Chemical Constituents from the Seeds of Lepidium apetalum Willd / 中国药学杂志

OBJECTIVE: To investigate the chemical constituents of the seeds of Lepidium apetalum Willd. METHODS: The compounds were isolated and purified by Diaion HP-20, Toyopearl HW-40, MCI Gel CHP-20, ODS, silica gel chromatography combined with Pre-HPLC and the structures were identified on the basis of spectral data and physiochemical properties. RESULTS: Sixteen compounds were isolated and identified from the water extract as catechol(1), protocatechuic aldehyde(2), 2-phenyl acetamide(3), methyl-5-hydroxypyridine-2-carboxlate(4), benzylcarbamic acid(5), N-benzylacetamide(6), raphanuside C(7), 1-phenyl-1, 2-ethanediol(8), 2-(4-hydroxyphenyl)-ethanol(9), isorhamnetin-7-O-α-L-rhamnopyranoside(10), kaempferol(11), methyl 2, 4, 6-trihydroxybenzoate(12), 2-(4-hydroxyphenyl)acetonitrile(13), syringic acid(14), protocatechuic acid(15), and methyl sinapate(16). CONCLUSION: Compounds 1-16 are isolated from this plant for the first time.

Lignans from seeds of Lepidium apetalum / 中草药

Objective To study the chemical constituents from the seeds of Lepidium apetalum. Methods Compounds were isolated from the water extract from the seeds of L. apetalum by using Diaion HP-20, Toyopearl HW-40, MCI Gel CHP-20, ODS, Silica gel chromatography and semi-preparative-HPLC. Results Nine compounds were isolated and identified as lepidiumlignan A (1), lepidiumlignan B (2), erythro-1-(4-O-β-D-glucopyranosyl-3-methoxyphenyl)-2-[4-(3-hydroxypropyl)-2,6-dimethoxyphenoxy]-1,3- propanediol (3), (7R,7’E,8S)-4,9-dihydroxy-3,3’,5-trimethoxy-4’,7-epoxy-8,5’-neolign-7’-en-9’-oic acid (4), spicatolignan B (5), (-)-pinoresinol-4-O-β-D-glucopyranoside (6), (-)-isolariciresinol (7), aegineoside (8), and (+)-syringaresinol-O-β-D-glucopyra- noside (9). Conclusion Compounds 1 and 2 are new compounds, named as lepidiumlignan A and lepidiumlignan B. Compounds 3-9 are isolated from the plants for the first time.

Cloning, sequence analysis, and prokaryotic expression of LaF3H gene from Lepidium apetalum / 中草药

Objective In order to study the key genes involved in flavonoid biosynthesis pathway, the flavanone-3-hydroxylase (F3H) gene was isolated from Lepidium apetalum, which is named as LaF3H. Meanwhile, the sequence analysis, prokaryotic expression, and purification were also performed. Methods Specific primers were designed according to LaF3H gene sequences in the transcriptome data of L. apetalum, and the cDNA sequence of LaF3H gene was isolated from L. apetalum. By construction the prokaryotic expression vector pET-32a-LaF3H, the recombinant LaF3H protein was expressed in Escherichia coli BL21 (DE3) cells under IPTG induction. Results The open reading frame (ORF) of LaF3H was 1 080 bp, which encoded a protein of 359 amino acid residues, with a predicted molecular mass of 40 320. Sequence analysis showed that LaF3H contains five conserved motifs of F3H protein. The phylogenetic analysis indicated that LaF3H protein showed the highest homology with F3H protein from cruciferous plants (such as AtF3H from Arabidopsis thaliana). The prokaryotic expression vector pET-32a-LaF3H was constructed and the recombinant LaF3H protein was successfully expressed in E. coli BL21 (DE3) cells. Furthermore, the recombinant LaF3H protein was purified through Ni2+ affinity chromatography. Conclusion The LaF3H gene was isolated from L. apetalum and the recombinant LaF3H protein was obtained. The results of this study provided the foundation for the further preparation of LaF3H antibody and detection of LaF3H enzyme activity, and were helpful for functional characterization of LaF3H gene involved in flavonoid biosynthesis pathway of L. apetalum.

Cloning, bioinformatic analysis, and prokaryotic expression of LaPMK gene in Lepidium apetalum / 中草药

Objective: To obtain the key enzyme gene involved in terpenoid biosynthesis pathway, phosphomevalonate kinase (PMK) gene was cloned from Lepidium apetalum, and sequence analysis and prokaryotic expression were performed. Methods: Based on the transcriptome data of L. apetalum, by designing specific primers of LaPMK gene, an open reading frame (ORF) of LaPMK gene was isolated from L. apetalum. Escherichia coli BL21 (DE3) cells were transformed with the prokaryotic expression vector pET32a-LaPMK and used for prokaryotic expression under IPTG induction. Results: LaPMK gene has ORF of 1 518 bp (GenBank accession number KT004541), which encoded a protein of 505 amino acid residues. Bioinformatic analysis indicated that LaPMK protein which located in cytoplasm had no transmembrane domain and signal peptide, and exhibited the specific N-terminal domain and C-terminal domain of GHMP kinase super family. Phylogenetic analysis indicated that LaPMK protein showed the highest homology, 92% similarity, with PMK protein from Brassica rapa. The recombinant LaPMK protein was successfully expressed in E. coli BL21 (DE3) cells. Conclusion: The LaPMK gene is cloned from L. apetalum, and the stable prokaryotic expression system of pET32a-LaPMK is constructed. This study will provide the fundamental information for the further purification and the antibody preparation of LaPMK protein be helpful for the functional researches of LaPMK gene in terpenoid biosynthesis pathway.

A new phenylacetamide from the seeds of Lepidium apetalum Willd / 药学学报

The chemical constituents of the seeds of Lepidium apetalum Willd. were investigated using chromatographic methods, including Diaion HP-20, Toyopearl HW-40, MCI Gel CHP-20, ODS, silica gel chromatography and semi-preparative-HPLC. Three compounds were isolated and their structures were elucidated by spectral data and physicochemical properties, which were identified as lepidiumamide A (1), cis-desulfoglucotropaeolin (2), trans-desulfoglucotropaeolin (3). Among those, compound 1 is a new phenylacetamide, compound 2 and 3 were isolated from this plant for the first time, and their configurations were also identified for the first time.