Authors:S. Thomas, S. K. Paul, A. Agarwal, and C. S. Mathela
Galantamine hydrobromide was subjected to oxidative stress degradation using hydrogen peroxide and analyzed as per the chromatographic conditions described in European Pharmacopoeia. The drug showed considerable degradation at ambient temperature resulting in the formation of two degradation products at relative retention times (RRTs) 0.63 and 2.52. The minor degradant at RRT 0.63 was identified as galantamine N-oxide. The principal degradant formed at RRT 2.52 was found to be unknown and has not been reported previously. The unknown impurity was identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS) followed by isolation using semi-preparative high-performance liquid chromatography (HPLC). The isolated impurity was characterized using one-dimensional, two-dimensional nuclear magnetic resonance spectroscopy (1D and 2D NMR) and elemental analysis (EA). The principal degradant was found to be formed due to the generation of bromine and subsequent attack on the aromatic ring via in situ reaction between hydrogen bromide and hydrogen peroxide. The unknown impurity was characterized as (4aS,6R,8aS)-5,6,9,10,11,12-hexahydro-1-bromo-3-methoxy-11-methyl-4aH-benzofuro [3a,3,2-ef]  benzazepin-6-ol.
Authors:J. Kuc, A. Grochowalski, S. Mach, and D. Placha
This paper is focused on the determination of three hexabromocyclododecane isomers, α, β, and γ, in the tissue of the most commonly consumed marine and farmed fish in Central Europe. The analytical procedure contains multiple steps: extraction, dialysis using semipermeable membranes, and a clean-up step using a silica gel column. Hexabromocyclododecane (HBCD) isomers were determined by sensitive isotope dilution liquid chromatography-tandem mass spectrometry (LC-MS/MS) method which is characterized by a low limit of detection (1 pg g−1 fresh weight). The recovery values obtained for individual isomers were in the range of 89–124%. The γ-HBCD isomer was detected in all samples. This isomer also had the largest percentage in comparison with the other isomers, except in salmon, in which the α-HBCD isomer was more abundant. The highest level of total HBCD was measured in mackerel (650 ± 195 pg g−1 fresh weight). Cluster analysis allowed the differentiation of groups of objects and the display of objects based on the degree of integration within the same group. Spearman’s rank correlation indicated no statistically significant difference between the levels of HBCD and the fat content in the fish samples.
High-performance liquid chromatography coupled with fluorescence (HPLC-FD) and tandem mass spectrometric detection (LC-MS/MS) was studied as a versatile tool for fast and reliable determination of nine regulated quinolones in food of animal origin (Council Regulation 2377/90/ECC). The sample pre-treatment protocol includes double step extraction with acetonitrile followed by solid phase extraction (SPE) cleanup on hydrophobic-lipophilic balance (HLB) cartridge. The separation of quinolones in HPLC-FD determination was performed on C18 Zorbax column with a gradient mixture of aqueous formic acid, methanol, and acetonitrile. A multi-wavelength excitation/emission program was used for sensitive quinolones detection. The separation efficiency of newly available chromatographic columns: Gemini C18 and Synergi Polar RP (fully porous particles), as well as Kinetex PFP and Poroshell 120 EC-C18 (core-shell particles), was studied in liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) analysis. Appropriate gradient elution program was designed for each column. Multiple reaction monitoring was used for selective determination of each quinolone. LC-MS/MS allowed quinolones determination in less than 5 min. Both methods showed detection limits below maximum residue limits for quinolones residues in food commodities.
Authors:L. Vlase, D.-S. Popa, A. Tero-Vescan, and N. Olah
A new liquid chromatography tandem mass spectrometry (LC-MS/MS) assay for the quantification of seven isoflavones (daidzin, genistin, ononin, daidzein, glycitein, genistein, and formononetin) and coumestrol in vegetable extracts was developed. The separation was performed on a Zorbax SB-C18 column with a mixture of methanol (solvent A) and 0.1% (υ/υ) acetic acid in water (solvent B) under gradient conditions at 50°C with a flow rate of 1 mL min−1. The detection of analytes was performed by electrospray ionization, negative ionisation, in non-reactive MS2 mode for aglycons or in reactive MS2 mode for glycosides. The method shows a good linearity (r2 > 0.9948) over the concentration range of 40–4000 ng mL−1 for all analytes, a good precision (CV < 11%) and accuracy (<10%). The method was successfully applied to quantify the isoflavones and coumestrol in vegetable extracts obtained from red clover (Trifolium pratense L., Fabaceae) and dyer's greenweed (Genista tinctoria L., Fabaceae) and can be used in the chemical characterization of vegetables with phytoestrogen content.
Authors:Y. Wang, M. Wang, J. Li, S. Yao, J. Xue, W. Zou, and C. Hu
An improved ion-pairing reversed-phase high-performance liquid chromatography method coupled with evaporative light scattering detection (HPLC-ELSD) was developed to determine spectinomycin and its related substances in commercial samples. The method was validated in accordance with International Conference on Harmonization (ICH) guidelines. The specificity of the HPLC-ELSD method was similar to that of the European Pharmacopoeia (Ph. Eur.) method, and repeatability and robustness were markedly improved relative to other reported methods due to our empirical evaluation of separation columns. Indeed, it is a more specific assay of spectinomycin than traditional microbiological techniques. The HPLC-ELSD method was used to evaluate the impurity profiles of eight compounds in seven spectinomycin batches from five different companies. Liquid chromatography coupled with electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) was employed to characterize the structures of these compounds. Though the HPLC-ELSD method was not as sensitive as the Ph. Eur. method, its limit of quantitation (LOQ) (0.16%) was lower than the disregard limit (0.3%) described by the Ph. Eur. 7.0. This suggests that the HPLC-ELSD method is appropriate for routine analysis of spectinomycin and its related substances.
Authors:Yan-Mei Zhong, Xun-Long Zhong, Ji-Hua Wang, and Liang Han
Patrinia scabiosaefolia Fisch. (PSF), a well-known traditional Chinese medicine, has been demonstrated to show therapeutic effects on inflammatory bowel disease. In this study, a rapid and sensitive method using ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC/Q-TOF-MS) was developed for identification of the major constituents in PSF. The separation analysis was performed on Waters Acquity UPLC system, and the accurate mass of molecules and their fragment ions were determined by Q-TOF-MS. Thirty-one constituents, including triterpenoids, iridoids, flavonoids, and organic acids were detected and tentatively deduced on the basis of their element compositions, tandem mass spectrometry (MS/MS) data, and relevant literatures. Twelve constituents were discovered for the first time in PSF. The results demonstrated that hederagenin-type and oleanolic acid-type saponins were the main constituents of PSF. Our work provides a certain foundation for further quantitation of major chemical constituents and in vivo pharmacokinetic studies of PSF. Moreover, the analytical approach developed herein has proven to be generally applicable for profiling the chemical constituents in traditional Chinese medicines (TCMs) and other complicated mixtures.
The composition and concentration of natural products largely depend on a plant part, development stage, cultivar, and growing conditions. This study evaluated the influence of cultivars and production systems on the composition of natural products (benzoxazinoids) in wheat aerial parts. The determination of benzoxazinoids was performed by combining pressurized liquid extraction, ultra-performance liquid chromatography, and tandem mass spectrometry. Six benzoxazinoids were identified and quantitated in wheat varieties. Significant differences were observed among the examined varieties. The average concentrations of total researched compounds were definitely higher in the organically produced spring wheat cultivars than in the winter ones. The content of these compounds in the same varieties grown under organic and conventional systems showed their higher content under the organic one. The main benzoxazinoids detected in wheat varieties were 2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one glucoside (DIMBOA-Glc) and 6-methoxy-2-benzoxazolinone (MBOA). The richest sources of benzoxazinoids were Brawura, Łagwa, and Kandela (52.46, 34.67, and 30.14 μg/g dry weight [DW], respectively).
Authors:Sinan Zazoğlu, Beril Anilanmert, Muhammed Aydin, and Salih Cengiz
A fast, reliable, inexpensive, and practical method with a low determination limit and high recovery has been developed for the determination of the marijuana metabolite in routine analysis. THC-COOH in urine was validated using liquid chromatography—tandem mass spectrometry (LC—MS/MS). Before an easy single-step extraction with Toxi-Tubes, basic hydrolysis was performed at 60 °C for 30 min. LC—MS/MS analysis takes 2.5 min for each sample, and the retention time of the analyte is 1.75 min. Specificity, linearity, limit of detection (LOD), limit of quantification (LOQ), accuracy, repeatability, and intermediate precision (inter-day) system suitability parameters were determined in the validation study. The recovery of the extraction method was 88.67 (±5.91). LOD and LOQ values were 1.41 and 5.00 ng mL−1, respectively. The method showed linear response between the values 5.00 and 500.00 ng mL−1. The repeatability was 9.64% (relative standard deviation, RSD%), and the intermediate precisions (RSDR%) were 10.73%, 13.74%, and 8.11% at 10.00, 100.00, and 200.00 ng mL−1 concentration levels, respectively. No statistically significant difference was found in ANOVA analysis, between three consecutive days in intermediate precision study, for 90% confidence level. HorRat values were between 0.34 and 0.61. The method was applied to CEDIA positive samples, obtained from the Trabzon Group Presidency of Turkish Council of Forensic Medicine, successfully.
Authors:X. Luo, C. Han, X. Deng, D. Zhu, Y. Liu, and Y. Yan
Seed germination is a new beginning for the crop life cycle, which is closely related to seed sprouting and subsequent plant growth and development, and ultimately affects grain yield and quality. Salt stress is one of the most important abiotic stress factors that restrict crop production. Therefore, it is highly important to improve crop salt tolerance and sufficient utilization of saline-alkali land. In this study, we identified the phosphorylated proteins involved in salt stress response by combining SEM, 2-DE, Pro-Q Diamond staining and tandem mass spectrometry. The results showed that salt stress significantly inhibited seed germination and starch degradation. In total, 14 phosphorylated protein spots (11 unique proteins) in the embryo and 6 phosphorylated protein spots (4 unique proteins) in the endosperm were identified, which mainly involved in stress/defense, protein metabolism and energy metabolism. The phosphorylation of some proteins such as cold regulated proteins, 27K protein, EF-1β and superoxide dismutase could play important roles in salt stress tolerance.
Palmatine is a compound with good water solubility extracted from Coptis chinensis, Fibraurea recisa Pierre, Cortex Phellodendri Chinensis. Palmatine has good antibacterial activity and mainly used for the treatment of bacterial dysentery, gynecological inflammation, surgical infection, and conjunctivitis. It has anti-diabetic, anti-oxidant, and cognitive-enhancing activities. In this study, we used UPLC-MS/MS to determinate palmatine in rat plasma, and investigated its pharmacokinetics. Coptisine was utilized as an internal standard (IS), and acetonitrile precipitation method was used to process the plasma samples. Chromatographic separation was achieved using a UPLC BEH C18 column using mobile phase of acetonitrile- 0.1% formic acid with gradient elution. Electrospray ionization (ESI) tandem mass spectrometry in multiple reaction monitoring (MRM) mode with positive ionization was applied. The results indicated that within the range of 1–500 ng/mL, linearity of palmatine in rat plasma was acceptable (r > 0.995), and the lower limit of quantification (LLOQ) was 1 ng/mL. Intra-day and inter-day precision RSD of palmatine in rat plasma were less than 14%. Accuracy range was between 93.7 and 107.1%, and matrix effect was between 101.6 and 109.4%. The method was successfully applied in the pharmacokinetics of palmatine in rats after oral and intravenous administration. The absolute bioavailability of the palmatine was 15.5% in rats.