柱前衍生化反相高效液相色谱法测定板蓝根中的氨基酸

建立了板蓝根中指标性成分精氨酸、脯氨酸和谷氨酸的柱前衍生化反相高效液相色谱分析方法。以2,4-二硝基氟苯为衍生化试剂,将氨基酸在碱性条件下衍生化,然后直接进样进行高效液相色谱测定。采用的色谱条件为：色谱柱Sinochrom ODS-BP柱(250 mm×4.6 mm i.d.,5 μm),流动相醋酸钠缓冲溶液(pH 6.4)-乙腈(体积比为850∶150),检测波长360 nm,采用外标法定量。该文同时对谷氨酸、精氨酸和脯氨酸进行定性,并对精氨酸和脯氨酸进行定量。精氨酸和脯氨酸的进样量分别为0.627～5.016 μg和0.874～7.000 μg时,其峰面积与进样量呈良好的线性关系(r分别为0.9996和0.9995)。精氨酸和脯氨酸的平均回收率分别为98.5%(相对标准偏差（RSD）为2.5%（n=8))和98.4%(RSD为2.3%（n=8))。对板蓝根样品进行了测定,结果表明,板蓝根中精氨酸的含量最高,脯氨酸次之,谷氨酸含量很少。该法简便、准确、可靠,适用于板蓝根药材的质量控制。     

Development of methods for determination of the residues of 15 pesticides in medicinal herbs Isatis indigotica Fort. by capillary gas chromatography with electron capture or flame photometric detection

Two multiresidue methods were developed for the determination of 15 pesticides (organochlorines, organphosphorus compounds, pyrethroids, and fungicides) in medicinal herbs Isatis indigotica Fort. and its formulations. The analytical procedure is based on ultrasonic assisted extraction and liquid-liquid extraction (LLE). After solvents were added, the raw material or granule sample was sonicated in an ultrasonic water bath and then centrifuged, filtered, and cleaned up by LLE. The infusion sample was extracted with petroleum ether by LLE. The pesticide residues were determined by capillary gas chromatography with electron-capture or flame photometric detection. Recoveries with the method at concentrations between 0.4 microg/kg and 10 mg/kg ranged from 70.2 to 119.5% for raw material, 73.2 to 105.1% for granule formulation, and 72.8 to 113.3% for infusion formulation. The relative standard deviation values were <20% for all of the pesticides studied. The pesticide detection limits were within the ranges 0.3-0.5 microg/L for endosulfan, 3-7.5 microg/L for pyrethroids, 0.7-32.5 microg/L for organophosphorus pesticides, and 0.1-0.6 microg/L for the other pesticides. The proposed methods are simple and rapid and provide simultaneous determination of pesticide residues in Isatis indigotica Fort. with acceptable recoveries and repeatability and an adequate limit of determination.     

Isolation,identification, biological estimation,and profiling of glucosinolates in Isatis indigotica roots

Abstract

Four main glucosinolates (progoitrin, epiprogoitrin, gluconapin, and neoglucobrassicin) were isolated and identified from Isatis indigotica roots (Isatidis Radix). Among these, neoglucobrassicin was the first indole glucosinolate isolated from this herb, and its structure was elucidated by NMR, MS, and X-ray diffraction analysis. The glucosinolate components of Isatidis Radix have been profiled by combined use of a conventional preparative column chromatography method and a new UHPLC-QTOF-MS/MS method, and validated by simultaneous analysis of intact and desulfurized GLS. A total of 16 glucosinolates (GLS) including 12 aliphatic GLS, two aromatic GLS, and two indole GLS were identified on the basis of their UV profiles, MS/MS data, and/or by comparison with reference substances. Among these nine GLS were reported from I. indigotica for the first time. The total GLS, four isolated GLS, and (R,S)-goitrin (a degraded product of progoitrin and epiprogoitrin) have been estimated in vitro for anti-influenza viral, anti-bacterial, and anti-inflammatory activities. All the glucosinolate samples were inactive against H3N2 virus and four pathogenic bacteria, and two previously unnoticed GLS (gluconapin and neoglucobrassicin) showed significantly anti-inflammatory activity against the respiratory burst of mouse macrophages.     

不同生长期中菘蓝多糖和氨基酸含量变化规律研究

分别采用苯酚-硫酸法、茚三酮比色法测定菘蓝中总多糖和游离总氨基酸含量.分析结果表明,在不同的生长季节,根中游离氨基酸、总多糖含量逐月上升,均在11月份含量最高;叶中游离氨基酸含量先上升,后下降,以8月份和10月份含量较高,总多糖含量逐月上升,在十一月份含量最高.菘蓝生长过程中,根和叶中的总多糖和游离总氨基酸含量随着发育进程而发生变化.这一变化规律为中药的规范化种植(GAP)提供了实验依据.     

Determination of indican, isatin, indirubin and indigotin in Isatis indigotica by liquid chromatography/electrospray ionization tandem mass spectrometry

The roots and leaves of Isatis indigotica, named 'Ban-Lan-Gen' and 'Da-Qing-Ye', respectively, are widely used for the treatment of influenza, viral pneumonia, mumps, pharyngitis, and hepatitis. The indoxyl derivatives detected in the roots and leaves of I. indigotica have been reported to be biologically active. In the present study, a liquid chromatography/electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) method was developed to determine indican, isatin, indirubin and indigotin in the roots and leaves of I. indigotica. The method has been validated for linearity, precision and accuracy. Using multiple reaction monitoring (MRM), the limits of detection (LODs) were determined as 0.004 ng for indican, 0.01 ng for isatin, 0.01 ng for indrubin and 0.03 ng for indigotin, while the limits of quantitation (LOQs) were 0.015 ng for indican, 0.04 ng for isatin, 0.04 ng for indirubin and 0.1 ng for indigotin. Compared with previously reported methods, the current method is more rapid, selective and sensitive. This is the first report of the LC/MS/MS determination of indican, isatin, indirubin and indigotin.     

Rhizonin A from Burkholderia sp. KCTC11096 and its growth promoting role in lettuce seed germination

We isolated and identified a gibberellin-producing Burkholderia sp. KCTC 11096 from agricultural field soils. The culture filtrate of plant growth promoting rhizobacteria (PGPR) significantly increased the germination and growth of lettuce and Chinese cabbage seeds. The ethyl acetate extract of the PGPR culture showed significantly higher rate of lettuce seed germination and growth as compared to the distilled water treated control. The ethyl acetate fraction of the Burkholderia sp. was subjected to bioassay-guided isolation and we obtained for the first time from a Burkholderia sp. the plant growth promoting compound rhizonin A (1), which was characterized through NMR and MS techniques. Application of various concentrations of 1 significantly promoted the lettuce seed germination as compared to control.     

Light production in alkaline mixtures of reducing agents and dimethylbiacridylium nitrate.

Abstract— Kinetic studies were made of light production and 0₂ absorption elicited by treatment of dimethylbiacridylium hydroxide [D(OH)₂] with reducing agents in alkaline aqueous solutions. D(OH)2 addition stimulated O₂ uptake which proceeded with zero-order kinetics until nearly all of the O₂ or of the D(OH)₂ was converted to end products. At the termination of the reactions with fructose as reductant the D(OH)₂ was converted to methylacridone and to dimethylbiacridene each compound accounting for approximately one-half the D(OH)₂ consumed. O₂ was reduced to H₂O₂. With hydroxylamine as the reducing agent the emitted light intensity was related to the first power of the D(OH)₂ concentration and the rate of O₂ uptake to the square root of the D(OH)₂. The disappearance of D(OH)₂ in these circumstances was by a first order or pseudo-first order rate. Fructose as a reducing agent by contrast resulted in an O₂ uptake nearly independent of D(OH)₂ concentration over a range from 1 × 10^-5M-1 × 10^-4M, while the light intensity and disappearance of D(OH)₂ followed a one-half order rate. O₂ uptake was by zero order kinetics and the oxidation of fructose proceeded at the same rate as was found with ferricyanide as oxidant. The kinetics, quantum yields and temperature dependence of the fructose reactions were compared with similar reactions employing H₂O₂ as the light eliciting reagent. The results are interpreted as indicating that D(OH)₂ acts as a chain initiator in a manner analogous to better known, radical producing compounds found to accelerate hydrocarbon autooxidations.     

Phenolic compounds from Merremia umbellata subsp. orientalis and their allelopathic effects on Arabidopsis seed germination

A bioassay-directed phytochemical study was carried out to investigate potential allelochemicals of the invasive plant Merremia umbellata subsp. orientalis (Hall. f.). Eight phenolic compounds, including a salicylic acid (SA)-derived new natural product, SA 2-O-β-D-(3',6'-dicaffeoyl)-glucopyranoside (1), and seven known ones 2-8 were isolated and identified from two bioactive sub-fractions of the acetone extract of this plant. The structure of new compound 1 was established by spectral and chemical methods. The potential allelopathic effects of these compounds at 0.5 and 1.0 mM concentrations on the germination of Arabidopsis seeds were tested. Results showed that 2 remarkably inhibited seed germination at concentrations as low as 0.5 mM. Compound 3 only moderately inhibited seed germination at 0.5 mM, but displayed strong inhibitory bioactivity at 1.0 mM concentration. Compounds 4 and 5 showed only slight inhibitory bioactivity at 1.0 mM, while the other compounds showed no obvious inhibitory effects.     

Simultaneous isolation of artemisinin and its precursors from Artemisia annua L. by preparative RP-HPLC

It is still a major challenge to simultaneously isolate artemisinin and its precursors, especially dihydroartemisinic acid and artemisinic acid, from herbal Artemisia annua. A rapid, economical and automatical chromatographic separation process to isolate and purify artemisinin, dihydroartemisinic acid and artemisinic acid at the same time on a preparative scale was developed. The procedure included solvent extraction of ground Artemisia annua leaves by refluxing and purification of crude extract by preparative reverse-phase high-performance liquid chromatography (RP-HPLC). Fractions containing artemisinin and its precursors were collected and identified by gas chromatography and mass spectrometry. High purity of artemisinin, dihydroartemisinic acid and artemisinic acid was obtained by preparative HPLC with a C(18) column and 60% acetonitrile in water as the mobile phase. The techniques described here are useful tools for the preparative-scale isolation of artemisinin and its precursors in a fast, cost-effective and environmental friendly manner.     

Monoterpenic and norisoprenoidic glycoconjugates of Vitis vinifera L. cv. Melon B. as precursors of odorants in Muscadet wines.

The volatile monoterpenic and norisoprenoidic compounds released by glycosidase enzyme hydrolysis of C18 reversed-phase isolates from the juice of Vitis vinifera L. cv. Melon B. have been qualitatively and quantitatively determined using GC-MS and GC-FID. The components analyzed were broadly similar to those previously reported for other varieties but the level of bound p-menth-1-en-7,8-diol was higher in this cultivar. Then the monoterpenic and norisoprenoidic volatiles released from the same glycosidic extracts under mild acid conditions, mimicking wine aging conditions, have been analyzed using GC-Olfactometry and GC-MS. The most odorous compounds detected were p-cymene, terpinen-4-ol, cis- and trans-vitispiranes, 1,6,6-trimethyl-1,2-dihydronaphtalene (TDN), beta-damascenone and riesling acetal. To assess their potential levels in corresponding wines after ageing, most of these odorants were generated by harsh acid hydrolysis from the precursors extracts and quantitatively determined using SPME and GC-MS/MS. For the development and application of this analysis, the odorants not commercialy available were synthesized. The total amounts of norisoprenoidic odorants generated by acid hydrolysis of the glycosidic extracts were shown to be proportional to the total amounts of these precursors.     

LC-DAD UV and LC-MS for the Analysis of Isoflavones and Flavones from In Vitro and In Vivo Biomass of Genista tinctoria L.

Conditions were determined for the separation of a complex set of isoflavones and flavones (free aglycones, monoglucosides, diglucosides and esters) by HPLC-DAD UV and MS detection. A good separation of all analytes was obtained and satisfactory peak shapes were achieved by gradient elution on an RP C18 column. The method was then applied to carry out qualitative and quantitative analysis of bioflavonoids present in the herb and in vitro cultures of Genista tinctoria L. Electrospray ionisation mass spectrometry in the negative mode was used to identify individual flavonoids in G. tinctoria plant material. Photodiode array detection was also utilised in flavonoid characterisation. All flavonoid compounds were then quantified using an internal standard. The method developed is useful for monitoring the process of flavonoid biosynthesis in biomass obtained through in vitro cultures.     

Spectral and photophysical studies of substituted indigo derivatives in their keto forms.

The spectral and photophysical properties of indigo derivatives with di-, tetra-, and hexa-substitution in their neutral (keto) form are investigated in solution. The study comprises absorption and emission spectra, together with quantitative measurements of quantum yields of fluorescence (phi(F)) and singlet oxygen formation (phi(Delta)) and fluorescence lifetimes. The energy difference between the HOMO and LUMO orbitals is dependent on the degree (number of groups) and relative position of substitution. The phi(F) and phi(Delta) values were found to be very low S(0) internal conversion yields and thus, with the other data, to determine the rate constants for all decay processes. From these, several conclusions are drawn. Firstly, the radiationless rate constants, k(NR) , clearly dominate over the radiative rate constants, k(F) , (and processes). Secondly, the main deactivation channel for the compounds in their keto form is the radiationless S(1) approximately approximately -->S(0) internal conversion process. Finally, although the changes are relatively small, internal conversion yield seems to be independent of the overall pattern of substitution. A more detailed investigation of the decay profiles with collection at the blue and red emission of the fluorescence band of indigo and one di-substituted indigo reveals the decays to be bi-exponential and that at longer emission wavelengths these appear to be associated with both rise and decay times indicating that two excited species exist, which is consistent with a keto-excited form giving rise (by fast proton transfer) to the enol-form of indigo. Evidence is presented which supports the idea that intramolecular (and possibly some intermolecular) proton transfer can explain the high efficiency of internal conversion in indigo.     

Studies in stability constants of schiff base hydrazone complexes with transition metal ions. Effect of ligand on seed germination

Spectrophotometric investigation of Cu (II), Ni(II), Co(II), and Fe(III) complexes with 2,4-dihydroxyacetophonone 2,4-dichlorobenzoylhydrazone (H₂L¹) and 2,4-didydroxy-5-nitroacetophenone 2,4-dichlorobenzoylhydrazone (H₂L²) shows 1: 1 and 1: 2 complex formation between the pH range of 3.0 to 6.0 and also studied by jobs variation method at 0.1 M ionic strength at 30 ± 1°C specrtophotometrically. The conditional stability constants are determined for 1: 1 complexes. Effect of H₂L¹ and H₂L² ligand and its complexes on seed germination is studied.     

Gas chromatographic determination of carboxylic acid chlorides and residual carboxylic acid precursors used in the production of some penicillins.

An improved gas chromatographic method is described for the simultaneous determination of carboxylic acid chlorides and related carboxylic acids used in the production of some commercial semisynthetic penicillins. The acid chloride reacts with diethylamine to form the corresponding diethylamide. Carboxylic acid impurities are converted to trimethylsilyl esters. The two derivatives are separated and quantitated in the same chromatographic run. This method, an extension of the earlier procedure of Hishta and Bomstein (1), has been applied to the acid chlorides used to make oxacillin, cloxacillin, dicloxacillin, and methicillin (Figure 1); it shows promise of application to other acid chlorides. The determination is more selective than the usual titration methods, which do not differentiate among acids with similar pK's. Relative standard deviations of the acid chloride determination are 1.0-2.5%. Residual carboxylic acid can be repetitively determined within a range of 0.6% absolute.     

Study of allelopathic effects of Eucalyptus erythrocorys L. crude extracts against germination and seedling growth of weeds and wheat

Allelopathic materials inside a tree can produce positive or negative change in the survival, growth, reproduction and behaviour of other organisms if they escape into the environment. To assess these effects, this work was carried out to evaluate the allelopathic impact of Eucalyptus erythrocorys L. on seed germination and seedling growth of two weeds: Sinapis arvensis L. and Phalaris canariensis L.; on one cultivated crop: Triticum durum L. Aqueous; and on ethanolic leaf extracts of E. erythrocorys L. The study was effected using four concentrations (10, 20, 25 and 30 μL/mL) while distilled water was used as a control. The results showed that the E. erythrocorys L. crude extracts had an inhibitory effect on seed germination and seedling growth of both studied weeds and wheat. The inhibition rate was increased by the increase in extract concentration. Only ethanolic extracts of E. erythrocorys L. induced a significant inhibition of seed germination of durum wheat. The effect of E. erythrocorys L. crude extracts was more severe on weeds than on durum wheat. These results indicate that the seedling growth, especially radicle elongation, was the more sensitive indicator to evaluate the effects of extracts than was the seed germination.     

Mesua ferrea L. seed oil based highly thermostable and biodegradable polyester/clay nanocomposites

Nanotechnology holds the prospective for pervasive and avant-garde changes to improve performance of materials. Mesua ferrea L. seed oil based highly thermostable and biodegradable polyester/clay nanocomposites have been prepared at different dose levels of nanoclay. The highly branched polyester resin was synthesized by condensation of 2,2-bis(hydroxymethyl) propionic acid with M. ferrea L. seed oil based carboxyl terminated pre-polymer. FTIR, rheometric, XRD, SEM and TEM studies unmasked well-dispersed partially exfoliated nanocomposites with good interfacial interaction. The results showed 2.6 and 6 times increase of tensile strength and elongation at break respectively, and about 150 °C increase of thermostability by the influence of nanoclay loading (1-5 weight %) compared to the pristine polymer. Two strains of Pseudomonas aeruginosa (PN8A1 and vs1) and one Bacillus subtilis strain (MTCC73) were used to study microbial degradation of pristine polymer and nanocomposite systems, with the revelation of enhanced degradation with increasing nanoclay loading.     

Enhanced production of L-DOPA in cell cultures of Mucuna pruriens L. and Mucuna prurita H

A comparative study on the production of 3,4-dihydroxyphenylalanine (L-DOPA) was carried out in cell cultures of two Mucuna species by elicitor treatment and precursor feeding. The influence of elicitors and the precursor molecule on L-DOPA production, polyphenol oxidase (PPO) and tyrosinase activities was also studied. Callus cultures were initiated in Mucuna pruriens L. and Mucuna prurita H. on MS medium supplemented with BAP and IAA at different concentrations. Suspension cultures were established in MS liquid medium supplemented with BAP, IAA, the elicitors methyl jasmonate, chitin and pectin or the precursor L-tyrosine at different concentrations for L-DOPA production. Compared to the controls, several-fold increases in L-DOPA concentration were observed in elicitor-treated and precursor-fed suspension cultures of both plant species. L-DOPA concentrations were comparatively higher in precursor-fed cultures than those receiving elicitor treatments. A parallel increase in tyrosinase and PPO levels was also observed. Loss of cell viability was observed at high concentrations of elicitor-treated cultures, whereas L-tyrosine did not cause any cell death. Compared to elicitor treatments, precursor feeding resulted in higher concentrations of L-DOPA production and tyrosinase activity. The efficacy of L-DOPA production was found to be higher for suspension cultures of M. pruriens compared to M. prurita in all treatments.     

Plasma activated water: the next generation eco-friendly stimulant for enhancing plant seed germination,vigor and increased enzyme activity,a study on black gram (Vigna mungo L.)

Plasma Chemistry and Plasma Processing - Chemical fertilization in agriculture is threatening to the ecosystem. Therefore, the use of eco-friendly stimulant for crop revolution is highly desirable....     

Macromolecular engineering of polylactones and polylactides. XV. Poly(D,L)-lactide macromonomers as precursors of biocompatible graft copolymers and bioerodible gels

The functional aluminum alkoxide, Et₂Al? O? (CH₂)₂? O-C(O)? C(CH₃)?CH₂, is a very effective initiator for the (D ,L )-lactide (LA) polymerization in toluene at 707deg;C. The coordination-insertion type of polymerization is living and exclusively yields linear P (D ,L )-lactide macromonomers of a predictable molecular weight and a narrow molecular weight distribution. IR and ¹H-NMR studies show that the methacryloyl group of the initiator is selectively and quantitatively attached to one chain end, whereas the second extremity is systematically a hydroxyl function resulting from the hydrolysis of the living growing site. α,ω-Dimethacryloyl-P(D ,L )-lactides, i.e., α,ω-macromonomers, have also been successfully synthesized by the additional control of the termination step, i.e., by reaction of Al alkoxide end groups with methacryloyl chloride. α-Macromonomer and α,ω-macromonomer P(D ,L )-lactides are easily free-radical copolymerized with 2-hydroxyethyl methacrylate (HEMA), resulting in a hydrophilic poly (HEMA) backbone grafted with hydrophobic P(D ,L )-lactide subchains and a biodegradable amphiphilic network, respectively. © 1994 John Wiley & Sons, Inc.