Certification of New Selenium-Enriched Yeast and Supplement Reference Materials for Selenomethionine Using Two Independent Measurement Strategies.

Selenium-enriched yeast possesses the unique ability of transforming chemical selenium, such as sodium selenite, into a biologically active form, which mitigates its toxic effects on the human body. The transformation product of this process, selenomethionine, can be safely and effectively absorbed and utilized by the human body; hence, it has been spiked into a selenium-enriched supplement. This study employs two distinct measurement strategies to determine the selenomethionine content in two candidate reference materials, a selenium-enriched yeast powder and supplement, using both organic and inorganic mass spectrometry. The concentrations of selenomethionine in the selenium-enriched yeast were determined using HPLC-ICP-MS and HPLC- ESI-MS/MS, with mass fractions measured at 718 mg SeMet kg-1 and 715 mg SeMet kg-1, respectively. Notably, both methods yielded consistent results for the selenium supplement, with a selenomethionine mass fraction of 59 mg SeMet kg-1. Ultimately, the certified values of these candidate reference materials were determined as 716 mg kg-1 and 59 mg SeMet kg-1 with expanded uncertainties of 36 mg SeMet kg-1 (k = 2) and 5 mg SeMet kg-1 (k = 2), respectively. The development of these candidate reference materials serves as a valuable reference for diverse methods aiming to determine the value of organic selenium speciation in complex food substrates.

Enhancement of Female Rat Fertility via Ethanolic Extract from Nigella sativa L. (Black Cumin) Seeds Assessed via HPLC-ESI-MS/MS and Molecular Docking.

The characteristic chemical composition of Nigella seeds is directly linked to their beneficial properties. This study aimed to investigate the phytochemical composition of Nigella sativa seeds using a 100% ethanolic extract using HPLC-ESI-MS/MS. Additionally, it explored the potential biological effects of the extract on female rat reproduction. Follicle Stimulating Hormone (FSH), Luteinizing Hormone (LH), Estrogen (E2), and Progesterone (P4) hormone levels were also assessed, along with the morphological and histological effects of the extract on ovarian, oviductal, and uterine tissues. Molecular docking was performed to understand the extract's activity and its role in regulating female reproduction by assessing its binding affinity to hormonal receptors. Twenty metabolites, including alkaloids, saponins, terpenes, flavonoids, phenolic acids, and fatty acids, were found in the ethanolic extract of N. sativa seeds through the HPLC-ESI-MS/MS study. The N. sativa seed extract exhibited strong estrogenic and LH-like activities (p < 0.05) with weak FSH-like activity. Furthermore, it increased the serum levels of LH (p < 0.05), P4 hormones (p < 0.001), and E2 (p < 0.0001). Molecular docking results displayed a strong interaction with Erß, LH, GnRH, and P4 receptors, respectively. Based on these findings, N. sativa seeds demonstrated hormone-like activities, suggesting their potential as a treatment for improving female fertility.

Anthocyanin accumulation and transcriptional regulation in purple flowering stalk (Brassica campestris L. var. purpurea Bailey).

KEY MESSAGE: 1. Purple flowering stalk (Brassica campestris L. ssp. chinensis L. var. purpurea Bailey) is a crop with the high-level anthocyanin. 2. Increased abundance of LBGs promoted the synthesis of anthocyanin. 3. TTG2 (WRKY) interacted with TTG1 (WD40), probably regulating anthocyanin accumulation by shaping a MBWW complex. Brassica crops are a class of nutrient-rich vegetables. Here, two Brassica Crops-Flowering Stalk cultivars, purple flowering stalk (Brassica campestris L. var. purpurea Bailey) and pakchoi (Brassica campestris ssp. chinensis var. communis) were investigated. HPLC-ESI-MS/MS analysis demonstrated that Cy 3-p-coumaroylsophoroside-5-malonylglucoside and Cy 3-diferuloylsophoroside-5-malonylglucoside were identified as the major anthocyanin in peel of purple flowering stalk. The transcript level of structural genes including C4H, CHS, F3H, DFR, ANS and UFGT, and regulatory genes such as TT8, TTG1, Bra004162, Bra001917 and TTG2 in peel of purple flowering stalk were significantly higher than that in peel of pakchoi. In addition, the TTG2(WRKY) interacted only with TTG1(WD40) and the interaction between TT8 (bHLH) and TTG1/Bra004162(MYB)/Bra001917(MYB) were identified. Else, the WD40-WRKY complex (TTG1-TTG2) could activate the transcript of TT12. Our study laid a foundation for the research on the anthocyanin accumulation in Brassica crops.

A simple method for simultaneous determination of organophosphate esters and their diester metabolites in dairy products and human milk by using solid-phase extraction coupled to liquid chromatography-tandem mass spectrometry.

Organophosphate esters (OPEs) and their diester metabolites have been frequently found in various environmental matrices and regarded as emerging environmental pollutants, whereas data on their occurrence in foods and human matrices are still limited. In this study, a novel and simple procedure was developed to simultaneously determine 14 OPEs and 6 diester metabolites in dairy products and human milk. After enzymatic hydrolysis by ß-glucuronidase/arylsulfatase, a freeze-dried milk sample was extracted with acetonitrile and purified by solid-phase extraction. Subsequently, all target compounds were determined by HPLC-ESI-MS/MS. Linearity, limits of detection (LODs), recovery, precision, and matrix effects of the proposed methodology were validated, and the parameters of HPLC-ESI-MS/MS were optimized. LODs for OPEs and their diester metabolites were from 0.001 to 0.02 ng/mL, and limits of quantification (LOQs) were 0.01-0.3 ng/mL. Average recoveries at two spiked levels ranged between 67.3 and 121%, with relative standard deviation lower than 20.7%. A test for matrix effects showed that most analytes presented signal suppression, and isotopically labeled ISs were essential for compensating for the matrix effects. Finally, OPEs and their metabolites both showed high detecting frequencies in real samples, which indicated that these emerging pollutants were ubiquitous in foods and the human body, and the impact of the diester metabolites on population exposure must be included in exposure assessment.

Development of a novel HPLC-ESI-MS/MS method to analyze the stereoselective pharmacokinetics and tissue distribution of isoconazole enantiomers in rats.

Isoconazole with an asymmetrical carbon is a broad-spectrum antimicrobial imidazole, but there is still lack of relevant report about the potential enantioselectivity in biological samples. The object of this research was to develop and validate a sensitive and effective high performance liquid chromatography-electrospray ionization coupled with tandem mass spectrometry (HPLC-ESI-MS/MS) method for stereoselective separation and determination of isoconazole enantiomers in Sprague-Dawley (SD) rat plasma and tissues. The greater enantioseparation of isoconazole enantiomers was obtained on a Chiralpak IC column with a mobile phase consisted of acetonitrile-10 mM aqueous ammonium acetate (90:10, v/v) under the reversed-phase mode. Subsequently, the studied compounds and internal standard (IS) were detected on a multiple reaction monitoring (MRM) mode with positive electrospray ionization source. The experimental and theoretical Electronic Circular Dichroism (ECD) spectra were employed to confirm the absolute configuration of isoconazole enantiomers. Eventually, after full method validation, the newly developed method was successfully applied to the study of enantioselectivity in plasma and tissues in SD rats. Results illustrated that the enantioselective differences in plasma were observed for the evidence that the concentrations of S-(-)-isoconazole were always higher than R-(+)-isomer. In terms of tissue distribution, liver, kidney, lung, spleen, and small intestine were the mainly distributed tissues and then followed by heart and muscle. This is the first study to reveal the stereoselective behavior of isoconazole enantiomers in vivo, which also provides reliable and valuable reference for further elucidating the enantioselective metabolisms of isoconazole enantiomers.

In vitro wound healing properties, antioxidant activities, HPLC-ESI-MS/MS profile and phytoconstituents of the stem aqueous methanolic extract of Dracaena reflexa Lam.

Column chromatography of the stem aqueous methanolic extract of Dracaena reflexa Lam. (DRSE) led to the isolation of five flavonoids, one phenolic glycoside, one triterpenoid and two steroidal saponins. Furthermore, 44 compounds were tentatively identified in the phytoconstituent profile of DRSE using HPLC-ESI-MS/MS. The antioxidant activity of DRSE was evaluated. In a DPPH radical scavenging assay, DRSE exhibited an IC50 value of 311.6 ± 10.10 µg/ml compared with the IC50 value of the standard Trolox (24.42 ± 0.87 µg/ml). The antioxidant activities of DRSE using ABTS assay and ferric reducing antioxidant power assay were 326.63 µm Trolox equivalents/mg extract and 208.67 µm Trolox equivalents/mg extract, respectively. The wound-healing activity of DRSE was studied by the scratch assay using Human Skin Fibroblast cells. After 24 h DRSE (at 10 and 20 µg/ml) decreased the wound width to 0.55 ± 0.37 and 0.47 ± 0.55 mm, respectively, compared with the wound width in the control cells (0.77 ± 0.17 mm). This result suggested that DRSE improved the wound-healing process by inducing the migration of fibroblasts. Moreover, a docking study was performed to evaluate the binding affinity of the identified phytoconstituents toward GSK-3ß relative to the co-crystalized inhibitor and curcumin with the possible involvement of this pathway in the wound-healing activity of the extract.

Determination and Removal of Selected Pharmaceuticals and Total Organic Carbon from Surface Water by Aluminum Chlorohydrate Coagulant.

In the present research, the removal of Total Organic Carbon (TOC) and erythromycin (ERY), fluoxetine (FLX), amoxicillin (AMO), colistin (COL), ethynylestradiol (EE), and diclofenac (DIC) from surface water by coagulation is studied. The concentration of selected pharmaceuticals in 24 surface water samples originating from some rivers located in Lesser Poland Voivodeship and Silesia Voivodeship, Poland, was determined. The removal of TOC and pharmaceuticals was carried out using the application of Design of Experiments (DOE), Response Surface Methodology (RSM), and by addition of aluminum chlorohydrate (ACH) as a coagulant. The study found that the concentration ranges of ERY, FLX, AMO, COL, EE, and DIC in analyzed water samples were 7.58−412.32, 1.21−72.52, 1.22−68.55, 1.28−32.01, 5.36−45.56, 2.20−182.22 ng/L, respectively. In some cases, concentrations lower than 1 ng/L were determined. In optimal conditions of coagulation process of spiked surface water (pH = 6.5 ± 0.1, ACH dose = 0.35 mL/L, Time = 30 min; R2 = 0.8799, R2adj = 0.7998), the concentration of TOC, ERY, FLX, AMO, COL, EE, and DIC was decreased by 88.7, 36.4, 24.7, 29.0, 25.5, 35.4, 30.4%, respectively. Simultaneously, turbidity, color, Total Suspended Solids (TSS), Chemical Oxygen Demand (COD), Total Nitrogen (Total N), and Ammonium-Nitrogen (N-NH4) were decreased by 96.2%, >98.0%, 97.8%, 70.0%, 88.7%, 37.5%, respectively. These findings suggest that ACH may be an optional reagent to remove studied pharmaceuticals from contaminated water.

Targeted phenolic profile of radler beers by HPLC-ESI-MS/MS: the added value of hesperidin to beer antioxidants.

The well-known health beneficial properties of beer are mainly due to phenolic antioxidants. Citrus-flavored beers represent a growing side-market in the beer industry, sparingly investigated to date. The phenolic profile of commercial radler beers (R1, R2) was investigated to evaluate the impact of the lemon juice added to beer in the industrial production. Results were compared to those obtained for opportunely chosen commercial beer (B) and lemonade (L). The study was carried out by an HPLC-MS/MS with an electrospray ionization source in selected ion recording mode, analyzing in a single chromatographic run 26 compounds belonging to the different phenolic classes of hydroxybenzoic, hydroxycinnamic and caffeoylquinic acids, flavonoids and prenylflavonoids. Different phenolic profiles were found for R1 and R2, mainly ascribed to different malt/hop/recipe used for the beer. High to very high level of hesperidin were found in the radlers, so that a major impact on phenolic antioxidants of the radlers was due to the lemon. Similarly, a major impact of the lemon aromas was found, D-limonene being the dominant peak resulting from the GC-MS analysis of the volatile fraction of the radlers.

Biodegradation and utilization of crop residues contaminated with poisonous pyrrolizidine alkaloids.

Disposal of noxious plant residues is a challenge for farmers and land management dealing with contaminated biomasses. Recent studies confirm the potential threat of transferring toxic plant constituents like pyrrolizidine alkaloids (PAs) from plant residues to non-toxic succeeding agricultural crops via the soil. We studied the degree of biochemical degradation of PAs in the two most important processes, composting and biomethanization. We used lab composting and biogas batches to investigate the potential of PA-degradation of two common PA-containing plants, Lappula squarrosa and Senecio jacobaea. The experiments demonstrated a virtually complete loss of PAs in three months during the composting process and a rapid decomposition of PAs from 3112.6 µg/kg to less than 21.5 µg/kg in L. squarrosa and from 6350.2 µg/kg to less than 539.6 µg/kg in S. jacobaea during biomethanization. The information obtained is a first guide on how to re-utilize PA-contaminated plant matter in a circular bioeconomy.

Isolation of bioactive components with soluble epoxide hydrolase inhibitory activity from Stachys sieboldii MiQ. by ultrasonic-assisted extraction optimized using response surface methodology.

Stachys sieboldii MiQ (SSM) is an important food and medicinal herb in Korea, used to improve memory of patients with senile dementia and cardiovascular diseases. However, little information on bioactive components from SSM or standardized extraction methods for these components is available. This study isolated and purified major components from SSM for the first time, and assessed their ability to inhibit soluble epoxide hydrolase (sEH). The results showed that acteoside is the most potent inhibitor of sEH, with an IC50 of 33.5 ± 0.5 µM. Additional active components, including harpagide, tryptophan, and 8-acetate-harpagide, along with acteoside, were tentatively identified using high-performance liquid chromatography photodiode array tandem mass spectrometry (HPLC-PDA-MS/MS) and quantified using an ultraviolet detector at 210 nm. Further, an ultrasonic-assisted extraction technique for extraction of four bioactive compounds in SSM was developed and optimized using response surface methodology (RSM). The optimal extraction conditions were: extraction time, 30.46 minutes; extraction temperature, 67.95 °C, and methanol concentration 53.85%. The prediction model of RSM was validated with laboratory experiments. The similarity between predicted and actual values was 97.84%. The extraction method is thus a rapid, environment-friendly, energy-saving method can be applied to extract bioactive components from SSM in large quantities.

Phytochemical Investigation of New Algerian Lichen Species: Physcia Mediterranea Nimis.

The present study provides new data concerning the chemical characterisation of Physcia mediterranea Nimis, a rare Mediterranean species belonging to the family Physciaceae. The phytochemical screening was carried out using GC-MS, HPLC-ESI-MS-MS, and NMR techniques. Hot extraction of n-hexane was carried out, followed by separation of the part insoluble in methanol: wax (WA-hex), from the part soluble in methanol (ME-hex). GC-MS analysis of the ME-hex part revealed the presence of methylbenzoic acids such as sparassol and atraric acid and a diterpene with a kaurene skeleton which has never been detected before in lichen species. Out of all the compounds identified by HPLC-ESI-MS-MS, sixteen compounds are common between WA-hex and ME-hex. Most are aliphatic fatty acids, phenolic compounds and depsides. The wax part is characterised by the presence of atranorin, a depside of high biological value. Proton 1H and carbon 13C NMR have confirmed its identification. Atranol, chloroatranol (depsides compound), Ffukinanolide (sesquiterpene lactones), leprolomin (diphenyl ether), muronic acid (triterpenes), and ursolic acid (triterpenes) have also been identified in ME-hex. The results suggested that Physcia mediterranea Nimis is a valuable source of bioactive compounds that could be useful for several applications as functional foods, cosmetics, and pharmaceuticals.

Cytokinin activity during early kernel development corresponds positively with yield potential and later stage ABA accumulation in field-grown wheat (Triticum aestivum L.).

MAIN CONCLUSION: Early cytokinin activity and late abscisic acid dynamics during wheat kernel development correspond to cultivars with higher yield potential. Cytokinins represent prime targets for marker development for wheat breeding programs. Two major phytohormone groups, abscisic acid (ABA) and cytokinins (CKs), are of crucial importance for seed development. Wheat (Triticum aestivum L.) yield is, to a high degree, determined during the milk and dough stages of kernel development. Therefore, understanding the hormonal regulation of these early growth stages is fundamental for crop-improvement programs of this important cereal. Here, we profiled ABA and 25 CK metabolites (including active forms, precursors and inactive conjugates) during kernel development in five field-grown wheat cultivars. The levels of ABA and profiles of CK forms varied greatly among the tested cultivars and kernel stages suggesting that several types of CK metabolites are involved in spatiotemporal regulation of kernel development. The seed yield potential was associated with the elevated levels of active CK levels (tZ, cZ). Interestingly, the increased kernel cZ levels were followed by higher ABA production, suggesting there is an interaction between these two phytohormones. Furthermore, we analyzed the expression patterns of representatives of the four main CK metabolic gene families. The unique transcriptional patterns of the IPT (biosynthesis) and ZOG (reversible inactivation) gene family members (GFMs) in the high and low yield cultivars additionally indicate that there is a significant association between CK metabolism and yield potential in wheat. Based on these results, we suggest that both CK metabolites and their associated genes, can serve as important, early markers of yield performance in modern wheat breeding programs.

Biodegradation of phenol by Chlamydomonas reinhardtii.

The data presented in this particular study demonstrate that the biodegradation of phenol by Chlamydomonas reinhardtii is a dynamic bioenergetic process mainly affected by the production of catechol and the presence of a growth-promoting substrate in the culture medium. The study focused on the regulation of the bioenergetic equilibrium resulting from production of catechol after phenol oxidation. Catechol was identified by HPLC-UV and HPLC-ESI-MS/MS. Growth measurements revealed that phenol is a growth-limiting substrate for microalgal cultures. The Chlamydomonas cells proceed to phenol biodegradation because they require carbon reserves for maintenance of homeostasis. In the presence of acetic acid (a growth-promoting carbon source), the amount of catechol detected in the culture medium was negligible; apparently, acetic acid provides microalgae with sufficient energy reserves to further biodegrade catechol. It has been shown that when microalgae do not have sufficient energy reserves, a significant amount of catechol is released into the culture medium. Chlamydomonas reinhardtii acts as a versatile bioenergetic machine by regulating its metabolism under each particular set of growth conditions, in order to achieve an optimal balance between growth, homeostasis maintenance and biodegradation of phenol. The novel findings of this study reveal a paradigm showing how microalgal metabolic versatility can be used in the bioremediation of the environment and in potential large-scale applications.

Phenolic Fingerprinting, Antioxidant, and Deterrent Potentials of Persicaria maculosa Extracts.

Persicaria maculosa (Polygonaceae) (known as lady's thumb) is an annual morphologically variable weed that is widely distributed in Chile. The purpose of this study was to investigate the antifeedant potential of methanolic (MeOH), ethanolic (EtOH), and dichloromethane (DCM) extracts from the aerial parts of this plant collected in the Valparaíso and Curicó provinces (Chile) and relate this activity to the antioxidant capacity and the presence of phenolic compounds in the extracts. A phenolic profile based on HPLC-ESI-MS/MS allowed the identification of 26 phenolic compounds, most of them glycosyl derivatives of isorhamnetin, quercetin, and kaempferol. In addition, the total phenolic content (TP), total flavonoids (TF), and antioxidant activity measured by 2,2-diphenyl-1-picrylhydrazyl (DPPH), superoxide anion scavenging (O2-), ferric-reducing antioxidant power (FRAP), and cupric-reducing antioxidant capacity (CUPRAC) of the extracts are reported. The antifeedant potentials of the plant extracts were tested against Epilachna paenulata, Pseudaletia adultera, Macrosiphum euphorbiae, and Diaphorina citri insects for the first time. The activity against the aphid M. euphorbiae was significant for the DCM extracts of plants from Valparaíso and Curicó (settling % = 23% ± 4% and 23% ± 5%, respectively). The antifeedant activities against the beetle E. paenulata and the lepidoptera P. adultera were significant for Valparaíso extracts, especially when tested against E. Paenulata (IFP = 1.0 ± 0.0). Finally, the MeOH and EtOH extracts from Valparaíso plants reduced the diet consumption of the psilid D. citri (p < 0.05). The results showed that P. maculosa is a good source of flavonoids with some antioxidant capacities and has potential interest as botanical eco-friendly alternative with deterrent activity.

Reductive Activity and Mechanism of Hypoxia- Targeted AGT Inhibitors: An Experimental and Theoretical Investigation.

O6-alkylguanine-DNA alkyltransferase (AGT) is the main cause of tumor cell resistance to DNA-alkylating agents, so it is valuable to design tumor-targeted AGT inhibitors with hypoxia activation. Based on the existing benchmark inhibitor O6-benzylguanine (O6-BG), four derivatives with hypoxia-reduced potential and their corresponding reduction products were synthesized. A reductase system consisting of glucose/glucose oxidase, xanthine/xanthine oxidase, and catalase were constructed, and the reduction products of the hypoxia-activated prodrugs under normoxic and hypoxic conditions were determined by high-performance liquid chromatography electrospray ionization tandem mass spectrometry (HPLC-ESI-MS/MS). The results showed that the reduction products produced under hypoxic conditions were significantly higher than that under normoxic condition. The amount of the reduction product yielded from ANBP (2-nitro-6-(3-amino) benzyloxypurine) under hypoxic conditions was the highest, followed by AMNBP (2-nitro-6-(3-aminomethyl)benzyloxypurine), 2-NBP (2-nitro-6-benzyloxypurine), and 3-NBG (O6-(3-nitro)benzylguanine). It should be noted that although the levels of the reduction products of 2-NBP and 3-NBG were lower than those of ANBP and AMNBP, their maximal hypoxic/normoxic ratios were higher than those of the other two prodrugs. Meanwhile, we also investigated the single electron reduction mechanism of the hypoxia-activated prodrugs using density functional theory (DFT) calculations. As a result, the reduction of the nitro group to the nitroso was proven to be a rate-limiting step. Moreover, the 2-nitro group of purine ring was more ready to be reduced than the 3-nitro group of benzyl. The energy barriers of the rate-limiting steps were 34-37 kcal/mol. The interactions between these prodrugs and nitroreductase were explored via molecular docking study, and ANBP was observed to have the highest affinity to nitroreductase, followed by AMNBP, 2-NBP, and 3-NBG. Interestingly, the theoretical results were generally in a good agreement with the experimental results. Finally, molecular docking and molecular dynamics simulations were performed to predict the AGT-inhibitory activity of the four prodrugs and their reduction products. In summary, simultaneous consideration of reduction potential and hypoxic selectivity is necessary to ensure that such prodrugs have good hypoxic tumor targeting. This study provides insights into the hypoxia-activated mechanism of nitro-substituted prodrugs as AGT inhibitors, which may contribute to reasonable design and development of novel tumor-targeted AGT inhibitors.

Development and Validation of an HPLC-ESI/MS/MS Method for the Determination of Amoxicillin, Its Major Metabolites, and Ampicillin Residues in Chicken Tissues.

A method for the simultaneous analysis of amoxicillin (AMO), amoxicillin metabolites, and ampicillin residues in edible chicken muscle, liver, and kidney samples via high-performance liquid chromatography-electrospray ionization tandem mass spectrometry (HPLC-ESI/MS/MS) was developed and verified. The extraction and purification procedures involved the extraction of the sample using a liquid-liquid extraction method with acetonitrile to eliminate the proteins. The chicken tissue extract was then injected directly onto an HPLC column coupled to a mass spectrometer with an ESI(+) source. The HPLC-ESI/MS/MS method was validated according to specificity, sensitivity, linearity, matrix effects, precision, accuracy, decision limit, detection capability, and stability, as defined by the European Union and Food and Drug Administration. The linearity was desirable, and the determination coefficients (r2 values) ranged from 0.9968 and 0.9999. The limits of detection and limits of quantification were 0.10-2.20 µg/kg and 0.30-8.50 µg/kg, respectively. The decision limits were 57.71-61.25 µg/kg, and the detection capabilities were 65.41-72.50 µg/kg, and the recoveries of the four target analytes exceeded 75% at the limits of quantification and exceeded 83% at 25, 50, and 100 µg/kg (n = 6 at each level), confirming the reliability of this method for determining these analytes and providing a new detection technology. For real sample analysis, this experiment tested 30 chicken tissue samples, only one chicken muscle, liver, and kidney sample were contaminated with 5.20, 17.45, and 7.33 µg/kg of AMO values, respectively, while other target compounds were not detected in the 30 tested chicken tissue samples.

Capillary-HPLC with tandem mass spectrometry in analysis of alkaloid dyestuffs - a new approach.

Development of the identification method of alkaloid compounds in Amur cork tree as well as not examined so far Oregon grape and European Barberry shrubs are presented. The novel approach to separation of alkaloids was applied and the capillary-high-performance liquid chromatography (capillary-HPLC) system was used, which has never previously been reported for alkaloid-based dyestuffs analysis. Its optimization was conducted with three different stationary phases (unmodified octadecylsilane-bonded silica, octadecylsilane modified with polar groups and silica-bonded pentaflourophenyls) as well as with different solvent buffers. Detection of the isolated compounds was carried out using diode-array detector (DAD) and tandem mass spectrometer with electrospray ionization (ESI MS/MS). The working parameters of ESI were optimized, whereas the multiple reactions monitoring (MRM) parameters of MS/MS detection were chosen based on the product ion spectra of the quasi-molecular ions. Calibration curve of berberine has been estimated (y = 1712091x + 4785.03 with the correlation coefficient 0.9999). Limit of detection and limit of quantification were calculated to be 3.2 and 9.7 ng/mL, respectively. Numerous alkaloids (i.e., berberine, jatrorrhizine and magnoflorine, as well as phellodendrine, menisperine and berbamine) were identified in the extracts from alkaloid plants and silk and wool fibers dyed with these dyestuffs, among them their markers.

Protective effect of Zizyphus lotus jujube fruits against cypermethrin-induced oxidative stress and neurotoxicity in mice.

CONTEXT: Cypermethrin (CYP) is a synthetic pyrethroid insecticide used worldwide in agriculture, home pest control. The toxicity of CYP is well studied in many organisms. OBJECTIVE: The aim of present study was to investigate the protective effect of Zizyphus lotus (Zizyp) fruit against neurotoxicity and oxidative stress induced by CYP in mice. MATERIALS AND METHODS: Mice were divided into four groups of six each: groups I and II were used as control and CYP control (20 mg/kg body weight). While, groups III was orally treated with Zizyphus lotus fruit (5 g/kg body weight) plus CYP (20 mg/kg body weight) for 18 days. Furthermore, HPLC-ESI-MS-MS (Q-Tof) and GC-MS were used to identify the compounds fraction. RESULTS: Antioxidant enzyme catalase (CAT), neurotoxicity enzyme acetylcholinesterase (AChE) activities and hydrogen peroxide (H2O2), malondialdehyde (MDA) levels were determined in the liver, kidney and heart. CYP caused decreased CAT activity, inhibition of AChE activity and increased the levels of H2O2 and MDA in heart, liver and kidney. CONCLUSION: Our results indicate that Zizyp fruit is markedly effective in protecting mice against CYP-induced biochemical changes. This protection may be due to its antioxidant property and scavenging ability against active free radicals.

Development of an LC-MS/MS-based assay to determine artemitin in rat plasma and its application in a pharmacokinetic study.

Artemitin, a significant flavonol compound existing in Laggera pterodonta (DC.) Benth., Artemisia rupestris L, etc., is the subject of attention by researchers owing to its pharmacological activities (such as antioxidative, anti-inflammatory and antiviral). In this work, a highly sensitive and specific high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (HPLC-ESI-MS/MS) assay combined with protein precipitation has been established and validated for determining artemitin concentration in rat plasma. Both artemitin and warfarin sodium (internal standard, IS) were separated on an Agela Venusil XBP Phenyl column through the isocratic elution mode of methanol-water containing 0.1% formic acid (80:20, v/v), at a flow rate of 0.4 mL/min. The MS/MS system was operated in a positive ion and ESI multiple reaction monitoring mode, and the multiple reaction monitoring transition was optimized as m/z 389.0 → 373.0 for artemitin and 309.2 → 163.0 for IS. The method showed good linearity in the range of 2.5-2000 ng/mL (R2 = 1.0000) and high sensitivity for artemitin with the lower limit of quantification of 2.5 ng/mL. The intra- and inter-day accuracies were 97.4-100.9 and 93.4-100.3%, respectively. The intra- and inter-day precisions were <4.8 and 6.5%, respectively. The extraction efficiency and absolute recovery were >66.5 and 71.3%, respectively. In addition, a good matrix effect of <9.5% was obtained. As a result, the method developed herein was successfully applied for the pharmacokinetic study of artemitin after an intravenous administration in rats.

Application of a sensitive and specific LC-ESI-MS/MS method for the simultaneous quantification of twelve bioactive components in dog plasma for an intravenous pharmacokinetic study of Yiqifumai Injection in beagle dogs.

Yiqifumai Injection is a lyophilized powder preparation widely used to treat coronary heart disease. However, its in vivo bioactive components and pharmacokinetic behavior remain unknown. Therefore a sensitive and specific LC-MS/MS was developed and validated for the simultaneous quantification of eight saponins and four lignans in beagle dog plasma. The plasma samples were pretreated by protein precipitation with methanol-acetonitrile (1:1, v/v). Chromatographic separation of all the 12 analytes and estazolam (internal standard, IS) was successfully accomplished on an Ultimate® XB-C8 column (100 × 2.1 mm, 3 µm) with a gradient elution system. The total running time was 8 min with a flow rate of 0.40 mL/min. Acquisition of mass spectrometric data was performed via positive electrospray ionization in multiple reaction monitoring mode. The assay was fully validated in terms of selectivity, linear range, lower limit of quantitation, precision, accuracy, matrix effect, recovery and stability. This validated method was successfully applied to the pharmacokinetics of 12 bioactive components after intravenous administration of Yiqifumai Injection to beagle dogs at a dose of 0.541 g/kg.