Purification, structure characterization and antioxidant activity of polysaccharides from Saposhnikovia divaricata / 中国天然药物

Polysaccharide from traditional Chinese herb, Saposhnikovia divaricata (Turcz.) Schischk. (SD) was extracted, fractionated and characterized in this work. Four fractions were prepared. Their molecular weight, monosaccharide compositions, linkage modes and structural properties were characterized with SEC-MALS-RI, HPAEC-PAD, GC-MS and NMR. SDP1 was assigned as a 1, 4-α-glucan with small amount of O-6 linked branches. SDP2 contained a big amount of the 1, 4-α-glucan and a small amount of arabinogalactan, while SDP3 possessed relatively lower amount of the 1, 4-α-glucan and a big amount of the arabinogalactan. SDP4 was defined as a pectic arabinogalactan. Four fractions showed antioxidant activities in both molecular and cellular levels and their activity was ranked as SDP4 ≈ SDP3>SDP2>SDP1. The 1, 4-α-glucan in SDP1 had the weakest, while SDP3 and SDP4 showed similar and the highest antioxidant activity. The arabinogalactan was the major component of both SDP3 and SDP4, which significantly contributed to the antioxidant activity of SDP.

Effect of P-glycoprotein on the transmembrane absorption and metabolism of quercitrin on Caco-2 cells / 国际药学研究杂志

Objective To explore the role of P-glycoprotein(P-gp)on cellular uptake and metabolism in the transmembrane transport of quercitrin.Methods Caco-2 cell monolayer and P-gp inhibitor Cyclosporin A(CysA)were used in the study.Quercitrin, quercetin,isorhamnetin and tamarixetin were determined by LC-MS to study cellular uptake and metabolism of quercitrin on Caco-2 cells.Results Uptake of quercitrin by Caco-2 cells:in different concentration groups of quercitrin coincubating with and without Cy?sA,intracellular accumulation presented the following characteristics:the amount of quercitrin first rose,reached the peak in 60 min and then declined to a steady-state in 120 min.And meanwhile there were significant differences between the two different processing groups incubating with and without CysA(P<0.05);quercetin was detected in all groups(3.0,9.0 and 27.0 mg/L).But in the higher concentration groups incubating with and without CysA,the intracellular quercetin presented a characteristics similar to its original glycosides and showed a significant difference(P<0.05),while the other groups showed no concentration-and time-dependence.At the same time,isorhamnetin and tamarixetin were detected in two higher concentration groups incubating with and without CysA, which showed the trend similar to the original glycosides but no significant difference was obtained between the two processing groups(P>0.05).Isorhamnetin and tamarixetin were not detected in the low and middle concentration groups.Transmembrane transport:on the basal lateral of all groups,the content of the quercitrin in 150 min incubation time showed a trend of continuous rise,and there was no significant difference between the two processing groups.Quercetin,isorhamnetin and tamarixetin were not detected.Conclu?sion Intact quercitrin could be absorbed into the Caco-2 cells and transported across the Caco-2 cell monolayer,and suffered a series of further metabolism in the Caco-2 cells and the basal side of Caco-2 cell monolayer,leading to different characteristics between intra?cellular accumulation and transmembrane transport.P-gp reduces the transmembrane transport of quercitrin by its efflux function,but did not involved in quercitrin metabolism.

Determination of morroniside concentration in rat plasma by high performance liquid chromatography-tandem mass spectrometry / 中国中药杂志

Morroniside, an iridoid glycoside extracted from Cornus officinalis, has multiple pharmacological effects such as neuroprotection. This study took the lead in establishing a method for determining morroniside concentration in rat plasma by high performance liquid chromatography-tandem mass spectrometry. Plasma samples were processed with protein precipitation method, with hyperoside as the internal standard. An Inertsil C8-3 column (2. 1 mm x 50 mm, 5 microm) was adopted, with a mobile phase composed of water (containing 1 mmol L-1 Sodium formate)-acetonitrile (gradient elution) at a flow rate of 0.4 mL . min -1. Electrospray ionization (ESI) was adopted in the positive ion mode for multi-reaction monitoring (MRM). Morroniside showed a good linear relationship ranging between 2-5 000 microg L-1 (r = 0. 995 7), with the minimum limit of quantification of 2 microg L-1. Its precise, accuracy, recovery and matrix effect were all in line with the biological sample measurement requirements. Therefore, the method described above was proved to be suitable for the determination of morroniside concentration in rat plasma. To use the method in the pharmacokinetic study on morroniside in rats, oral administration dose shall be set at 20 mg . kg - to map the plasma concentration-time curve. Main pharmacokinetic parameters were calculated by DAS 2. 0. Specifically, AUC0-inifinity was (587.6 +/- 290. 7) microg min L-1, Cmax was (334.2+/-148.0) microg L-1, Tmax was (0.6 +/-0.3) h, t1/2 was (0.7+/-0.3) h.

Research progress of enhancing quantitative sensitivity by using LC-MS(n) with derivatization method in bio-matrices / 药学学报

Liquid chromatography/mass spectrometry (LC-MS(n)) has been essential to a large number of quantitative analytical applications in drug research, and especially in the drug PK/PD research, due to its high sensitivity and high specificity. But following the appearance of drugs with high activity and low dosage and the especial structural compounds, a number of limitations of LC-MS(n) have been noted. Derivatization changes the structure of drugs and therefore changes their physical and chemical properties, resulting in high ionization efficiency, low matrix effect and low disturbance by inorganic salts and endogenous compounds in LC-MS(n). In this article, recent progress in the research of the chemical derivatization strategy with LC-MS(n) is reviewed on breakthrough of some LC-MS(n) limitations, in particular focusing on the applications involving some drugs in bio-matrices.

Quantification of peramivir in rat plasma by liquid chromatography/tandem mass spectrometry employing precolumn derivatization and its application in pharmacokinetics / 军事医学科学院院刊

Objective:To develop a liquid chromatography/tandem mass spectrometry employing precolumn derivatization method for determination of peramivir in rat plasma.Methods: The sample preparation consisted of a protein precipitation extraction followed by derivatization with hydrochloric acid (10 mol/L) methanol (10∶90, v/v) and determined by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). The positive ion SRM mode was performed and the precursor-to-the-product ion transitions of m/z 343→284 and 299→152 were used to measure the derivative of peramivir and Ro 64-0802 (I.S.). The chromatographic separation was achieved using a Zorbax RX-C8 (2.1 mm × 150 mm × 5 μm) analytical column with an isocratic mobile phase composed of acetonitrile-water-formic acid (30∶70∶0.1, v/v/v, 0.2 ml/min). Results:The method was linear over a concentration range of 10 to 10 000 ng/ml. The LLOQ was 10 ng/ml. The average inter-day/intra-day precision values ranged from 5.0% to 7.1%, respectively, while the accuracy values ranged from 89.9% to 106.1%.Conclusion: In this method, retention time is greatly improved, the matrix effects are decreased by chemical derivatization. This method has been successfully applied to the preclinical and clinical research of peramivir.

Quantitative determination of metoprolol by liquid chromatography mass spectrometry in rat plasma / 军事医学科学院院刊

Objective:To establish a highly sensitive, rapid and selective liquid chromatography mass spectrometry (LC-MS) method for the determination of metoprolol in rat plasma.Methods:A simplified liquid-liquid extraction with acetidin was employed for the sample preparation. The separation was carried out on a Thermo ODS-C_(18)(5 μm,100 mm×2.1 mm).The mobile phase consisted of acetonitrile-methanol-water(20∶20∶60). Propranolol was used as the internal standard. The detection was performed on a liquid chromatography mass spectrometry by selected ion monitoring(SIM) scan mode electrospray ionization(ESI).Results and Conclusion:The range of calibration curve was 0.1-50 ng/ml and the limit of quantification was 0.1 ng/ml. The intra- and inter-day precision RSD was less than 15%.This method is sensitive, simple,rapid and suitable for the pharmacokinetic study of metoprolol.

Determination of decapeptide LXT-101 in plasma by HPLC-MS/MS and its pharmacokinetics in Beagle dogs / 药学学报

This paper developed a sensitive and specific liquid chromatography-electrospray ionization mass spectrometry (HPLC-MS/MS) method for the determination of decapeptide LXT-101 in Beagle dog plasma. Plasma samples spiked with internal standard (IS) were treated with acetonitrile to precipitate the protein. Selected reaction monitoring (SRM) using the precursor --> product ion combinations of m/z 472.1-->587.9 and m/z 502.8-->633.8 were used to quantify LXT-101 and IS, respectively. The linear calibration curves were obtained in the concentration range of 0.5 - 500.0 ng x mL(-1). The limit of quantification (LOQ) was 0.5 ng x mL(-1). The inter-day and intra-day precision (RSD) across three validation run over the entire concentration range was below 10.9%, and the accuracy (RE) was within +/- 1.8%. The main pharmacokinetic parameters of LXT-101 after muscle injection of 20 microg x kg(-1) were as follows, AUC(0-t): (176.8 +/- 116.7) microg x h x L(-1), MRT(0-t): (2.52 +/- 0.53) h, T(1/2): (1.4 +/- 0.3) h; CL: (0.16 +/- 0.09) L x h(-1) x kg(-1), and Vd: (0.30 +/- 0.16) L x kg(-1), respectively. The method is proved to be specific, sensitive and suitable for the investigation of LXT-101 pharmacokinetics in Beagle dog.

Stability studies of sulfated polyguluronic acid ester / 药学学报

<p><b>AIM</b>To test the stability of marine polysaccharide drug sulfated polyguluronic acid ester.</p><p><b>METHODS</b>Four methods including high performance gel chromatography (HPGC), poly-acrylamide gel electrophoresis (PAGE), UV scan of absorbance between 200 and 800 nm and gelatin nephelometry were established. Samples were tested in high temperature, high humidity, strong light and accelerated test conditions. The methods were used to test the changes of the parameters including molecular weight, molecular weight distribution, absorbance between 200 and 800 nm, free sulfate, with which we could estimate the stability of sulfated polyguluronic acid ester could be estimated.</p><p><b>RESULTS</b>The four methods were suitable to test the stability of sulfated polyguluronic acid ester and the sample were stable in the conditions as before except in high temperature.</p><p><b>CONCLUSION</b>Sulfated polyguluronic acid ester has good stability.</p>

Identification of the metabolites of penehyclidine hydrochloride raceme in rats by LC-MS/MS and ion cluster / 药学学报

<p><b>AIM</b>To study the metabolites of penehyclidine hydrochloride (PH) raceme, a new anticholinerigic drug invented by the Institute of Pharmacology and Toxicology, Academy of Military Medical Sciences.</p><p><b>METHODS</b>Three healthy rat urine samples were collected within 24 h after a single i.m. dose of PH raceme and PH-d5 [(5 + 5) mg.kg-1] simultaneously. The eight metabolites of PH raceme were identified by the methods of LC-MS/MS, GC-MS, FAB-MS and the stable isotope ion cluster. Mass spectrometry was operated in the positive mode for the method of LC-MS/MS.</p><p><b>RESULTS</b>M1 and M1* were identified as the oxygenated products of PH in the cyclopentyl group; M2 and M2* were as the hydroxylated products of PH in the cyclopentyl group; M3 and M3* were as the oxygented and hydroxylated products of PH at the meta-position of cyclopentyl group; M4 and M4* were identified as the dihydroxylated metabolites of PH, the hydroxylated position were at the cyclopentyl group and quiniuclidinol ring of PH. Among them, M1 and M1*, M2 and M2*, M3 and M3*, M4 and M4* were the isomers of each other.</p><p><b>CONCLUSION</b>These characteristics can be used for future structure elucidation in studies of the metabolites of PH optical isomers. The structure data of PH metabolites provide important information for the clinical use and for developing better anticholinerigic drug.</p>

RP-HPLC determination of loganin in mouse plasma and its pharmacokinetics / 药学学报

<p><b>AIM</b>To develop a method for determination of loganin in mouse plasma by using high-performance liquid chromatography. The method was employed to study pharmacokinetics of loganin.</p><p><b>METHODS</b>An RP-C18 was used as the stationary phase. The mobile phase consisted of methanol-water (30:70), at the flow-rate of 0.8 mL.min-1. The UV absorbance detector was set at 240 nm. Plasma samples were treated with solid phase extraction.</p><p><b>RESULTS</b>The recovery of loganin in mouse plasma was 86.0%-91.5%. The calibration curve in plasma was linear over the range of 0.01-5.00 micrograms.mL-1. The limit of quantitation was 10 ng.mL-1. The RSDs of intra-day and inter-day (n = 5) were less than 15%. The pharmacokinetic parameters were Cmax = 6.8 micrograms.mL-1, Tmax = 30 min, T1/2 alpha = 26.1 min, T1/2 beta = 29.01 min.</p><p><b>CONCLUSION</b>The method is accurate, sensitive and suitable for pharmcokinetic study of loganin. The absorption and elimination of loganin were rapid after ig in mice.</p>