RESUMO
Magnetic solid phase extraction with the functionalization of protein onto micro- or nano-particles as a probe is favorable for the discovery of new drugs from complicated natural products. Herein, we aimed to develop a rapid method by immobilizing halogenated alkane dehalogenase (Halo)-tagged calcium-sensing receptor (CaSR) directly out of crude cell lysates onto the surface of magnetic microspheres (MM) with no need to purify protein. Thereby we achieved CaSR-functionalized MM for revealing adsorption characteristics of agonist neomycin and screening ligands from herbal medicine Radix Astragali (RA). About 43.87 mg CaSR could be immobilized per 1 g MM within 30 min, and the acquired CaSR-functionalized MM showed good stability and activity for 4 weeks. The maximum adsorption capacity of neomycin on CaSR-functionalized MM was determined as 4.70 × 10-4 ~ 3.96 × 10-4 mol/g within 277 ~ 310 K, and its adsorption isotherm characteristics described best by the Temkin model were further validated using isothermal titration calorimetry. It was inferred that CaSR's affinity for neomycin was driven by electrostatic forces in a spontaneous process when the system reached an equilibrium state. Moreover, the ligands from the RA extract were screened, three of which were assigned as astragaloside IV, ononin, and calycosin based on HPLC-MS. Our findings demonstrated that the functionalization of a receptor onto magnetic materials designed as an affinity probe has the capability to recognize its agonist and capture the ligands selectively from complex matrices like herbs.
Assuntos
Neomicina , Receptores de Detecção de Cálcio , Microesferas , Adsorção , Ligantes , Fenômenos MagnéticosRESUMO
Our initial studies detected elevated levels of 3,4-dihydroxyphenyllactic acid (DHPLA) in urine samples of patients with severe heart disease when compared with healthy subjects. Given the reported anti-inflammatory properties of DHPLA and related dihydroxylated phenolic acids (DPAs), we embarked on an exploratory multi-centre investigation in patients with no urinary tract infections to establish the possible pathophysiological significance and therapeutic implications of these findings. Chinese and Caucasian patients being treated for severe heart disease or those conditions associated with inflammation (WBC ≥ 10 ×109/L or hsCRP ≥ 3.0 mg/L) and/or hypoxia (PaO2 ≤ 75 mmHg) were enrolled; their urine samples were analyzed by HPLC, HPLC-MS, GC-MS and biotransformation assays. DHPLA was detected in urine samples of patients, but undetectable in healthy volunteers. Dynamic monitoring of inpatients undergoing treatment showed their DHPLA levels declined in proportion to their clinical improvement. In DHPLA-positive patients' fecal samples, Proteus vulgaris and P. mirabilis were more abundant than healthy volunteers. In culture, these gut bacteria were capable of reversible interconversion between DOPA and DHPLA. Furthermore, porcine and rodent organs were able to metabolize DOPA to DHPLA and related phenolic acids. The elevated levels of DHPLA in these patients suggest bioactive DPAs are generated de novo as part of a human's defense mechanism against disease. Because DHPLA isolated from Radix Salvia miltiorrhizae has a multitude of pharmacological activities, these data underpin the scientific basis of this medicinal plant's ethnopharmacological applications as well as highlighting the therapeutic potential of endogenous, natural or synthetic DPAs and their derivatives in humans.
Assuntos
Cardiopatias , Inflamação , Humanos , Suínos , Animais , Hipóxia , Di-HidroxifenilalaninaRESUMO
Siraitia grosvenorii (Swingle) C. Jeffrey (SG) is widely used as a natural sweetener and traditional medicine for respiratory diseases. The anti-respiratory compounds in the plant and their mechanism remain elusive due to the lack of a high-throughput screening method. In this work, immobilization of the muscarinic-3 acetylcholine receptor (M3R) was used to establish an affinity chromatographic strategy for synchronously recognizing the flavour components in the SG extract binding to this receptor and evaluating their anti-asthmatic effect. The accuracy of the method was assessed by in vivo experiments. Mogroside V (Mog V) and 11-oxomogroside V (11-O MogV) were identified as functional flavour compounds binding to M3R. Their association constants were determined to be 3.32 × 104 and 2.40 × 104 M-1 by the injection amount-dependent method. The binding energies of the two compounds to M3R were calculated to be -80.52 and -48.20 kJ/mol by molecular dynamics simulation. The synergistic application of the two flavour compounds exhibited stronger anti-asthma activity than the original SG extract. These results indicated that immobilized M3R is a powerful alternative for the identification of flavour compounds in plants. Mog V and 11-O Mog V are the main functional flavour compounds contributing to SG's anti-asthma function. We reasoned that the two compounds have the potential to become functional food additives. This work has the possibility to contribute considerably to the pursuit of functional flavour compounds from natural plants in the field of functional food development.
Assuntos
Antiasmáticos , Cucurbitaceae , Triterpenos , Colinérgicos , Cucurbitaceae/química , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Receptores Colinérgicos , Triterpenos/análiseRESUMO
α-Asaronol from Acorus tatarinowii (known as "Shichangpu" in Traditional Chinese medicine) has been proved to possess more efficient antiepileptic activity and lower toxicity than α-asarone (namely "Xixinnaojiaonang" as an antiepileptic drug in China) in our previous study. However, the molecular mechanism of α-asaronol against epilepsy needs to be known if to become a novel antiepileptic medicine. Nuclear magnetic resonance (NMR)-based metabolomics was applied to investigate the metabolic patterns of plasma and the brain tissue extract from pentylenetetrazole (PTZ)-induced seizure rats when treated with α-asaronol or α-asarone. The results showed that α-asaronol can regulate the metabolomic level of epileptic rats to normal to some extent, and four metabolic pathways were associated with the antiepileptic effect of α-asaronol, including alanine, aspartate, and glutamate metabolism; synthesis and degradation of ketone bodies; glutamine and glutamate metabolism; and glycine, serine, and threonine metabolism. It was concluded that α-asaronol plays a vital role in enhancing energy metabolism, regulating the balance of excitatory and inhibitory neurotransmitters, and inhibiting cell membrane damage to prevent the occurrence of epilepsy. These findings are of great significance in developing α-asaronol into a promising antiepileptic drug derived from Traditional Chinese medicine.
RESUMO
Screening and identification of potential compounds from herbal medicine is a prevailing way to find a lead for the development of innovative drugs. This promotes the development of new methods that are feasible in complex matrices. Here, we described a one-step reversible methodology to immobilize nuclear peroxisome proliferator-activated receptor gamma (PPARγ) onto amino microsphere coated with a DNA strand specifically binding to the receptor. The specific interaction allowed us to achieve the immobilization of PPARγ by mixing the DNA modified microspheres with E. coli lysates expressing the receptor. Characterization of the immobilized receptor was carried out by morphology and binding specificity analysis. Feasibility of immobilized PPARγ in the drug-receptor interaction analysis was performed by an injection amount-dependent method. Besides, immobilized PPARγ was also applied in screening modulators of the receptor from Coptidis Rhizoma extract. The binding of the screened compounds to PPARγ was examined by time-resolved fluorescence resonance energy transfer assay. The results showed that immobilized PPARγ was stable for thirty days with a high-specificity of ligand recognition at the subtype receptor level. Berberine and palmatine were the bioactive compounds of Coptidis Rhizoma specifically binding to PPARγ. The two compounds exhibited half maximal inhibitory concentrations of 4.11 and 2.98 µM during their binding to the receptor. We concluded that the current method is possible to become a common strategy for the immobilization of nuclear receptors, and the immobilized receptor is a high throughput method for recognizing and separating the receptor modulators from complex matrices including herbal medicine.
Assuntos
Coptis chinensis/química , Medicamentos de Ervas Chinesas/química , PPAR gama/química , Berberina/química , Alcaloides de Berberina/química , Transferência Ressonante de Energia de Fluorescência , Medicina Herbária , Humanos , Ligação ProteicaRESUMO
Drug discovery based on natural products like medicinal herbs remains challenging due to the technique limitations for rapidly screening and validating leads. To address the challenges, we employ the immobilized ß2- adrenergic recepotor (ß2-AR), an identified target of asthma, as the stationary phase in chromatographic column to screen compounds extracted from Stemonae Radix, Playtycodonis Radix, and Glycyrrhizae Radix et Rhizoma. To analyze binding properties of the extracted compounds to the immobilized receptors, we measured their retention behavior in the receptor chromatography and compared with six clinical asthma drugs. We identified tuberostemonine, platycodin D, and glycyrrhizic acid as the potential leads against asthma by our ß2-AR chromatography coupled with mass spectrum (MS). The association constants of the three compounds to ß2-AR were 2.85 × 10-5, 2.55 × 10-4, and 4.07 × 10-6 M with the dissociation rate constants of 6.91 ± 0.35, 11.88 ± 0.60, and 9.49 ± 0.64 min-1, respectively. Tuberostemonine, a pentacyclic Stemona alkaloids, presented the most optimum values of binding efficiency index (BEI) and surface efficiency index (SEI) as close to the diagonal of SEI-BEI optimization plane when it is compared with platycodin D, glycyrrhizic and the six clinical drugs. Our results suggest that tuberostemonine is a promising natural product to be developed for treating asthma because it exhibits better drug-like binding properties to ß2-AR than the clinical drugs. As such, we demonstrate a chromatographic strategy to identify bioactive natural products based on the ß2-AR immobilization, which can be widely adopted to screen natural products from mixture of herbal extracts.
Assuntos
Produtos Biológicos , Medicamentos de Ervas Chinesas , Cromatografia , Descoberta de Drogas , Glycyrrhiza , Receptores Adrenérgicos beta 2RESUMO
Inspired by the traditional Chinese herbal pair of Polygala tenuifolia-Acori Tatarinowii for treating epilepsy, 33 novel substituted cinnamic α-asaronol esters and analogues were designed by Combination of Traditional Chinese Medicine Molecular Chemistry (CTCMMC) strategy, synthesized and tested systematically not only for anticonvulsant activity in three mouse models but also for LDH inhibitory activity. Thereinto, 68-70 and 75 displayed excellent and broad spectra of anticonvulsant activities with modest ability in preventing neuropathic pain, as well as low neurotoxicity. The protective indices of these four compounds compared favorably with stiripentol, lacosamide, carbamazepine and valproic acid. 68-70 exhibited good LDH1 and LDH5 inhibitory activities with noncompetitive inhibition type, and were more potent than stiripentol. Notably, 70, as a representative agent, was also shown as a moderately positive allosteric modulator at human α1ß2γ2 GABAA receptors (EC50 46.3⯱â¯7.3⯵M). Thus, 68-70 were promising candidates for developing into anti-epileptic drugs, especially for treatment of refractory epilepsies such as Dravet syndrome.
Assuntos
Anisóis/química , Anticonvulsivantes/química , Cinamatos/química , Medicamentos de Ervas Chinesas/química , Ésteres/química , L-Lactato Desidrogenase/antagonistas & inibidores , Polygala/química , Regulação Alostérica , Animais , Anisóis/farmacologia , Anticonvulsivantes/farmacologia , Carbamazepina/química , Carbamazepina/farmacologia , Cinamatos/farmacologia , Dioxolanos/química , Dioxolanos/farmacologia , Desenho de Fármacos , Medicamentos de Ervas Chinesas/farmacologia , Ésteres/farmacologia , Humanos , Medicina Tradicional Chinesa , Camundongos , Estrutura Molecular , Neuralgia/prevenção & controle , Receptores de GABA-A/metabolismo , Relação Estrutura-Atividade , Ácido Valproico/química , Ácido Valproico/farmacologiaRESUMO
The past decade has witnessed the great promise of strategies for ligand discovery based on surface-immobilized GPCRs. We present here a method for preparation of immobilized GPCRs. Key features include covalent immobilization with high specificity and robust application in drug-receptor interaction analysis and ligand screening. In our example assay using beta2-adrenergic receptor (ß2-AR), the human DNA repair protein O6-alkylguanine-DNA alkyltransferase (hAGT) fusion receptor expressed in Escherichia coli was directly captured onto polyethylene glycol polyacrylamide (PEGA) resin. We observed even distribution and physiological functions of ß2-AR on the resin. The immobilized ß2-AR as a stationary phase enabled us to rapidly determine the binding of four drugs to ß2-AR. By coupling this assay to mass spectrometry, we screened rosmarinic acid as a bioactive compound targeting ß2-AR in Fructus Perillae. We concluded that O6-benzylguanine derivative-functionalized supporter is promising for specific immobilization of hAGT-tagged proteins; immobilized receptor chromatography has great potential in screening receptor-binding leads from herbal plants or traditional medicine recipes.
Assuntos
Cinamatos/farmacologia , Depsídeos/farmacologia , Descoberta de Drogas , Guanina/análogos & derivados , Ensaios de Triagem em Larga Escala , Receptores Adrenérgicos beta 2/metabolismo , Cinamatos/química , Depsídeos/química , Guanina/química , Guanina/metabolismo , Humanos , Ligantes , Perilla/química , Receptores Adrenérgicos beta 2/análise , Propriedades de Superfície , Ácido RosmarínicoRESUMO
Bornyl caffeate was initially discovered as a bioactive compound in medicinal plants. Despite the promising pharmacological activities including anti-tumor and antibacterial activities, the pharmacokinetics of the compound remain open. This work developed a high performance liquid chromatography-tandem mass spectrometric method for the determination of bornyl caffeate and caffeic acid (major metabolite and a main unit of bornyl caffeate) in vivo. Successful application of the method included identification of its metabolites and investigation on the drug pharmacokinetics. A total of 30 compounds were identified as the metabolites of bornyl caffeate in rats. We attributed these metabolites to phase I metabolic routes of reduction, oxidation, hydrolysis and phase II metabolic reactions of glucuronidation, sulfation, O-methylation and glycine. Glucuronidation, sulfation, O-methylation and reduction were the main metabolic pathways of bornyl caffeate. The method presented a linear range of 1-4000 ng mL-1. The pharmacokinetic profile of bornyl caffeate was found to be a three compartment open model, while caffeic acid fitted to a two compartment open model when it was administered alone or served as the main metabolite of bornyl caffeate. The time to peak concentration (T max) and the maximum plasma concentration (C max) of bornyl caffeate were 0.53 h and 409.33 ng mL-1. Compared with original caffeic acid, the compound displayed an increased half-life of elimination (T 1/2ß), area under the concentration time curve from 0 to t (AUC0-t ) and area under the concentration time curve from 0 to ∞ (AUC0-∞), a decreased half-life of absorption (T 1/2α) and an identical C max. Taking together, we concluded that bornyl caffeate is able to rapidly initiate therapeutic effect and last for a relatively long time in rats; metabolic pathways of O-methylation and reduction is key to interpret the mechanism and toxicity of bornyl caffeate.
RESUMO
INTRODUCTION: The distribution of metabolites in the different root parts of Cortex Moutan (the root bark of Paeonia suffruticosa Andrews) is not well understood, therefore, scientific evidence is not available for quality assessment of Cortex Moutan. OBJECTIVE: To reveal metabolomic variations in Cortex Moutan in order to gain deeper insights to enable quality control. METHODS: Metabolomic variations in the different root parts of Cortex Moutan were characterised using high-performance liquid chromatography combined with mass spectrometry (HPLC-MS) and multivariate data analysis. The discriminating metabolites in different root parts were evaluated by the one-way analysis of variance and a fold change parameter. RESULTS: The metabolite profiles of Cortex Moutan were largely dominated by five primary and 41 secondary metabolites . Higher levels of malic acid, gallic acid and mudanoside-B were mainly observed in the second lateral roots, whereas dihydroxyacetophenone, benzoyloxypaeoniflorin, suffruticoside-A, kaempferol dihexoside, mudanpioside E and mudanpioside J accumulated in the first lateral and axial roots. The highest contents of paeonol, galloyloxypaeoniflorin and procyanidin B were detected in the axial roots. Accordingly, metabolite compositions of Cortex Moutan were found to vary among different root parts. CONCLUSION: The axial roots have higher quality than the lateral roots in Cortex Moutan due to the accumulation of bioactive secondary metabolites associated with plant physiology. These findings provided important scientific evidence for grading Cortex Moutan on the general market.
Assuntos
Medicamentos de Ervas Chinesas/química , Metaboloma , Metabolômica/métodos , Paeonia/química , Extratos Vegetais/análise , Raízes de Plantas/química , Cromatografia Líquida de Alta Pressão , Espectrometria de Massas , Estrutura Molecular , Especificidade de Órgãos , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificaçãoRESUMO
A novel high performance liquid chromatography-electrochemical detector (HPLC-ECD) analytical system was developed in this study by integratedly utilizing ionic liquid (IL) of 1-butyl-3-methylimidazolium bromide and an additive of gold nanoparticles. The resulted pilot study was first performed to assess the effects of 1-butyl-3-methylimidazolium bromide and gold nanoparticles on the chromatographic characteristics of five phenolic acids in Xuebijing injection, including danshensu (DSS), protocatechuic acid (PA), protocatechuic aldehyde (PAH), hydroxy safflower yellow A (HSYA) and ferulic acid (FA). It was notable to observe that retainability of the phenolic acids were markly lowered by IL addition. Compared with the cases without IL addition, the retention times of DSS, PA, PAH, HSYA and FA have decreased 2.851, 1.532, 1.53, 0.818 and 0.552 min, respectively when 0.6% IL in the mobile phase. In addition, the corresponding theoretical plate numbers and peak areas for these compounds were significantly increased. Area response for DSS, PA, PAH, HSYA and FA were enhanced by 772%, 628%, 584%, 703% and 600%, respectively. It was observed that nano-gold catalysis power enabled peak areas of DSS, PAH, FA and PA to enhance 5.7, 6.2, 8.5 and 66.5 times relative to the case with addition of IL. Altogether, the optimized HPLC-ECD system was successfully applied to the pharmacokinetics study of Xuebijing injection with underlying applicability to in vivo and in vitro analysis of a variety of natural product from Chinese medicine plants, TCM formulae and associated patent TCM preparation.
Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Medicamentos de Ervas Chinesas/química , Medicamentos de Ervas Chinesas/farmacocinética , Hidroxibenzoatos/análise , Hidroxibenzoatos/farmacocinética , Animais , Medicamentos de Ervas Chinesas/administração & dosagem , Técnicas Eletroquímicas/métodos , Ouro/química , Hidroxibenzoatos/administração & dosagem , Imidazóis/química , Injeções , Líquidos Iônicos/química , Masculino , Nanopartículas/química , Ratos , Ratos Sprague-Dawley , Sensibilidade e EspecificidadeRESUMO
Gallic acid (GA) and its metabolites are polyphenolic compounds present in daily diets and herbal medicines. To understand the GA effects on the endogenous metabolism of mammals, we systematically analyzed the metabonomic responses of rat plasma, liver, urine, and feces to a single GA dosage of 120 and 600 mg/kg, which were below the no-obvious-adverse-effect-level of 1 g/kg for rats. Clinical chemistry and histopathological assessments were conducted to provide complementary information. Our results showed that GA intake induced significant metabonomic changes in multiple rat biological matrices. Such changes were more outstanding in liver than in the other matrices and clearly showed dose- and time-dependence. The results suggested GA-induced promotion of oxidative stress as the major effect. High-dose GA caused significant metabolic changes involving glycogenolysis, glycolysis, TCA cycle, and metabolism of amino acids, purines, and pyrimidines, together with gut microbiota functions. Low-dose GA only caused some urinary metabonomic changes and to a much less degree. The GA-induced liver metabonomic changes were not completely recoverable within a week, although such recovery completed in plasma, urine, and feces within 80 h. These findings provided new essential information on the effects of dietary polyphenols and demonstrated the great potential of this nutrimetabonomics approach.
Assuntos
Ácido Gálico/farmacologia , Fígado/efeitos dos fármacos , Metaboloma , Administração Oral , Aminoácidos/sangue , Aminoácidos/urina , Animais , Ciclo do Ácido Cítrico , Relação Dose-Resposta a Droga , Fezes/química , Glicogenólise , Glicólise , Fígado/metabolismo , Espectroscopia de Ressonância Magnética , Masculino , Estresse Oxidativo , Purinas/sangue , Purinas/urina , Pirimidinas/sangue , Pirimidinas/urina , Ratos , Ratos Wistar , Fatores de TempoRESUMO
Identification of new compounds especially those with new skeletons from plant kingdom has long been a vital aspect for understanding phytochemistry, plant metabolisms and discovering new bioactive compounds. In this study, we identified and isolated three novel polyphenolic compounds, origanine A-C, from a well-researched plant Origanum vulgare L. using the hyphenated LC-DAD-SPE-NMR/MS methods. Based on the combined information from UV-visible, accurate mass and 2D NMR spectra together with computational calculations, we found that these compounds all had a novel skeleton of cyclohexenetetracarboxylic acids attached with some well-known bioactive moieties including 3,4-dihydroxyphenyl, 4-(ß-d-glucopyranosyloxy)benzyl alcohol (gastrodin), and 3-(3,4-dihydroxyphenyl)lactic acid (danshensu) residues. These findings provided crucial information to fill the gaps in our knowledge in terms of the plant secondary metabolism. This study also indicated the necessity for further research in plant secondary metabolism for even well-studied plants and demonstrated the powerfulness of the hyphenated LC-DAD-SPE-NMR/MS methods for comprehensive analysis of plant metabolites in particular for discovering new natural compounds.
Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Espectroscopia de Ressonância Magnética/métodos , Espectrometria de Massas/métodos , Origanum/química , Extratos Vegetais/química , Polifenóis/química , Extração em Fase Sólida/métodos , Extratos Vegetais/isolamento & purificação , Polifenóis/isolamento & purificaçãoRESUMO
A receptor affinity chromatographic selection method was developed for screening the bioactive compounds binding to beta(2)-adrenoceptor (beta(2)-AR) in Coptidis rhizome. The bioactive compounds were analyzed by molecular recognition with a beta(2)-AR affinity column. The retention compounds eluted from the beta(2)-AR column were separated online with reverse-phase high-performance liquid chromatography by column switching technology, and identified by a coupled ion-trap mass spectrometer. Four compounds were screened as the bioactive compounds of Coptidis rhizome and identified as 2,9,10-trimethoxy-3-hydroxyl-protoberberine (jateorhizine), 2,3-methylenedioxy-9-methoxy-protoberberine, 2,3,9,10-tetramethoxy-protoberberine (palmatine) and 2,3-methylenedioxy-9,10-dimethoxy-protoberberine (berberine). The association constants of jatrorrhizine, palmatine and berberine to the beta(2)-AR were determined by the zonal elution method with standards. Berberine and palmatine had only one type of binding site on the immobilized beta(2)-AR. Their association constants were (2.28+/-0.11)x10(4)/M and (3.00+/-0.10)x10(4)/M, respectively. Jatrorrhizine had at least two type of binding sites on the immobilized beta(2)-AR, and the corresponding association constants were (2.20+/-0.09)x10(-4)/M and (6.78+/-0.001)x10(5)/M.
Assuntos
Cromatografia de Afinidade/métodos , Medicamentos de Ervas Chinesas/química , Pulmão/química , Receptores Adrenérgicos beta 2/química , Animais , Coptis chinensis , Medicamentos de Ervas Chinesas/metabolismo , Pulmão/metabolismo , Ligação Proteica , Coelhos , Receptores Adrenérgicos beta 2/metabolismoRESUMO
Plant metabonomic analysis is essential for understanding plant systems responses to osmotic stresses. To understand the comprehensive metabolic responses of Salvia miltiorrhiza Bunge (SMB) to continuous and exhaustive water depletion, we characterized the SMB metabonomic variations induced by three different drying processes using the combined NMR and LC-DAD-MS method. NMR results showed that SMB extracts were dominated by 29 primary metabolites such as sugars, carboxylic acids and amino acids, which were comprehensively reported for the first time, and 8 secondary metabolites including polyphenolic acids and diterpenoids. LC-DAD-MS methods detected 44 secondary metabolites, among which 5 polyphenolic acids together with genipin, umbelliferone and tormentic acid were found for the first time in this plant. We found that aqueous methanol was efficient in extracting both primary metabolites and polyphenolic acids, whereas chloroform-methanol was effective in selectively extracting diterpenoids. We further found that air- and sun-drying markedly affected both primary and secondary metabolisms of SMB by enhancing tanshinone and glutamate-mediated proline biosynthesis and altering carbohydrate and amino acid metabolisms. The shikimate-mediated biosynthesis of polyphenolic acids was promoted by air-drying but suppressed by sun-drying. These findings fill the gap of our understandings to the metabolic responses of S. miltiorrhiza Bunge to water depletion and demonstrated effectiveness of the combined NMR and LC-DAD-MS methods in plant metabonomic analysis.
Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Metabolômica/métodos , Ressonância Magnética Nuclear Biomolecular/métodos , Salvia miltiorrhiza/metabolismo , Água/metabolismo , Interpretação Estatística de Dados , Medicamentos de Ervas Chinesas/química , Análise de Fourier , Liofilização , Espectrometria de Massas , Análise Multivariada , Pressão Osmótica , Raízes de Plantas/química , Raízes de Plantas/metabolismo , Análise de Componente Principal , Salvia miltiorrhiza/química , Estresse FisiológicoRESUMO
OBJECTIVE: To study the influence of processing on metabolism of the main component of bitter almond-amygdalin in rat. METHOD: The blood was collected at different times after amygdalin given by injection and oral, bitter almond and its processed production given by oral respectively, and then detected by both HPLC and HPLC-MS(n) methods after extraction pretreatment. RESULT: After injection, amygdalin was absorbed in prototype to blood rapidly, while the other three kinds of medicine given by oral were all not detected the prototype of amygdalin, but two metabolites were detected which were isomers of prunasin confirmed by mass spectrometry. The metabolic pathway of prunasin in processed bitter almond group was markedly different from the bitter almond group. CONCLUSION: Processing has a significant effect on bitter almond metabolic processes in rats.
Assuntos
Amigdalina/metabolismo , Prunus/química , Amigdalina/análise , Animais , Feminino , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
Metabolite identification is a key step for metabonomics study. A fully automated hyphenation of HPLC-diode-array detector (DAD) mass spectrometry (MS) solid phase extraction (SPE)-NMR spectroscopy (HPLC-DAD-MS-SPE-NMR) is one of the most efficient methods to determine the structure of a given unknown metabolite in a complex mixture (metabonome) and hence represents one of the most important analytical techniques for the further development of metabonomics. In this review, some recent applications of this technique in identifying novel and trace metabolites in plant extracts and drug metabolism have been discussed. Modification of this hyphenated technique, enabling multiple trappings of strong polar metabolites for biofluids, needs further development.
Assuntos
Metabolômica , Preparações de Plantas/química , Ácido Quínico/química , Química Farmacêutica , Cromatografia Líquida de Alta Pressão , Lasers Semicondutores , Espectroscopia de Ressonância Magnética/métodos , Estrutura MolecularRESUMO
The molecular compositions of rosemary ( Rosmarinus officinalis L.) extracts and their dependence on extraction solvents, seasons, and drying processes were systematically characterized using NMR spectroscopy and multivariate data analysis. The results showed that the rosemary metabonome was dominated by 33 metabolites including sugars, amino acids, organic acids, polyphenolic acids, and diterpenes, among which quinate, cis-4-glucosyloxycinnamic acid, and 3,4,5-trimethoxyphenylmethanol were found in rosemary for the first time. Compared with water extracts, the 50% aqueous methanol extracts contained higher levels of sucrose, succinate, fumarate, malonate, shikimate, and phenolic acids, but lower levels of fructose, glucose, citrate, and quinate. Chloroform/methanol was an excellent solvent for selective extraction of diterpenes. From February to August, the levels of rosmarinate and quinate increased, whereas the sucrose level decreased. The sun-dried samples contained higher concentrations of rosmarinate, sucrose, and some amino acids but lower concentrations of glucose, fructose, malate, succinate, lactate, and quinate than freeze-dried ones. These findings will fill the gap in the understanding of rosemary composition and its variations.