High-performance Chromatography Technique Research Articles

Carotenoids from Different Pumpkin Varieties Exert a Cytotoxic Effect on Human Neuroblastoma SH-SY5Y Cells.

Plants, including pumpkins (Cucurbita spp.), are an interesting source of nutrients and bioactives with various health benefits. In this research, carotenoid extracts obtained from the pulp of eight pumpkin varieties, belonging to the C. moschata and C. maxima species, were tested for cytotoxicity on SH-SY5Y neuroblastoma cells. The results showed that pumpkin bioactives exert a cytotoxic action against the tested cells, in particular Butternut extract at a 100 μM (53.69 μg/mL) concentration after 24 h of treatment and Mantovana extract at 50 μM (26.84 μg/mL) after 48 h. Moreover, the carotenoid extracts also showed interesting in vitro antioxidant activity, evaluated by ABTS and ORAC assays. To fully characterize the qualitative and quantitative profile of carotenoids in the tested extracts, a high-performance chromatographic technique was performed, revealing that pumpkin pulp carotenoids were mainly represented by β-carotene, mono- and di-esterified hydroxy- and epoxy-carotenoids. Moreover, the carotenoid dataset was also useful for discriminating samples from two different species. In conclusion, the results of the present study highlight the potential anti-cancer activity of pumpkin carotenoid extracts and the possibility of using them as chemotherapeutic adjuvants.

Detection of some Volatile Oils using High-Performance Chromatography Technique and Study Antibacterial Activity of Black Seed Extract Nigella sativa L.

The antibacterial activity of crude extract of Nigella sative extract was studied on the some pathogenic bacteria by using paper disc diffusion. The tested strains were: Escherichia coli, Enterococcus faecalis, staphylococcus aureus (MRSA) and psendomonas aeruginosa. The results were showed that the crude extract produced inhibition zone in all tested bacteria ranging from (20 mm) in S.aureus and MRSA and (12 mm) in P.aureginusa. Some Volatile Oils were determined using a high-performance liquid chromatograph(hplc), and three extraction methods (aqueous, methanol and ethanol) were used. The following Volatile Oils were estimated ((Limonene, Anethole, -α-pinene, Estagole) reached(35.61,86.40,41.69,35.61) sequentially While the aqueous extraction recorded the lowest averages of (30.52, 71.42, 32.82, 25.89). In conclusion, the current article was attitude to assess the effectiveness of N. sativa in front of all prevalent illnesses caused by strains of microorganisms that are drug-resistant.

Detection and Analysis of Xenobiotics in Food Products and Environmental and Biological Samples by High-Performance Chromatographic Techniques Coupled with High-Resolution Mass Spectrometry.

Detection and Analysis of Xenobiotics in Food Products and Environmental and Biological Samples by High-Performance Chromatographic Techniques Coupled with High-Resolution Mass Spectrometry.

A syn-ent-labdadiene derivative with a rare spiro-β-lactone function from the male cones of Wollemia nobilis

An undescribed labdane-like diterpene with a rare spiro-β-lactone function was identified from the ethanol extract of the male cones of the coniferous tree Wollemia nobilis. This spirolabdadienolide (IUPAC name: syn-ent-8(17),13-labdadien-19,18-olid-15-oic acid methyl ester; trivial name: wollemolide), was isolated by means of traditional and high performance chromatography techniques and structurally elucidated through NMR and MS. In addition, six further known metabolites were evidenced in the extract. Wollemolide, which may be considered an additional chemotaxonomic marker, and 4′-O-methyl-scutellarein, a simple flavonoid, had not been isolated in our previous phytochemical study on the same plant organ. This demonstrates how the molecular pattern of a plant species is in continuous movement and changes with the passing of time according to the climate of the year.

Prevalence of toxigenic fungi in common medicinal herbs and spices in India.

Mycotoxins are unavoidable contaminants of food grains, feeds, medicinal herbs, and spices, posing as health threat to animals and humans. The objective of this study was to screen medicinal herbs and spices for fungi and mycotoxin contamination and evaluate their safety. Sixty-three samples were examined for fungal contamination and fungal load determined using standard microbiological method. Aflatoxin and citrinin were detected using thin layer chromatography and high-performance chromatography technique. Fifty-eight out of the 63 samples were contaminated, while five were free from fungal contamination. Analysis revealed that 47 % of the samples had a fungal load above 1 × 103 cfu/g which is the permissible limit set by World Health Organization. The samples Mesua ferrea-II and Terminalia chebula-III had the highest fungal load, i.e., 5.0 × 104 cfu/g. A total of 187 fungi were isolated, out of which 28 were toxigenic which included 19 aflatoxin-producing Aspergillus flavus and 9 citrinin-producing Penicillium citrinum. The natural contamination with aflatoxin B1 was detected only in one sample, i.e., Arachis hypogaea (groundnut) which was present beyond the permissible limit. Though toxigenic fungi were isolated, mycotoxins were not detected from any of the medicinal herbs and spices. Medicinal herbs and spices are susceptible to toxigenic fungi; however, they also possess intrinsic factors that inhibit mycotoxin contamination. This study provides a basis in assessing the degree of fungal and potential mycotoxin contamination in medicinal herbs and spices.Electronic supplementary materialThe online version of this article (doi:10.1007/s13205-016-0476-9) contains supplementary material, which is available to authorized users.

Application of HPCCC, UHPLC-PDA-ESI-MS3 and HPLC-PDA methods for rapid, one-step preparative separation and quantification of rutin in Forsythia flowers

In this study, the flowers of Forsythia × intermedia, Forsythia suspensa and Forsythia ovata are presented as valued raw materials for isolation of rutin, an industrial and pharmacologically active flavonoid. An efficient strategy based on three high-performance chromatographic techniques was established to screen and purify the target compound from the plant matrices. In the first step, an UHPLC-PDA-ESI-MS3 assay documented the relatively simple phytochemical profiles of the flower extracts and led to the identification of ten polyphenols classified as caffeoyl acid derivatives, flavonoids, phenylethanoids and lignans, with rutin as the major constituent. Significant levels of rutin (20.0–57.6mg/g dw) were found in the flowers by the validated HPLC-PDA method, with the highest values observed for F. suspensa. Finally, a novel, fast, single-step HPCCC method was developed and successfully applied for the separation of rutin from the plant samples. The ethyl acetate-n-butanol-water (4:1:5, v/v/v) was selected as the optimum two-phase solvent system for both analytical and preparative normal (NP) and reversed-phase (RP) HPCCC separations. Under optimal RP-HPCCC conditions, 155 mg of the crude methanol extract was separated in a single 50min run, yielding 14.6mg of rutin with 97.2% HPLC purity and 95.9% recovery. The process efficiency of the HPCCC rutin isolation was demonstrated to be superior to the existing HSCCC methods. The results indicated the usefulness of Forsythia flowers and the developed RP-HPCCC methodology for practical and commercial applications.

Nitric oxide inhibition of free radical-mediated lipid peroxidation in photodynamically treated membranes and cells.

Of the numerous biological activities attributed to nitric oxide ((*)NO), relatively little is known about its ability to intercept lipid-derived free radicals and thus protect cells against the damaging effects of lipid peroxidation, particularly in photodynamic settings. To address this, we asked how the (*)NO donor spermine-NONOate (SPER/NO) would affect porphyrin (PpIX)-photosensitized, iron/ascorbate-amplified chain peroxidation in cholesterol (Ch)/phospholipid (0.8:1.0, mol/mol) liposomes. Several Ch oxidation products (ChOX) were monitored by high performance chromatographic techniques. When added immediately before irradiation, SPER/NO (0.4 mM) had no effect on accumulation of 5alpha-hydroperoxide, a primary singlet oxygen-derived ChOX, but strongly suppressed the secondary species arising from postphotooxidation chain reactions, including 7alpha/7beta-hydroperoxides, 7alpha/7beta-hydroxides, and 5,6-epoxides. Metabolism of exogenous 5-aminolevulinate to PpIX in COH-BR1 tumor cells sensitized them to ChOX photogeneration and necrotic photokilling. When present during irradiation, active (but not decomposed) SPER/NO strongly inhibited both effects. These findings support the hypothesis that suitably presented NO, by intercepting lipid-derived radicals, can antagonize the antitumor effects of photodynamic therapy and other oxidative therapies.

19] Analysis of synthetic peptides by high-performance liquid chromatography

High-performance liquid chromatography (HPLC) has proved extremely versatile for the isolation/purification of peptides varying widely in their sources, quantity, and complexity. High-performance chromatographic techniques are suited to the purification of a single peptide from the kind of complex peptide mixtures encountered following solid-phase peptide synthesis, where impurities are closely related to the peptide of interest (deletion, terminated, or chemically modified peptides), perhaps missing only one amino acid residue, and, hence, may be difficult to separate. This chapter focuses on HPLC applications of particular interest to researchers utilizing solid-phase synthesis methods: (1) problem-solving approaches to difficult peptide separations, (2) HPLC monitoring of formation of desired product, (3) and HPLC as a diagnostic tool to detect unwanted side-chain modification. The use of reversed-phase chromatography (RP-HPLC), the most important mode of HPLC for synthetic peptidepurification, is focussed; in addition, the exciting potential of a novel mixed-mode HPLC technique, hydrophilic interaction/cation-exchange chromatography (HILIC/CEC), is demonstrated.

A bioassay-directed investigation of Sydney Harbour sediment

A coal tar-contaminated sediment sample from Sydney Harbour, Nova Scotia, Canada was examined for genotoxicity using a bioassay-directed fractionation approach. An organic solvent soluble fraction, prepared from the marine sediment by ultrasonic extraction, was fractionated into compound classes using a sequential alumina and Sephadex LH20 clean-up procedure followed by high performance chromatographic techniques. The PAH-containing fractions were analyzed using gas chromatography-mass spectrometry (GC-MS) and reversed phase-high performance liquid chromatography (RP-HPLC). The composition of the fractionated material was also characterized using Salmonella typhimurium microbiological assays to identify fractions with mutagenic potency. The fraction containing the PAH was the major source of mutagenicity requiring microsomal activation. Compounds responsible for some of the mutagenic activity in the PAH-containing fraction, including benzo[a]pyrene, benzo[ghi]perylene and indeno[1,2,3-cd]pyrene, were identified through a combination of RP-HPLC followed by GC-MS of individual subfractions.

Interrelation between the charge isoforms of mammalian ornithine decarboxylase

Ornithine decarboxylase (ODC) isolated from a variety of tissues has been separated, using DEAE ion-exchange chromatography, into multiple peaks of activity that appear to be related to control of this enzyme stability. Reports of these charge isoforms in current literature are generally unclear as to whether these represent a covalent post-translational modification or merely an alteration in structural conformation or association. In this study we investigated the relationship of this form separation to the degree of enzyme polymerization, interaction with other proteins and buffer components, and the multiple isoelectric forms of this enzyme noted in denaturing concentrations of urea. High-performance chromatography techniques were used to demonstrate that two of the major enzyme forms, ODC I and II, are really monomers of the enzyme, while minor peaks of activity frequently observed to elute after ODC II contain various dimeric enzyme states. Pyridoxal 5′-phosphate (0.05 m m) added to isolated enzyme preparations composed of I and II monomers induced the formation of I and II dimers as well as a mixed I–II dimer. All three dimer forms were observed to be natural components of freshly isolated crude cell homogenates. The charge distinction between the monomer forms I and II was found to be maintained during ion-exchange chromatography in the presence of 8 m urea, and the enzyme isoforms demonstrated distinct bands on isoelectric focusing gels run in the presence of 9 m urea. Thus, although some of the multiple ornithine decarboxylase forms identified by ion-exchange chromatography of crude mammalian cell homogenates are related to enzyme conformation, the two major forms are distinctly charged protein states that can be visualized using two-dimensional gel electrophoresis of highly purified samples.

Direct in vitro effect of thyroid hormones on 25-hydroxyvitamin D3 metabolism in the perfused rat kidney.

We have shown that thyroid hormones have a direct effect on the renal metabolism of 25-hydroxyvitamin D3 (25OHD3). Using a new high performance chromatographic technique based upon a cyano-bonded packing, we studied the metabolism of [26,27-3H]25OHD3 in the isolated perfused rat kidney in vitro. We measured the rate of synthesis of 24,25-dihydroxyvitamin D3 [24,25-(OH)2D3], 24-oxo-25-hydroxyvitamin D3 [24-oxo-25OHD3], and 24-oxo-23,25-dihydroxyvitamin D3 [24-oxo-23,25-(OH)2D3] by the kidneys of vitamin D-replete [D(+)] rats and the rate of synthesis of 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] by the kidneys of vitamin D-deplete [D(-)] rats. Synthesis of 24,25-(OH)2D3 increased significantly at 2 and 6 h of perfusion in perfused kidneys treated with T3, T4, and TSH as compared to untreated controls. Synthesis of 24-oxo-25OHD3 increased at 2 and 6 h and that of 24-oxo-23,25-(OH)2D3 increased at 6 h in T3- and TSH-treated groups. Perfused kidneys from vitamin D(-) rats treated with T3, T4, and TSH synthesized significantly decreased amounts of 1,25-(OH)2D3 at 2, 4, and 6 h. TRH treatment of kidneys from either vitamin D(+) or D(-) rats caused no significant changes in synthesis of any vitamin D metabolites when compared with appropriate controls. These results suggest T3, T4, and perhaps TSH, but not TRH, have a direct effect on renal 25OHD3 metabolism.

Electrochemical Detection in Hight-Performance Liquid Chromatography

I. INTRODUCTION Modern analytical chemistry must satisfy very diverse demands. Among the most important and simultaneously most difficult tasks are the continuous monitoring of substances (chiefly in industrial and environmental control) and analyses of very complicated organic and inorganic systems, often at trace concentration levels (mainly in clinical analysis, pharmaceutical research and industry, petrochemistry, etc.). For these purposes, suitable detectors must be available. Industrial and laboratory continuous analyzers have a long history. Combination of continuous monitoring with high-performance chromatographic techniques is a much more recent technique.

Separation and quantification of angiotensins and some related peptides by high-performance liquid chromatography.

A high-performance chromatographic technique for the separation of angiotensins and some related peptides is described. Complete separation of angiotensin I, angiotensin II, tetradecapeptide and the tetrapeptide Leu-Val-Tyr-Ser is achieved in a single step, using reversed-phase high-performance liquid chromatography. The application of this technique for the detection of renin activity in crude biological samples, employing the artificial renin substrate tetradecapeptide, is demonstrated.