Compositional Characteristics of Glucuronide Conjugates in Regional Glechoma hederacea var. longituba Herbal Extracts Using a Set of Polyphenolic Marker Compounds.

Glechoma hederacea var. longituba (GHL) is one of many herbal plants widely used in hot herbal teas and in oriental prescriptions to treat various diseases. Although the beneficial effects of GHL may be influenced by differences in the composition of active constituents in the herbal extracts, there are few reports on the compositional characteristics of GHL herbal extracts to date. In this study, liquid chromatography-mass spectrometry technology was used for comparative analysis of constituents in hot-water extracts of GHL samples obtained from various cultivating provinces in South Korea. A set of marker panel consisting of nine polyphenolic compounds, including glucuronide conjugates in particular, was constructed and used to monitor the compositional characteristics in each GHL extract. Our findings show that some of the marker compounds, including rosmarinic acid, were persistently observed as major constituents in the analyses of the 22 GHL sample extracts, whereas, interestingly, other marker compounds such as polyphenol-glucuronic acid conjugates displayed dramatic differences in compositional ratios. This chromatographic approach using the marker compound panel can be applied to qualitatively and quantitatively evaluate compositional characteristics in the GHL extracts, and can also be useful for quality assays of the GHL herbal plant in medicinal and industrial fields.

Identification of Polyphenol Glucuronide Conjugates in Glechoma hederacea var. longituba Hot Water Extracts by High-Performance Liquid Chromatography-Tandem Mass Spectrometry (HPLC-MS/MS).

Glechoma hederacea var. longituba (GHL) is one of many herbal plants distributed worldwide and is known to contain various biologically useful antioxidant constituents. GHL has been used in folk remedies for various treatments and as favorable tea beverages. However, research on the precise analysis of ingredients in GHL extracts remains insufficient. In this study, compositional analysis has been conducted on polyphenolic ingredients in GHL hot water extracts. GHL samples collected from growing regions were incubated in hot water at 100 °C for 1 h. The polyphenolic constituents in the hot water extracts were analyzed using high performance liquid chromatography-high resolution mass spectrometry (HPLC-HR MS) and tandem mass spectrometry (HPLC-MS/MS) in negative ion mode. As a result, a total of seven compounds were identified as the major polyphenolic constituents. Interestingly, four constituents out of the identified substances were confirmed to be polyphenol glucuronide conjugates, in which glucuronidation was known to be an important metabolic process in polyphenol aglycone along with methylation and sulphation. This study can be applied for the quality control and standardization of GHL herbal samples and the monitoring of metabolic processes involved in the polyphenolic conjugates.

Proteome analysis of hairy root from Panax ginseng C.A. Meyer using peptide fingerprinting, internal sequencing and expressed sequence tag data.

As an initial step to the comprehensive proteomic analysis of Panax ginseng C. A. Meyer, protein mixtures extracted from the cultured hairy root of Panax ginseng were separated by two-dimensional polyacrylamide gel electrophoresis (2-DE). The protein spots were analyzed and identified by peptide finger printing and internal amino acid sequencing by matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) and electrospray ionization quadrupole-time of flight mass spectrometry (ESI Q-TOF MS), respectively. More than 300 protein spots were detected on silver stained two-dimensional (2-D) gels using pH 3-10, 4-7, and 4.5-5.5 gradients. Major protein spots (159) were analyzed by peptide fingerprinting or de novo sequencing and the functions of 91 of these proteins were identified. Protein identification was achieved using the expressed sequence tag (EST) database from Panax ginseng and the protein database of plants like Arabidopsis thaliana and Oryza sativa. However, peptide mass fingerprinting by MALDI-TOF MS alone was insufficient for protein identification because of the lack of a genome database for Panax ginseng. Only 17 of the 159 protein spots were verified by peptide mass fingerprinting using MALDI-TOF MS whereas 87 out of 102 protein spots, which included 13 of the 17 proteins identified by MALDI-TOF MS, were identified by internal amino acid sequencing using tandem mass spectrometry analysis by ESI Q-TOF MS. When the internal amino acid sequences were used as identification markers, the identification rate exceeded 85.3%, suggesting that a combination of internal sequencing and EST data analysis was an efficient identification method for proteome analysis of plants having incomplete genome data like ginseng. The 2-D patterns of the main root and leaves of Panax ginseng differed from that of the cultured hairy root, suggesting that some proteins are exclusively expressed by different tissues for specific cellular functions. Proteome analysis will undoubtedly be helpful for understanding the physiology of Panax ginseng.