje, an Elapidae member, has been analyzed from this point of view. Understanding the fully biochemical role of its enzymes has determined the scientists to find new separation and identification methods. Objective: Our goal was to develop an optimal HPLC analytical method for separation and identification of Naja haje snake venom components, known for its neurotoxic activity. In addition, we wanted to find out if crude snake venom could inhibit the development of both Gram-positive and Gram-negative bacterial cultures. Materials and Method: Analysis of venom was performed on a HPLC system using a C16 column with UV detection at 210 nm. The analysis was done using two mobile phases, containing different concentrations of acetonitrile and trifluoroacetic acid aqueous solution at different pH values. An elution gradient was set at a flow of 0.60 mL/min. Bactericidal activity was quantified by measuring inhibition diameter around an aseptically disk filled with crude venom using Staphylococcus aureus and Escherichia coli. Results: An optimal HPLC analytical method has been developed by changing different parameters such as the pH value of mobile phase A or the elution gradient. The best resolution were obtained at a pH value of 7.4, in gradient varying from 5% to 45% in mobile phase B. Microbiological studies of the venom showed that Gram-positive bacteria growth was inhibited by crude venom, while on Gram-negative bacteria growth no effect was observed. Inhibition zone is dose-dependent and fresh crude venom is with 30% more potent than venom freeze and kept at -55°C. Conclusions: A comprehensive catalog of venom composition may serve as a starting point for studying structurefunction correlations of individual toxins for the development of new research tools and drugs of potential clinical use.

Degradation of amino acids by leaching in feeds for shrimp

The feed for shrimp are one of the most expensive in the aquaculture sector, mainly because this type of feed should have high stability in water. This study aimed to evaluate the stability of amino acids in commercial feeds with different protein contents intended for larval and juvenile shrimp subjected to leaching. The feed samples were exposed to the leaching process during the time period of 04, 08 and 12 hours. The analyses of degradation of amino acids were performed using an elution gradient in HPLC system. In all diets evaluated it was found that lysine and histidine are essential amino acids which suffered less degradation processes. It’s important to mention that arginine is considered an important amino acid for growth of shrimp. In both feeds with 35% protein (RA35 and RB35) the losses of arginine were 79 and 89% respectively. The results obtained in this study indicate that the leaching process significantly reduced the content of amino acids in the feeds. The physical structure of the feed doesn't prevent the degradation process of amino acids in the leaching process.