Comparative analysis of chloroplast genomes of Pulsatilla species reveals evolutionary and taxonomic status of newly discovered endangered species Pulsatilla saxatilis.

BACKGROUND: Pulsatilla saxatilis, a new species of the genus Pulsatilla has been discovered. The morphological information of this species has been well described, but its chloroplast genome characteristics and comparison with species of the same genus remain to be reported. RESULTS: Our results showed that the total length of chloroplast (cp.) genome of P. saxatilis is 162,659 bp, with a GC content of 37.5%. The cp. genome contains 134 genes, including 90 known protein-coding genes, 36 tRNA genes, and 8 rRNA genes. P. saxatilis demonstrated similar characteristics to other species of genus Pulsatilla. Herein, we compared cp. genomes of 10 species, including P. saxatilis, and found that the cp. genomes of the genus Pulsatilla are extremely similar, with a length of 162,322-163,851 bp. Furthermore, The SSRs of Pulsatilla ranged from 10 to 22 bp in length. Among the four structural regions of the cp. genome, most long repeats and SSRs were detected in the LSC region, followed by that in the SSC region, and least in IRA/ IRB regions. The most common types of long repeats were forward and palindromic repeats, followed by reverse repeats, and only a few complementary repeats were found in 10 cp. genomes. We also analyzed nucleotide diversity and identified ccsA_ndhD, rps16_trnK-UUU, ccsA, and rbcL, which could be used as potential molecular markers for identification of Pulsatilla species. The results of the phylogenetic tree constructed by connecting the sequences of high variation regions were consistent with those of the cp. gene phylogenetic tree, and the species more closely related to P. saxatilis was identified as the P. campanella. CONCLUSION: It was determined that the closest species to P. saxatilis is P. campanella, which is the same as the conclusion based on pollen grain characteristics, but different from the P. chinensis determined based on morphological characteristics. By revealing information on the chloroplast characteristics, development, and evolution of the cp. genome and the potential molecular markers, this study provides effective molecular data regarding the evolution, genetic diversity, and species identification of the genus Pulsatilla.

Two novel ketone alkaloids from Portulaca oleraceaL. and their anti-inflammatory activities.

Eleven compounds were obtained from Portulaca oleracea L., including two novel ketone alkaloids, (1, 2), 4-hydroxy-3-methoxybenzamide (3) (isolated for the first time), ß-adenosine (4), oleracrylimide A and B (5, 6), oleracein H, C, D, Q and A (7-11). The two novel ketone alkaloids were identified as 5-acetyl-5-methylcyclopent-2-ene-1-carboxamide (1), named oleraciamide H, and (2 R,3S,4R,5R)-5-((R)-1,2-dihydroxyethyl)-3,4-dihydroxytetrahydrofuran-2-yl glycinate (2), named oleracone Q by spectroscopic methods, including 1D, 2D NMR and compound fingerprints. Additionally, their anti-inflammatory activities were tested via RAW 264.7 cells induced by LPS and found that they could significantly inhibit the release of IL-1ß and TNF-α.

Four alkaloids from Portulaca oleracea L. and their anti-inflammatory.

Four alkaloids were isolated from Portulaca oleracea L., including two new compounds, 2-(4-amino-6-hydroxy-1,6-dihydropyrimidin-5-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol, named Olerapyrimidine (1) and (2 R,3S,4S,5R,6S)-2-(hydroxymethyl)-6-((6-hydroxypyridin-3-yl)oxy)tetrahydro-2H-pyran-3,4,5-triol, named Olerapyridine (2), and two known compounds including 1H-imidazole (3) and (5S, 6 R, 7S, 8 R)-5-amino-(2Z,4Z)-1,2,3-trihydroxybuta-2,4-dienyloxy-pentane-6,7,8,9-tetraol (4) from P. oleracea for the first time. Their structures were determined by spectroscopic methods, including UHPLC-ESI-Q-TOF/MS, 1D and 2D NMR spectra. Both Olerapyrimidine and Olerapyridine at 20 µM could inhibit the inflammatory factors, IL-1ß and TNF-α in the RAW 264.7 cells induced by LPS.

Two novel alkaloids from Portulaca oleracea L. and their anti-inflammatory bioactivities.

Two novel alkaloids were identified as (E)-N-(4-3,4-dihydroxy-6-(hydroxymethyl)-5-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-2,5-dihydroxyphenyl)-3-(4-hydroxyphenyl)acrylamide (1), named Oleracrylimide D, (E)-N-(4-3,4-dihydroxy-6-(hydroxymethyl)-5-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-2,5-dihydroxyphenyl)-3-(4-hydroxy-3-methoxyphenyl)acrylamide (2), named Oleracrylimide E, isolated from Portulaca oleracea L. The structures were identified by spectroscopic methods, including 1D NMR, 2D NMR, and UHPLC-ESI-QTOF/MS methods, also, the anti-inflammatory bioactivities of the compounds were studied by ELISA method.

A new alkaloid and two organic acids from Portulaca oleracea L. and their bioactivities.

A new alkaloid, identified as 1-benzyl-2-nitroso-1,2,3,4-tetrahydroisoquinoline-6,7-diol, named oleraisoquinoline (1), and five organic acids and two esters, identified as 5-(hydroxymethyl)furan-2-carboxylic acid (2), 1H-pyrrole-2,5-dicarboxylic acid (3), (7E,10E)-octadeca-7,10-dienoic acid (4), (10E,13E)-octadeca-10,13-dienoic acid (5), (7E,10E)-hexadeca-7,10-dienoic acid (6), methyl tridecanoate (7) and methyl (9E,12E)-octadeca-9,12-dienoate (8), were isolated from Portulaca oleracea L., among which compounds 2 and 4‒7 were isolated for the first time. Moreover, the anti-inflammatory activities of compounds 1‒3 were studied, especially, compound 1 presented good inhibitory effects on the production of inflammatory factors IL-1ß and TNF-α.

Two new alkaloids from Portulaca oleracea L. and their anti-inflammatory and anticholinesterase activities.

Two new alkaloids identified as 2-(((S,Z)-1-(1H-azirin-1-yl)-5-methylhex-1-en-3-yl)oxy)-6-(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol and (S,Z)-1-(1H-azirin-1-yl)-5-methylhex-1-en-3-ol, named olerazirin A (1), olerazirin B (2), together with five known alkaloids, identified as cyclo (L-Val-L-Ala) (3), cyclo-(glycyl-L-leucine) (4), cyclo-(Gly-Phe) (5), cyclo (Ser-Phe) (6), (3S,6S)-3-[(1R)-1-hydroxyethyl]-6-(phenyl-methyl)-2,5-piperazinedione (7) were obtained from Portulaca oleracea L. using a range of chromatographic techniques, 1D and 2D NMR, and high-resolution electrospray ionisation time-of-flight mass spectroscopic methods, in which the compounds 3-7 were isolated from P. oleracea for the first time. In addition, the results showed that the compounds 1 and 2 have anti-inflammatory activities and compounds 1-3 and 5-7 exhibit the anticholinesterase activities.

Two novel carbon skeleton compounds from Portulaca oleracea L. and their effects on nitric oxide.

Two new carbon skeleton compounds, identified as 2-(1-(3-hydroxy-2,2,6-trimethyl-6-vinyltetrahydro-2H-pyran-3-yl)-2-methylpropoxy)-6-(hydroxymethyl)tetrahydro-2H-pyran-3,4,5-triol named Olerapyran A (1), and (E)-3-(6-acetyl-2-methyl-5-((3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)cyclooctylidene)butan-2-one named Oleraoctyl (2), were first isolated from Portulaca oleracea L., then their structures were determined using spectroscopic methods, including UHPLC-ESI-QTOF/MS, 1D and 2D NMR. In addition, the activities of Olerapyran A and Oleraoctyl inhibiting nitric oxide (NO) were studied.

Pharmacokinetics of oleracimine in rats by ultra-high-performance liquid chromatography

The novel alkaloid, oleracimine, presented remarkable anti-inflammatory bioactivity, and therefore, its pharmacokinetics was investigated in rat plasma after intravenous and oral administration by using a rapid ultra-high-performance liquid chromatography (UHPLC) method with UV detection at 270 nm. The analysis was performed on a shim-pack ODS column (75 mm×2 mm, 1.6 µm particle size, Shimadzu, Japan) column using isocratic elution with a mobile phase consisting of methanol-water (62:38, v/v) within 3 min. The results indicated that oleracimine was rapidly distributed with Tmax for 11.7 min after oral administration, which presented the double-peak phenomenon in the pharmacokinetic profile with a higher oral absolute bioavailability of 55.1% ± 7.83%.

Tissue distribution and excretion of the five components of Portulaca oleracea L. extract in rat assessed by UHPLC

The aim of the present study was to investigate the tissue distribution and excretion of five components of Portulaca oleracea L. extract (POE) in rat following oral administration. A rapid, sensitive and specific ultra-high performance liquid chromatography (UHPLC) method with puerarin as the internal standard was used for the quantitative analysis of five components of POE, including caffeic acid (CA), p-coumaric acid (p-CA), ferulic acid (FA), quercitrin (QUER) and hesperidin (HP) in rat tissues including the liver, intestine, stomach, muscle, heart, lung, brain, kidney and spleen, urine and feces. The results show that onlyp-CA and FA were found in nearly all tissues with low cumulative ratios, and CA was higher in the intestine and stomach with a slightly higher cumulative ratio in the urine and feces after 24 h. HP and QUER were found at low levels in the tissues with low cumulative ratios.

O objetivo do presente estudo foi investigar a distribuição tecidual e excreção de cinco componentes de extrato Portulaca oleracea L. (POE) em ratos após administração oral. Um método analítico rápido, sensível e específico para quantificação de cinco componentes de POE (ácido cafeico (CA), ácidop-cumárico (p-CA), ácido ferúlico (FA), quercitrina (QUER) e hesperidina (HP)) por cromatografia líquida de ultra eficiência (UHPLC), empregando puerarina como padrão interno de referência. Os compostos foram quantificados em diferentes tecidos dos animais, sendo eles fígado, intestino, estômago, músculo, coração, pulmão, cérebro, rim e baço, urina e fezes. Os resultados mostraram que apenas p-CA e FA foram encontradas em todos os tecidos com baixas taxas cumulativas e CA apresentou níveis mais altos no intestino e estômago com a taxa cumulativa um pouco mais elevada na urina e nas fezes após 24 h. HP e QUER apresentaram baixas concentrações nos tecidos com baixas taxas cumulativas.

Pharmacokinetic study of isoquercitrin in rat plasma after intravenous administration at three different doses

The aim of this study is to develop a simple and specific HPLC method using vitexin as the internal standard to investigate the pharmacokinetics of isoquercitrin (ISOQ) after three different doses administrated intravenously to rats. The pharmacokinetic parameters were calculated by both compartmental and non-compartmental approaches. The results showed that ISOQ fitted a three-compartment open model. The values of AUC increased proportionally within the range of 5-10 mg·kg-1. Moreover, a half-life, b half-life, ªCL, MRT0-t and MRT0→∞ of ISOQ in rats showed significant differences between 20 mg·kg-1 and other doses, indicating that ISOQ presented dose-dependent pharmacokinetics in the range of 5-10 mg·kg-1 and non-linear pharmacokinetics at higher doses.

O objetivo deste estudo é desenvolver um método simples e específico de HPLC usando vitexina como padrão interno para investigar a farmacocinética do isoquercitrina (ISOQ) após três doses diferentes administradas por via intravenosa a ratos. Os parâmetros farmacocinéticos foram calculados pelas abordagens compartimental e não compartimental. Os resultados mostraram que ISOQ se encaixa no modelo de três compartimentos. Os valores de AUC aumentaram proporcionalmente na faixa de 5-10 mg·kg-1. Além disso, a meia-vida, b meia-vida, ªCL, MRT0-t and MRT0→∞ de ISOQ em ratos mostraram diferenças significativas entre 20 mg·kg-1 e outras doses, o que significa que ISOQ apresenta farmacocinética dose-dependente no intervalo de 5-10 mg·kg-1 e farmacocinética não linear em doses mais elevadas.

HPLC method for the simultaneous determination of four compounds in rat plasma after intravenous administration of Portulaca oleracea L. extract

The objective of the present study was to develop a simple and selective HPLC method for the simultaneous determination of hesperidin (HP), caffeic acid (CA), ferulic acid (FA) and p-coumaric acid (p-CA) in rat plasma after intravenous administration of Portulaca oleracea L. extract (POE). With the hyperoside as the internal standard, the sample pretreatment procedure involved simple single-step extraction with methanol of 0.2 mL plasma. The mobile phase consisted of methanol-acetonitrile-tetrahydrofuran-0.5% glacial acetic acid (5:3:18:74, v/v/v/v). The calibration curves were linear over the range of 0.1-25 µg mL-1, 0.1-25 µg mL-1, 0.1-25 µg mL-1and 0.015-3 µg mL-1 for HP, CA, FA and p-CA, respectively. The method developed was suitable for the pharmacokinetic study of HP, CA, FA and p-CA in rats after intravenous administration of POE.

O objetivo do estudo foi desenvolver um método simples e específico de HPLC para a determinação simultânea de hesperidina (HP), ácido caféico (CA), ácido ferúlico (FA) e ácido p-cumárico (p-CA) em plasma de rato após a administração intravenosa de extrato Portulaca oleracea L. (POE) empregando hyperosídeo como padrão interno de referência. Metanol foi empregado para os analitos em plasma (0,2 mL). A fase móvel isocrática foi composta por metanol-acetonitrila-tetraidrofurano-0,5% ácido acético glacial (5:3:18:74, v/v/v/v). Curvas de calibração foram lineares na faixa de concentração de 0,1-25 µg mL-1, 0,1-25 µg mL-1, 0,1-25 µg mL-1 e 0,015-3 µg mL-1 para HP, CA, FA e p-CA, respectivamente. O método desenvolvido foi adequado para estudo farmacocinético de HP, CA, FA e p-CA em ratos após a administração intravenosa de POE.