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The study evaluated the therapeutic potential of ethanolic leaf extract of Piliostigma foveolatum (Dalzell) Thoth. (EEBF), its toluene, ethylacetate, methanol soluble fractions (viz. TFBF, EFBF, MFBF), and isolated phytoconstituents against lung cancer. Four compounds were isolated from MFBF by column chromatography and preparative HPLC. Structures were elucidated by IR, 13C-NMR, 1H-NMR, mass spectroscopy and identified as Quercetin, Kaempferol, Isorhamnetin, and ß-glucogallin. EEBF and its biofractions exhibited remarkable antiproliferative activity with GI50<85µg/mL, while isolated Quercetin, Kaempferol, Isorhamnetin, and ß-Glucogallin displayed GI50 values of 56.15 ± 1.16 μ M, 68.41 ± 3.98 μ M, 55.08 ± 0.57 μ M and 58.99 ± 12.39 μ M respectively. MFBF demonstrated significant apoptotic activity with 42.24 ± 0.57% cells in early and 4.61 ± 0.88% cells in late apoptosis comparable to standard Doxorubicin. Kaempferol exhibited 23.03 ± 0.37% cells in early and 2.11 ± 0.55% cells in late apoptosis, arresting Hop-62 cells in S-phase. In silico molecular docking, revealed that isolated constituents effectively bound to the same binding site of caspase-3 as Doxorubicin, highlighting their apoptotic mode of action. [ABSTRACT FROM AUTHOR]

Passiflora edulis Var. flavicarpa (passion fruit) generates vast waste (60–70%) in the form of peel and seed after the juice extraction. The study aimed to isolate Scirpusin B (SB) from passion fruit (PF) seed waste collected from Northeast India and to analyse its anti-radical, antibacterial, anti-diabetic, and anti-oral cancer activities. Scirpusin B was isolated following hydro-alcoholic extraction, fractionation, and column chromatography. The isolated fraction was further identified through NMR and mass spectroscopy. SB exhibited significant antiradical activity against six standard antioxidant compounds, indicating its commercial application. SB inhibited α-amylase (IC50 Value: 76.38 ± 0.25 µg/mL) and α-glucosidase digestive enzymes (IC50 Value: 2.32 ± 0.04 µg/mL), signifying its antidiabetic properties. In addition, SB showed profound antibacterial activity against eight gram-positive and gram-negative bacteria reported for the first time. Furthermore, SB inhibited SAS and TTN oral cancer cell proliferation up to 95% and 83%, respectively. SB significantly inhibited colonies of SAS and TTn cells in the clonogenic assay, attributing to its anticancer properties. The PI-FACS assay confirmed the ability of SB (75 µM) to kill SAS and TTn cells by 40.26 and 44.3% in 72 h. The mechanism of SB inhibiting oral cancer cell proliferation was understood through western blot analysis, where SB significantly suppressed different cancer hallmark proteins, such as TNF-α, survivin, COX-2, cyclin D1, and VEGF-A. The present study suggests that SB isolated from PF seed can add noteworthy value to the waste biomass for various industrial and medical applications. [ABSTRACT FROM AUTHOR]

This article proposes a solvent system screening strategy for compounds with similar UV absorption in complex samples by UV spectrophotometer. There is no need to calculate the partition coefficient value of each compound, only the partition coefficient of the whole sample. The partition coefficient value should be close to 1 in order to obtain as many high‐speed counter‐current chromatography fractions as possible. Then, preparative HPLC was used to purify the high‐speed counter‐current chromatography fractions. Based on the above strategy, seven c‐glycosyl flavonoids and an amino acid were successfully obtained from barley seedlings through high‐speed counter‐current chromatography fractionation with ethyl acetate/n‐butanol/water (8:2:10, v:v:v) system followed by preparative HPLC purification. The research shows that high‐speed counter‐current chromatography could be well developed as a tool for fractionation before purification, and greatly improves the separation efficiency. [ABSTRACT FROM AUTHOR]

Herein, two iridoid glucosides aucubin (1) and ajugol (2), and two phenyl ethanoids, verbascoside (3) and poliumoside (4) were isolated from the methanol extract of the aerial parts of Verbascum speciosum and used to study about their anticancer activity for the first time. The structures of all compounds were elucidated using spectroscopic data (IR, 1D and 2D NMR, LC‐TOF/MS). Antiproliferative activities of Aucubun (1) and Verbascoside (3) were tested against A‐549 (human colon cancer), MDA‐MD‐453 (human breast cancer) and 3T3‐L1 (mouse fibroblast)cell lines by XTT assay. In addition, the anticarcer mechanism of action of aucubin (1) was investigated on MDA‐MB‐453 cells for the first time. XTT result showed that both applied compounds exhibited antiproliferative effect at different dose ranges depending on the cancer type, as well as selectivity between cancer and healty cell lines. Flow cytometry analyzes revealed that aucubin (1) exerts its cytotoxic effect in MDA‐MB‐453 cells by directing cells to early apoptosis and inhibiting the P13K/AKT signaling pathway. [ABSTRACT FROM AUTHOR]

Drug impurities are important factors that affect drug safety and efficacy. The aim of this study is to separate and confirm the structure of two degradation impurities of esomeprazole sodium, designated X and Y. The impurities X and Y were successfully isolated using preparative HPLC by developing separation methods with the help of ACD/Labs AutoChrom software. There was a steady increase in X and Y impurities in forced esomeprazole sodium degradation. Impurity X was confirmed as 6‐methoxy‐1h‐benzo[d]imidazole‐2‐yl‐4‐amino‐3,5‐dimethylpyridinecarboxylate and impurity Y as 6‐methoxy‐1h‐benzo[d]imidazole‐2‐yl‐4‐hydroxy‐3,5‐dimethylpyridinecarboxylate using nuclear magnetic resonance spectrometry, infrared spectroscopy, and high‐resolution mass spectrometry. These findings provide a comprehensive understanding of the impurity profile of esomeprazole sodium because these impurities are reported for the first time. Based on our results, active pharmaceutical ingredient manufacturers can further control process parameters to reduce impurity generation, and drug production manufacturers can optimize the packaging and storage conditions of esomeprazole sodium. [ABSTRACT FROM AUTHOR]

This study provides a rapid and accurate method for screening steroid 5-alpha reductase (S5αR) inhibitors in Impatiens balsamina Linn (IB). using at-line LC-QTOF-ESI-MS/MS coupling S5αR inhibitory assay. IB (Balsaminaceae) is an annual herbaceous plant cultivated in tropical and subtropical regions. It has been used in traditional Chinese and Thai medicine for treatment of hair loss and various skin conditions, potentially through anti-androgenic mechanisms. A combined approach of S5αR inhibitory assay and LC-QTOF-ESI-MS/MS was developed to rapidly screen for target biomarkers and guide their isolation using preparative HPLC. The toxicity of both the extract and isolated biomarkers was evaluated on skin cells, keratinocytes, and fibroblasts. Eight bioactive compounds were identified as two naphthoquinone, two fatty acid derivatives, three nitrogenous compounds and one aromatic derivative. The most potent bioactive markers, identified as 2-methoxy-1,4-naphthoquinone (2MN) and impateinol, were targeted and isolated using preparative HPLC, yielding 5.0 % and 3.5 %, respectively. These compounds exhibited S5αR inhibitory activity higher than that of finasteride drug by 10 and 2 times, respectively. Both the isolated biomarkers and the extract demonstrated a broad therapeutic index. The developed method in this study proved to be both rapid and accurate, making it suitable for screening and targeting S5αR inhibitors in herbal plants or complex matrix samples. It facilitated the fast-guided isolation of bioactive compounds, highlighting its potential for future applications in drug discovery research., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Introduction: Reactive oxygen species (ROS) and ultraviolet (UV) light are significant factors to impair skin disorders. Impatiens balsamina Linn. (IB), a traditional Chinese and Thai herbal medicine, has long been used to treat skin and nail diseases, potentially due to its radical-scavenging properties. However, specific antioxidant compounds in IB have not been well defined., Objective: This work aims to rapidly identify, target, and isolate antioxidant biomarkers in IB using at-line LC-ESI-QTOF-(MS/MS) coupled with a simple DPPH assay and comprehensively investigate the antioxidant activities of IB extract and isolated biomarker., Methodology: Following liquid chromatography (LC), the eluent of IB extract was split into two streams (9:1 ratio). The majority was fractionated for DPPH assay in 96-well plates, whereas 10% underwent chemical identification using ESI-QTOF-MS. Antioxidants in IB were identified, targeted, and promptly isolated through transfer from analytical LC to preparative HPLC. IB and the isolated biomarkers were evaluated for antioxidant effects using various antiradical assays and in suppressing ROS induced by UV in skin cells, keratinocytes, and fibroblasts., Results: Thirty-one chemical constituents were identified, with four tentatively identified as potent antioxidants. Kaempferol emerged as a potential antioxidant biomarker in IB, exhibiting superior antioxidant activity in various in vitro assays compared with positive controls. Both IB extract and kaempferol effectively reduced UVB-induced ROS in skin cells., Conclusion: This study represents the first comprehensive identification of antioxidants and chemical constituents in IB, pinpointing kaempferol as a key antioxidant biomarker. Its rapid identification using at-line techniques holds promise for advancing bioactive compound discovery in herbal medicine., (© 2024 John Wiley & Sons Ltd.)

A bio-guided selection and identification of the most active compounds obtained from Algerian Pistacia atlantica desf. Subsp. atlantica. An antioxidant activity guided fractionation was performed on buds' extract using extensive chromatographic and spectroscopic techniques. The antioxidant potentials of isolated compounds and other unpurified fractions were evaluated against DPPH radicals using TLC plates and test tubes. The results showed that all isolated compounds and fractions exhibited eminent DPPH scavenging potential. Two coumarins (7-ethoxycoumarin and 7-hydroxy-5-methoxycoumarin) and two flavonoids (3′,5,7-trihydroxy-4′-methoxyflavanone and 5,6,7,4′-tetrahydroxyflavonol-3-O-rutinoside) were isolated for the first times from the titled subspecies. These results confirm that the species of P. atlantica is far from being exhausted of active compounds, especially polyphenols. [ABSTRACT FROM AUTHOR]

• Five polyphenols from apple leaf were successfully isolated by HSCCC-prep-HPLC. • Optimization of HSCCC conditions was realized by response surface methodology. • The experimental value was in good consistence with the predicted value. • Polyphenols except phloridzin showed significant neuronal protection effect. Considering comprehensive utilization of natural products, isolation and activity determination processes of bioactive compounds are essential. In this study, a combined high-speed countercurrent chromatography (HSCCC) with preparative HPLC method was developed to isolate the five antioxidant polyphenols from 75% ethanol extract of Malus pumila Mill. leaves. The HSCCC conditions were optimized by response surface methodology (RSM) considering two response indexes including retention of stationary phase and analysis time. The optimal HSCCC conditions were flow rate of 2.11 mL/min, revolution speed of 717 rpm, and temperature of 25℃, with a solvent system of ethyl acetate/methanol/water (10:1:10, v/v/v). The unseparated fractions obtained from HSCCC were subjected to preparative HPLC for further isolation. As a result, phloridzin (15.3 mg), isoquercitrin (2.1 mg), quercetin 3- O -xyloside (1.9 mg), quercetin-3- O -arabinoside (4.0 mg), and quercitrin (2.0 mg) were isolated from 200.0 mg extracts. The purities of these compounds were all above 92%. Their chemical structures were identified by mass spectrometer and nuclear magnetic resonance. The five isolated compounds were further investigated for their rat hippocampal neuroprotective effects against hydrogen peroxide-induced oxidative stress. No cytotoxicity was observed in all tested concentrations. While all five compounds except phloridzin showed significantly neurogenic activities and neuroprotective effects, especially at the concentration of 0.5 mg/L. These results demonstrate that RSM is a suitable technique for optimisation of HSCCC and the isolated polyphenols can be used as antioxidants in pharmaceutical and food products. [ABSTRACT FROM AUTHOR]

[Display omitted] • Purification of individual anthocyanins from black carrots. • Higher stability of non-acylated anthocyanins than acylated anthocyanins when exposed to light. • Acylated anthocyanins was found to undergo cleavage of the glycosidic bond. Black carrot anthocyanins have gained increasing attention as natural coloring agent, owing to their higher stability than anthocyanins from berries. The stability has been attributed to their higher degree of acylation. This study investigated the impact of acylation on the stability of individual anthocyanins during storage in light and darkness. We hypothesized that the acylated anthocyanins would be more stable than the non-acylated ones. The major five anthocyanins were fractioned by semi-preparative HPLC and stored at pH 4.5 in light and darkness to investigate how acylation affected the stability. The stability was evaluated by absorption spectroscopy and mass spectrometry (MS). Two of the anthocyanins were non-acylated; 3-xylosyl(glucosyl)galactoside and cyanidin 3-xylosylgalactoside, and three were acylated; cyanidin 3-xylosyl(sinapolyglucosyl)galacto-side, cyanidin 3-xylosyl(feruloylglu-cosyl)galactoside, and cyanidin 3-xylosyl(coumaroyl-glucosyl)galactoside. Both methods (spectroscopy and MS) showed a clear effect of acylation when stored in light, but surprisingly the two non-acylated anthocyanins, showed higher stability than the three acylated ones. [ABSTRACT FROM AUTHOR]

As per ICH Q1A (R2), stress testing (i.e., acidic, basic, oxidative, thermolytic, hydrolytic, and photolytic) was carried out at room temperature with a single batch of Sacubitril Sodium of the drug substance. Though the drug is resistant to oxidation, thermal, and photolysis conditions, two degradation products were formed during base stress. By using gradient preparative HPLC and a normal C-18 column, the degradants were separated, and the obtained SAC D-1 and SAC D-2 were precisely characterized by LCMS and extensive NMR (including 2D) spectroscopic methods. Study of NMR spectra (1D, 2D) and Mass spectrometry confirmed the suggested structures for the products 5-([1,1'-biphenyl]-4-yl)-4-(3-carboxy propanamide)-2-methyl pentatonic acid (SAC D-1) and (3S)-5-([1,1'-biphenyl]-4-ylmethyl)-3-methyl pyrrolidin-2-one (SAC D-2). [ABSTRACT FROM AUTHOR]

The current study was concentrated to detection of anti-cancer compound from marine Streptomyces akiyoshiensis GRG 6 (KY457710) against MCF-7 breast cancer cells. The partial purification of TLC was moved three different active spots. Among the spots, the third spot of the extract was shown excellent anti-cancer properties against MCF-7 human breast cancer cells. The available anti-cancer compounds present in the partially purified extract was confirmed by GC–MS data interpretation. Three fractions of the compounds were purified by preparative HPLC and their respective spectrum was confirmed by analytical HPLC. Active anti-cancer fraction was separately purified by preparative HPLC and the cytotoxicity result was shown with 100 µg/mL concentration. The anti-cancer compound of pyrrolo [1,2-a]pyrazine-1,4-dione, hexahydro-3 was detected by LC-MS analysis after interpretation with GC–MS data. Further, the morphological damages and increased proliferation with more death cells of pyrrolo[1,2-a]pyrazine-1,4-dione, hexahydro-3 treated MCF-7 were observed by phase contrast and fluorescence microscope techniques. Finally, the nuclear condensation and increased necrotic cells of the pyrrolo[1,2-a]pyrazine-1,4-dione, hexahydro-3 treated MCF-7 cells was clearly identified by fluorescence microscope techniques using Hoechst 33,342 stain. Therefore, our result was prove, the pyrrolo[1,2-a]pyrazine-1,4-dione, hexahydro-3 rich Streptomyces akiyoshiensis GRG 6 (KY457710) as a potential anti-cancer agent against MCF-7 breast cancer cells.

Chiral amphiphiles are useful for controlling the structures and properties of supramolecular assemblies, but their stereocontrolled synthesis is generally difficult, because their long alkyl chains tend to bring unfavorable effects on the solubility, reactivity, and crystallinity of molecules. Typical examples are amphiphilic 1,2‐amino alcohols (S)‐1 and (1S,2S)‐2 developed by our group, which were known to serve as chiral reaction media for controlling the stereochemistry of asymmetric photoreactions. We previously developed synthetic schemes for these 1,2‐amino alcohols, but their synthetic efficiencies were unsatisfactory (13 steps with 2% overall yield for (S)‐1; eight steps with 8% yield for (1S,2S)‐2). As the main reason of such low efficiencies, the stereocontrolling methods we previously employed (diastereomer‐salt crystallization for (S)‐1; stereoselective reactions for (1S,2S)‐2) were not ideal. Here, we report highly improved synthetic schemes for (S)‐1 and (1S,2S)‐2 based on the enantioselective high performance liquid chromatography (HPLC) separation of intermediates in preparative scales. Compared with the previous schemes, the new schemes are advantageous in fewer number of steps, higher overall yield, and lower risk of racemization (seven steps with 15% overall yield for (S)‐1; seven steps with 26% overall yield for (1S,2S)‐2). [ABSTRACT FROM AUTHOR]

The present study is intended to explore the pharmacological significance of the crude root extract of Sterculia urens Roxb. Further the bio-active compounds were isolated and characterized using chromatographic and spectroscopic techniques. Soxhlet extraction apparatus was utilised for isolating the chemical constituents from the root extract using a series of solvents such as n-hexane, ethyl acetate (EA), methanol (ME) and water (WA). The pharmacological activities such as inhibition of DPPH radical activity, a-amylase enzyme activity, albumin denaturation along with anti-bacterial and thrombolytic activities. The isolation of purified bio-active constituents was carried using preparative HPLC technique and the purified compounds were characterized using spectroscopic techniques like NMR and Mass spectroscopy. Among the crude root extracts, methanolic extract shows high DPPH radical scavenging activity with IC50 concentration of 26.74±0.08 µg/mL. The IC50 concentrations in a-amylase enzyme inhibition activity was 263.96±0.90, 127.73±1.23 and 223.54±4.76 µg/mL for EA, ME, WA extracts respectively. The ME extract shows high albumin denaturation inhibition assay than other extracts with IC50 concentration as 137.09±0.20 µg/mL which is very close to standard ascorbic acid. The ME shows high % clot lysis than other extracts and results were comparable with 100 µL of streptokinase standard. The preparative HPLC followed by spectral analysis confirm that two known alkaloids (Sterculinine I & II) and three known flavonoids (Gossypetin, Apigenin and 6 Hydroxyluteolin) were purified and characterised from the root ME of Sterculia urens Roxb. The purified and identified compounds were reported for the first time in Sterculia urens Roxb. [ABSTRACT FROM AUTHOR]

The current study was concentrated to detection of anti-cancer compound from marine Streptomyces akiyoshiensis GRG 6 (KY457710) against MCF-7 breast cancer cells. The partial purification of TLC was moved three different active spots. Among the spots, the third spot of the extract was shown excellent anti-cancer properties against MCF-7 human breast cancer cells. The available anti-cancer compounds present in the partially purified extract was confirmed by GC–MS data interpretation. Three fractions of the compounds were purified by preparative HPLC and their respective spectrum was confirmed by analytical HPLC. Active anti-cancer fraction was separately purified by preparative HPLC and the cytotoxicity result was shown with 100 µg/mL concentration. The anti-cancer compound of pyrrolo [1,2-a]pyrazine-1,4-dione, hexahydro-3 was detected by LC-MS analysis after interpretation with GC–MS data. Further, the morphological damages and increased proliferation with more death cells of pyrrolo[1,2-a]pyrazine-1,4-dione, hexahydro-3 treated MCF-7 were observed by phase contrast and fluorescence microscope techniques. Finally, the nuclear condensation and increased necrotic cells of the pyrrolo[1,2-a]pyrazine-1,4-dione, hexahydro-3 treated MCF-7 cells was clearly identified by fluorescence microscope techniques using Hoechst 33,342 stain. Therefore, our result was prove, the pyrrolo[1,2-a]pyrazine-1,4-dione, hexahydro-3 rich Streptomyces akiyoshiensis GRG 6 (KY457710) as a potential anti-cancer agent against MCF-7 breast cancer cells. [ABSTRACT FROM AUTHOR]

Thymoquinone (TQ) is a monoterpene ketone which is an important bioactive substance in Nigella sativa L. seed. This important natural compound has many potential medical applications for cardiovascular diseases, diabetes, stroke, neurodegenerative diseases and different types of cancer. To date, no studies have focused on development of a reliable industrial method for purification of TQ from this plant as a major challenge. In this study, an efficient extraction and purification method was developed for preparation of TQ from Nigella sativa L. seeds using preparative high-performance liquid chromatography (HPLC). To this end, a two-step procedure was applied for the first time for extraction of TQ: first, maceration using methyl tert-butyl ether (MTBE) was performed; then, liquid-liquid extraction using methanol successfully removed the majority of the impurities. Next, preparative HPLC was carried out for separation and purification of TQ using a C18 column and the mobile phase of methanol and water containing 0.1% trifluoroacetic acid (TFA).The collected peak from preparative HPLC was analyzed by the analytical HPLC as well as GC-MS instrument. Results of HPLC analysis proved the purity of the collected TQ by 97%, while the results of GC-MS identified the main peak appeared at around 15.6 min as TQ by library searching, and determined its purity by 97% based on peak heights. Overall, this method has a potential for industrialization to prepare purified TQ for medicinal applications. [ABSTRACT FROM AUTHOR]

Abstract Background Khat (Catha edulis Forsk) is an evergreen shrub of the Celastraceae family. It is widely cultivated in Yemen and East Africa, where its fresh leaves are habitually chewed for their momentary pleasures and stimulation as amphetamine-like effects. The main psychostimulant constituents of khat are the phenylpropylamino alkaloids: cathinone, cathine and norephedrine. Results In this study, simple procedures based on preparative HPLC and salting-out assisted liquid–liquid extraction (SALLE) based methods were developed respectively for large scale isolation and the extraction of psychoactive phenylpropylamino alkaloids; cathinone, cathine and norephedrine, from khat (Catha edulis Forsk) chewing leaves, a stimulant and drug of abuse plant. The three khat alkaloids were directly isolated from the crude oxalate salt by preparative HPLC–DAD method with purity > 98%. In addition, a modified (SALLE) method has been developed and evaluated for the extraction efficiency of psychoactive phenylpropylamino alkaloids from khat (Catha edulis Forsk) chewing leaves. An in situ two steps extraction protocol was followed without dispersive SPE clean up. The method involves extraction of the samples with 1% HAc and QuEChERS salt (1.0 g of CH3COONa and 6.0 g of MgSO4) followed by subsequent in situ liquid–liquid partitioning by adding ethyl acetate and NaOH solution. The optimized method allowed recoveries of 80–86% for the three alkaloids from khat sample with relative standard deviation (RSD) values less than 15% and limits of detection (0.85–1.9 μg/mL). Conclusion The method was found to be simple, cost-effective and provides cleaner chromatogram with good selectivity and reproducibility. The SALLE based protocol provided as good results as the conventional extraction method (ultrasonic assisted extraction followed by solid phase extraction, UAE–SPE) and hence the method can be applicable in forensic and biomedical sectors.

Purpose: Zygophyllum fabago L. (Z. fabago) is a widespread perennial herb which is used as a medicinal plant in traditional medicine of Iran, Turkey and China. The present study was a survey on phytochemical constituents and biological activities of this plant. Methods: Methanolic extract of the roots was fractionated over a C-18 pre-packed cartridge (Sep-pak) and chromatographic separation was performed on a reversed-phase preparative HPLC. Structural elucidation of the isolated compounds was carried out using UV, 1H-NMR and 13C-NMR spectral analyses. Furthermore, the chemical compositions of the essential oil of the aerial parts were identified by GC-MS analysis. Antiproliferative and antioxidant activities of all extracts from aerials were determined by MTT and DPPH assays, respectively. Results: Phytochemical investigation on the plant roots led to the isolation and identification of two the 60% methanol-water Sep-pak fraction, a prenylated flavone glycoside, 6-C-prenyl-7-O-[ β -D-4'''-O-acetyl-glucopyranosyl-(1'''→2'')-β-D-glucopyranosyl] apigenin, which was named as a Zygocaperoside and also, other flavonoid, was named as the Isorhamnetin -3-O glucoside. None of the extracts showed antiproliferative effect against cancerous cells. However, among the extracts, methanolic extract indicated antioxidant activity. Moreover, essential oils of flowers and leaves of plant have high amounts of sesquiterpene hydrocarbons and diterpenoides. Conclusion: The results of present study introduce Z. fabago roots as a new source of flavonoid glycosides and suggest it as an appropriate candidate for further pharmacological studies.

• Two-step preparative HILIC fractionation of lithium ion battery electrolyte. • Extraction of potentially hazardous organofluorophosphate decomposition products. • Effective removal of electrolyte matrix for toxicological investigations. The expansion of lithium ion battery (LIB) application is accompanied by the growth of battery pack sizes. This progression emphasizes the consideration of electrolyte safety as well as environmental aspects in case of abuse, accident, or recycling. Hexafluorophosphate is one of the most commonly used conducting salt anions in electrolytes. It has great potential to degrade to various acidic and non-acidic organo(fluoro)phosphates with presence of water and during battery cell operation. Consequently, toxicological investigation on these organo(fluoro)phosphates has emerged because they either have structural similarities as chemical warfare agents or play a widespread physiological role as phosphates in the human body. This circumstance underlines the need of isolated examination of these compounds for safety assessment. In this work, we used hydrophilic interaction liquid chromatography for the extraction of acidic organofluorophosphates from thermally aged LIB electrolytes. The developed two-step fractionation method provided high separation selectivity towards acidic head groups, which allowed the separation of undesired matrix and target compounds. These findings facilitate isolated toxicological investigations on organofluorophosphates that are beneficial for environmental and safety research, the battery cell industry, and human safety surveillance in regard to aged LIB electrolytes. [ABSTRACT FROM AUTHOR]

Aim: The present study reports the degradation behavior of new antihypertensive drug Riociguat under various stress conditions as per International Conference on Harmonization guidelines ICH, Q2(R1). Materials and Methods: Riociguat was subjected to stress degradation under hydrolytic (acidic, alkaline and neutral), oxidative, photolytic and thermal stress conditions to investigate the inherent stability. A rapid, accurate, precise and robust HPLC method was developed on Waters Symmetry C18 Column (150mm X 4.6 mm, 5μ) using isocratic elution of 10 mm ammonium acetate buffer pH 5.7 and acetonitrile in the ratio of 70:30 with the flow rate at 1.0 mL/min. The detection was performed at 254nm. Results: The drug was found to be degraded in alkaline and oxidative condition whereas it was stable under acidic, neutral hydrolytic, thermal and photolytic conditions. Two degradation products (DP1, DP2) under alkaline condition and one under oxidative condition (DP3) were characterized by LC-HR-MS/MS with accurate mass measurements. Degradation products (DP1, DP2 and DP3) were isolated by preparative HPLC and were characterized by ¹H NMR, 13C NMR, APT and IR Techniques. Conclusion: Using spectral data analysis, alkaline degradation product DP1 wascharacterized as 2-(1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl)-N5-methylpyrimidine-4,5,6-triamine and DP2 was characterized as 2-(1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl)- 6-amino-7-methyl-7H-purin-8(9H)-one while oxidative degradation product DP3was characterized as methyl 2-(1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl)-4,6- diaminopyrimidin-5-ylmethylcarbamate-N-oxide. The developed chromatographic method was validated in terms of specificity, linearity, accuracy, precision as per ICH guidelines. The robustness of the method was studied with 2-level fractional factorial design 24-1. [ABSTRACT FROM AUTHOR]

Laccifer lacca (Kerr) (LL) has widely been used as pigmenting, dying and coloring agent in chemical industry. Although, it has wide range of industrial applications, but unfortunately, due to lesser availability of data, it has been ignored. Keeping in mind, the broad application of Laccifer lacca (Kerr), we tried to report the in vitro anti-cancer effects along with its chemical characterization. This work is divided into two sections, first section to assess the anti-cancer activity of dye of Laccifer lacca (Kerr) employing SRB assay. In second section, the active fraction was processed to isolate the active chemical entities using preparative gradient HPLC-UV. The detector was set at 275 nm and 360 nm as reference wavelength. Results of ethyl acetate fraction (EA) showed promising anti-cancer spectrum for MDA-MB-231 (<10 µg/mL) and SiHa (<10 µg/mL) cell lines. Furthermore, laccaic acid-A was identified from EA of Laccifer lacca. In conclusion, our results suggested being the first report for isolation of laccaic acid-A from Laccifer lacca. We characterized the isolated compound be UV, IR, NMR and Mass spectrometry. We also propose that this is the first report for isolation of laccaic acid- A using preparative HPLC with a good yield and purity. [ABSTRACT FROM AUTHOR]

An improved process for the synthesis of latanoprost having excellent optical purity (de 99.9%, [α]D20 = +35.37° (c = 0.90, acetonitrile)) without use of preparative HPLC is described. This process involves effective purification of hydroxyl intermediate (5A) through solvent crystallization followed by inhibition of inversion of the chiral center at C-15 position. This was possible due to judicious use of diol intermediate (6) for double bond reduction prior to hydroxyl protection. [ABSTRACT FROM AUTHOR]

Flavonol glycosides are important components of tea leaves, contributing to the bioactivities as well as bitterness and astringency of tea. However, the standards of many flavonol triglycosides are still not available, which restricts both sensory and bioactivity studies on flavonol glycosides. In the present study, we established a simultaneous preparation method of seven flavonol triglycoside individuals from tea leaves, which consisted of two steps: polyamide column enrichment and preparative HPLC isolation. The structures of seven flavonol triglycoside isolates were identified by mass and UV absorption spectra, four of which were further characterized by nuclear magnetic resonance spectra, namely, quercetin-3-O-glucosyl-rhamnosyl-glucoside, quercetin-3-O-rhamnosyl-rhamnosyl-glucoside, kaempferol-3-O-glucosyl-rhamnosyl-glucoside and kaempferol-O-rhamnosyl-rhamnosyl-glucoside. The purities of all isolated flavonol triglycosides were above 95% based on HPLC, and the production yield of total flavonol glycosides from dry tea was 0.487%. Our study provides a preparation method of flavonol triglycosides from tea leaves, with relatively low cost of time and solvent but high production yield.

This study aimed to isolate, prepare and identify the main flavonoids from a standardized Smilax glabra flavonoids extract (SGF) using preparative HPLC, MS, 1H NMR and 13C NMR, determine the contents of these flavonoids using UPLC, then compare their pharmacological activities in vitro. We obtained six flavonoids from SGF: astilbin (18.10%), neoastilbin (11.04%), isoastilbin (5.03%), neoisoastilbin (4.09%), engeletin (2.58%) and (−)-epicatechin (1.77%). The antioxidant activity of six flavonoids were evaluated by determining the 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical and 2,2′-Azinobis (3-ethylbenzothiazoline-6-sulphonic acid) diammonium salt (ABTS+) radical scavenging activity and ferric reducing antioxidant power (FRAP). In addition, the anti-inflammatory activity of six flavonoids were evaluated by determining the production of cytokines (IL-1β, IL-6), nitric oxide (NO) using enzyme linked immunosorbent assay and the NF-κB p65 expression using Western blotting in lipopolysaccharide (LPS)-stimulated RAW264.7 cells. The results showed that (−)-epicatechin, astilbin, neoastilbin, isoastilbin and neoisoastilbin had strong antioxidant activities, not only in DPPH and ABTS+ radicals scavenging capacities, but in FRAP system. Furthermore, all the six flavonoids could significantly inhibit the secretion of IL-1β, IL-6, NO (p < 0.01) and the protein expression of NF-κB p-p65 (p < 0.01) in LPS-stimulated RAW264.7 cells. This study preliminarily verified the antioxidant and anti-inflammatory activities of six flavonoids in S. glabra.

Plant extracts are complex matrices and, although crude extracts are widely in use, purified compounds are pivotal in drug discovery. This study describes the application of automated preparative-HPLC combined with a rapid off-line bacterial bioassay, using reduction of a tetrazolium salt as an indicator of bacterial metabolism. This approach enabled the identification of fractions from Dodonaea viscosa that were active against Staphylococcus aureus and Escherichia coli, which, ultimately, resulted in the identification of a clerodane type diterpenoid, 6β-hydroxy-15,16-epoxy-5β, 8β, 9β, 10α-cleroda-3, 13(16), 14-trien-18-oic acid, showing bacteriostatic activity (minimum inhibitory concentration (MIC) = 64–128 µg/mL) against test bacteria. To the best of our knowledge, this is the first report on antibacterial activity of this metabolite from D. viscosa.

This study reports a comprehensive investigation on the presence of flavonoids, limonoids and dietary fiber determined by HPLC in all the by-products of the industrial transformation of orange. Seeds were the richest source of bioactive molecules, with flavanones being the most abundant, followed by phenolic acids (238 mg/kg). However, p-hydroxybenzoic acid and caffeic acid were highly represented also in the exhausted peels and pulps (560 mg/kg). Limonoids were determined exclusively in seeds. Among all the by-products it was found that waste water is extremely rich of hesperidin (19,500 mg/kg). For this reason an only eco-friendly preparative HPLC method for the recovery and purification of hesperidin from waste water was developed. Dietary fiber was determined in exhausted peels and pulps which resulted to be rich sources of insoluble dietary fiber. The results show that the by-products here investigated represent important sources of nutraceuticals as ingredients useful for functional food preparations.

Abstract Genetic analysis of Paleoamerican human remains suggests that people first entered the Americas sometime between ∼14,000 and ∼16,000 years ago. Evaluation of these data requires unequivocal archaeological evidence in a solid geological context that is well dated. Accurately determining the age of late Pleistocene sites is thus crucial in explaining when and how humans colonized the Americas. There are, however, significant challenges to dating reliability, especially when vertebrate fossils (i.e. bones, teeth and ivory) are often the only datable materials preserved at sites. We re-dated vertebrate fossils associated with the North American butchering sites of Wally's Beach (Canada), La Prele [also known as Fetterman (Wyoming)], Lindsay (Montana), and Dent (Colorado). Our work illustrates the crucial importance of sample chemical preparation in completely removing contaminants derived from sediments or museum curation. Specifically, our work demonstrates that chromatographic methods, e.g. preparative High Performance Liquid Chromatography and column chromatography using XAD resins, are currently the only efficient methods for removing environmental and museum-derived contaminants. These advanced techniques yield demonstrably more accurate AMS 14C measurements that refine the ages of these four sites and thereby contribute to advancing our understanding of human dispersals across North America during the late Pleistocene. Highlights • The arrival time for humans into North America is still an extremely debated topic. • Butchering sites can reveal the presence of humans even if stone artifacts are absent. • Radiocarbon dates can be inaccurate because of incomplete removal of contaminants. • Chromatographic methods are the most efficient to remove contaminants from bones. • These new dates help to build stronger chronologies for the peopling of the Americas. [ABSTRACT FROM AUTHOR]

In order to enrich and separate three coumarins (columbianetin acetate, osthole and columbianadin) from Angelicae Pubescentis Radix (APR), an efficient method was established by combining macroporous resins (MARs) with preparative high-performance liquid chromatography (PHPLC). Five different macroporous resins (D101, AB-8, DA-201, HP-20 and GDX-201) were used to assess the adsorption and desorption characteristics of three coumarins. The result demonstrated that HP-20 resin possessed the best adsorption and desorption capacities for these three coumarins. Moreover, the adsorption dynamics profiles of three coumarins were well fitted to the pseudo second order equation (R2 > 0.99) for the HP-20 resin. The adsorption process was described by the three isotherms models including Langmuir (R2 > 0.98, 0.046 ≤ RL ≤ 0.103), Freundlich (R2 > 0.99, 0.2748 ≤ 1/n ≤ 0.3103) and Dubinin Radushkevich (R2 > 0.97). The contents of columbianetin acetate, osthole and columbianadin in the product were increased 10.69-fold, 19.98-fold and 19.68-fold after enrichment, respectively. Three coumarins were further purified by PHPLC and the purities of them reached above 98%. Additionally, the anti-inflammatory effects of these three coumarins were assessed by Lipopolysaccharide (LPS)-induced RAW 264.7 cells. It was found that the production of NO and MCP-1 was obviously inhibited by three coumarins. Columbianetin acetate, osthole and columbianadin could be used as potentially natural anti-inflammatory ingredients in pharmaceutical products. It was concluded that the new method combining MARs with PHPLC was efficient and economical for enlarging scale separation and enrichment of columbianetin acetate, osthole and columbianadin with anti-inflammatory effect from the APR extract.

St. John's Wort (Hypericum perforatum L.) is one of the most studied plant species in the family Hypericaceae. The aim of this study was the identification of the constituents of methanol extract of H. perforatum and optimization of conditions for their isolation. The main components of the methanol extract were isolated on preparative ZORBAX Eclipse XDB C18 column with solvent system consisting of methanol and 1x10-2 M ammonium acetate in water. Constituents of the extract were identified by comparing their retention times with the retention times of the standards, with the literature data and the UV spectra. By varying the conditions of chromatography, the optimal conditions for isolation of the methanol extract constituents were determined: mobile phase consisting of methanol and 1x10-2 M ammonium acetate in water in ratio 1 : 1, sample concentration 100 mg/mL, sample volume 30 µL, flow 2 mL/min. Under these conditions 7 components of the methanol extract were isolated.

Background: Khat ( Catha edulis Forsk) is an evergreen shrub of the Celastraceae family. It is widely cultivated in Yemen and East Africa, where its fresh leaves are habitually chewed for their momentary pleasures and stimulation as amphetamine-like effects. The main psychostimulant constituents of khat are the phenylpropylamino alkaloids: cathinone, cathine and norephedrine. Results: In this study, simple procedures based on preparative HPLC and salting-out assisted liquid-liquid extraction (SALLE) based methods were developed respectively for large scale isolation and the extraction of psychoactive phenylpropylamino alkaloids; cathinone, cathine and norephedrine, from khat ( Catha edulis Forsk) chewing leaves, a stimulant and drug of abuse plant. The three khat alkaloids were directly isolated from the crude oxalate salt by preparative HPLC-DAD method with purity > 98%. In addition, a modified (SALLE) method has been developed and evaluated for the extraction efficiency of psychoactive phenylpropylamino alkaloids from khat ( Catha edulis Forsk) chewing leaves. An in situ two steps extraction protocol was followed without dispersive SPE clean up. The method involves extraction of the samples with 1% HAc and QuEChERS salt (1.0 g of CHCOONa and 6.0 g of MgSO) followed by subsequent in situ liquid-liquid partitioning by adding ethyl acetate and NaOH solution. The optimized method allowed recoveries of 80-86% for the three alkaloids from khat sample with relative standard deviation (RSD) values less than 15% and limits of detection (0.85-1.9 μg/mL). Conclusion: The method was found to be simple, cost-effective and provides cleaner chromatogram with good selectivity and reproducibility. The SALLE based protocol provided as good results as the conventional extraction method (ultrasonic assisted extraction followed by solid phase extraction, UAE-SPE) and hence the method can be applicable in forensic and biomedical sectors. [ABSTRACT FROM AUTHOR]

A purification sequence including a Gilson CPC 250 PRO device coupled to PrepHPLC hyphenated with a MS triggering fraction collector was applied to isolate secoiridoid glycosides from a complex methanolic extract of Centaurium erythraea . This species is widely used for ethnomedicinal purposes around the Mediterranean Sea. The solvent system ethyle acetate/ethanol/water 7.5/3/5 was determined using shake-flask method targeting swertiamarin, the major secoiridoid of the extract. Optimization of CPC experimental parameters enabled the injection of 4 g of extract with a flow rate of 40 mL/min at 3000 rpm to provide a secoiridoid glycosides enriched fraction. 130 mg of this latter was submitted to a second step of purification by preparative HPLC (gradient water/formic acid (19:1) (A) and methanol (B) as follows: 0 min, 85% A; 8 min, 60% A; 12 min, 55% A; 35 min, 55% A; 40 min, 10% A; 50 min, 10% A; 52 min, 85% A; 55 min, 85% A) to give swertiamarin (36 mg, yield 27.7%, purity 98.2%). Other secoiridoid glycosides (sweroside, gentiopicroside, secologanol, secoxyloganin) were also isolated in minor amounts. As these monoterpene derivatives are responsible for several biological activities, their quick recovery with high yield and purity may serve as a model for further scale-up and industrial development. [ABSTRACT FROM AUTHOR]

Some bacteria produce antimicrobial chemicals in their immediate environments. These antimicrobial agents are enzymes, polypeptides or non-protein organic compounds. In this study, a bacterial isolate that produces antifungal chemical(s) was isolated from an over incubated nutrient agar plate that was exposed to air. The bacterium is aerobic, Gram positive bacilli; capsule and endospore producing. It ferments glucose and sucrose but not lactose, galactose, mannitol and sorbitol; it is citrate, indole, methyl red and Voges Prauskauer negative. Using agar gel diffusion technique, the cell-free culture supernatant resulting from centrifugation of a six day bacterial culture showed antifungal activity against filamentous fungi but not yeasts. Heating the cell-free supernatant in 90°C water bath and digestion with different proteases had no negative impact on the antifungal activity. A segment of 16S rRNA gene of the bacterial isolate was amplified. The nucleotide sequence of the amplicon was used to identify the bacterium as being very similar to Bacillus amyloliquefaciens subsp. plantarum strain FZB42. Five bacterial peptides were isolated and identified from cell free supernatant of the bacterial culture using a suite of techniques, including flash chromatography, HPLC, NMR and mass spectrometry. One of the five peptides has been previously reported in literature to possess antifungal activity. [ABSTRACT FROM AUTHOR]

A fast and efficient preparative HPLC-PDA method was developed for the separation and isolation of four rare isomeric kaempferol diglycosides from leaves of Prunus spinosa L. The separation procedure of the enriched diglycoside fraction of the 70% ( v/v ) aqueous methanolic leaf extract was first optimised on analytical XBridge C18 column (100 mm × 4.6 mm i.d., 5 μm) and central composite design combined with response surface methodology was utilized to establish the optimal separation conditions. The developed method was directly transferred to preparative XBridge Prep C18 column (100 mm × 19 mm i.d., 5 μm) and the final separation was accomplished by isocratic elution with 0.5% acetic acid-methanol-tetrahydrofuran (75.2:16.6:8.2, v/v/v ) as the mobile phase, at a flow rate of 13.6 mL/min, in less than 12 min for a single run. Under these conditions, four flavonoid diglycosides: kaempferol 3- O -α- l -arabinofuranoside-7- O -α- l -rhamnopyranoside, kaempferol 3,7-di- O -α- l -rhamnopyranoside (kaempferitrin), and reported for the first time for P. spinosa kaempferol 3- O - β - d -xylopyranoside-7- O -α- l -rhamnopyranoside (lepidoside) and kaempferol 3- O -α- l -arabinopyranoside-7- O -α- l -rhamnopyranoside, were isolated in high separation yield (84.8–94.5%) and purity (92.45–99.79%). Their structures were confirmed by extensive 1D and 2D NMR studies. Additionally, the UHPLC-PDA-ESI–MS 3 qualitative profiling led to the identification of twenty-one phenolic compounds and confirmed that the isolates were the major components of the leaf material. [ABSTRACT FROM AUTHOR]

Purpose: Zygophyllum fabago L. (Z. fabago) is a widespread perennial herb which is used as a medicinal plant in traditional medicine of Iran, Turkey and China. The present study was a survey on phytochemical constituents and biological activities of this plant. Methods: Methanolic extract of the roots was fractionated over a C-18 pre-packed cartridge (Sep-pak) and chromatographic separation was performed on a reversed-phase preparative HPLC. Structural elucidation of the isolated compounds was carried out using UV, 1H-NMR and 13C-NMR spectral analyses. Furthermore, the chemical compositions of the essential oil of the aerial parts were identified by GC-MS analysis. Antiproliferative and antioxidant activities of all extracts from aerials were determined by MTT and DPPH assays, respectively. Results: Phytochemical investigation on the plant roots led to the isolation and identification of two the 60% methanol-water Sep-pak fraction, a prenylated flavone glycoside, 6-C-prenyl-7-O-[β-D-4‴-O-acetyl-glucopyranosyl-(1‴→2″)-β-D-glucopyranosyl] apigenin, which was named as a Zygocaperoside and also, other flavonoid, was named as the Isorhamnetin -3-O glucoside. None of the extracts showed antiproliferative effect against cancerous cells. However, among the extracts, methanolic extract indicated antioxidant activity. Moreover, essential oils of flowers and leaves of plant have high amounts of sesquiterpene hydrocarbons and diterpenoides. Conclusion: The results of present study introduce Z. fabago roots as a new source of flavonoid glycosides and suggest it as an appropriate candidate for further pharmacological studies. [ABSTRACT FROM AUTHOR]

This paper discusses the development of a separation method for isolation of Tm from a half-gram irradiated erbium target in support of stockpile stewardship and astrophysics research. The developed procedure is based on cation exchange separation using alpha-hydroxyisobutyric acid (α-HIBA) as chelating agent. It is able to achieve either a decontamination factor of 1.4(4) × 10 with 68.9(3) % recovery or 95.4(3) % recovery with a decontamination factor of 5.82(7) × 10 for a mock 500-mg target containing 17.9 mg thulium in a single pass-through at room temperature. [ABSTRACT FROM AUTHOR]

Phorbol esters (PEs) are well known as the main toxic compounds in Jatropha curcas Linnaeus (JCL), the seed oil of which has been considered as a major feedstock for the production of biodiesel. In the present study, we investigated a series of PEs extracted from JCL seed kernels with methanol (MeOH), and identified more than seven components contained in the PEs. The isolation of main five components of a series of PEs was revised using a semi-preparative reversed phase HPLC analysis of ODS-3 column. The five peaks of components were successfully isolated, and peaks of J2, J3, J5, and J7 were assigned to be Jatropha factors C 1 , C 2 , C 3, and C 4/5 , but J6 was a mixture of Jatropha factor C 6 and its isomer based on the data of UV and LC–MS/MS, and J2 was identified using 1 H NMR analysis. By characterization using LC–MS/MS analysis, all components of a series of PEs were elucidated to be the 12-deoxy-16-hydroxyphorbol esters composed of isomeric form of dicarboxylic groups with same m / z value of 380. [ABSTRACT FROM AUTHOR]

Eight bufadienolides were successfully isolated and purified from ChanSu by high-speed counter-current chromatography (HSCCC) combined with preparative HPLC (prep-HPLC). First, a stepwise elution mode of HSCCC with the solvent system composed of petroleum ether - ethyl acetate - methanol - water (4:6:4:6, 4:6:5:5, v/v) was employed and four bufadienolides, two partially purified fractions were obtained from 200 mg of crude extract. The partially purified fractions III and VI were then further separated by prep-HPLC, respectively, and another four bufadienolides were recovered. Their structures were confirmed by ESI-MS and ¹H-NMR spectra.