Bohyoung Lee

The alteration of components in the fermented Hwangryunhaedok-tang and its neuroprotective activity

Background: Hwangryunhaedok-tang is a traditional herbal prescription that has sedative activity, hypotensive and anti-bacterial effects. Objective: In this study, we investigated the alteration of contents of components in Hwangryunhaedok-tang, antioxidant activity and neuroprotective activity by fermentation with Lactobacillus acidophilus KFRI 128. Materials and Methods: Contents of three marker compounds (geniposide, berberine and palmatine) and unknown compounds in the Hwangryunhaedok-tang (HR) and the fermented Hwangryunhaedok-tang (FHR) were measured and compared using the established high-performance liqued chromatograph coupled with a photodiode (HPLC-DAD) method. The antioxidant activity of HR and FHR were determined by DPPH free radical and hydrogen peroxide (H2O2) scavenging assay. Also, the neuroprotective activities of HR and FHR against glutamate-induced oxidative stress in a mouse hippocampal cell line (HT22) were evaluated by MTT assay. Results: The contents of geniposide and palmatine were decreased but the content of berberine was increased in the FHR. And the contents of unknown compounds (1), (2), (3), (4) and (5) in the HR were altered by fermentation. Electron donating activity (EDA, %) value of FHR was higher than HR for DPPH radical scavenging activity and H2O2 scavenging activity, respectively. In the MTT assay, FHR showed more potent neuroprotective activity than HR by 513.90%. Conclusion: The FHR using microorganism could convert compounds in HR and enhance the antioxidant and neuroprotective activity.

Neuroprotective activity of the methanolic extract of Lonicera japonica in glutamate-injured primary rat cortical cells.

Background: We previously reported that the extracts of several Korean medicinal plants showed neuroprotective activity in glutamate-injured primary culutres of rat cortical cells. Objective: Among them, the effect of the methanolic extract of Lonicera japonica flower on the glutamate-induced neuronal cell death and its potential mechanism of action was investigated. Results: Treatment by the methanolic extract of L. japonica flower significantly protected neuronal cells against glutamate-induced excitotoxicity. It decreased the calcium influx that accompanies the glutamate induced excitotoxicity of neuronal cells, and inhibited the subsequent overproduction of nitric oxide, reactive oxygen species and peroxide to the level of control cells. In addition, it preserved cellular activity of superoxide dismutase, an antioxidative enzyme reduced by glutamate insult. Conclusions: According to this data, the methanolic extract of L. japonica flower significantly protected neuronal cells against glutamate excitotoxicity via antioxidative activity.

Simultaneous determination of 11 major components in Palmul-tang by HPLC-DAD and LC-MS-MS.

Palmul-tang, a traditional herbal medicine, is composed of eight herbs (Ginseng radix, Glycyrrhizae radix, Hoelen, Atractylodis rhizoma, Angelicae gigantis radix, Cnidii rhizoma, Paeoniae radix and Rehmanniae radix) and exhibits various bioactivities, including antiallergic and antitumor effects. In this study, an effective, reliable and accurate high-performance liquid chromatography method has been developed for the simultaneous determination of 11 marker components in Palmul-tang: hydroxymethylfurfural, albiflorin, paeoniflorin, ferulic acid, nodakenin, ginsenoside Rg1, decursinol, glycyrrhizin, 6-gingerol, ginsenoside Rg3 and decursin. All calibration curves of the 11 components indicated excellent linearity (correlation coefficient > 0.9997) within the test range. The limits of detection and quantification of each component were in the ranges of 0.08-1.03 and 0.23-3.11 µg/mL, respectively. The intra-day and inter-day relative standard deviation values were within 1.65 and 2.71%, respectively. The mean recovery values were 94.49 to 101.10%. The established method was successfully applied to the simultaneous determination of 11 major components in 12 commercial samples of Palmul-tang. The developed analytical method is simple and suitable for the quality control of Palmul-tang.

Qualitative and quantitative simultaneous determination of six marker compounds in soshiho-tang by HPLC-DAD-ESI-MS

Soshiho-tang, one of the herbal prescriptions, has been used in treatment of chronic liver disease. In this study, the efficient high-performance liquid chromatography coupled with diode array detection and electrospray ionization tandem mass spectrometry (HPLC-DAD-ESI-MS) method was developed and validated for simultaneous determination of six marker compounds, namely homogentisic acid, baicalin, glycyrrhizin, saikosaponin A, 6-gingerol and ginsenoside Rg3 in Soshiho-tang. The analysis of six marker compounds was carried out using a C18 column (SHISEIDO S-5 μm, 4.6 mm I.D. × 250 mm) and gradient elution with water − 0.1% trifluoroacetic acid and acetonitrile. The analytical method was validated for linearity, precision and accuracy. Calibration curve for six marker compounds showed good linear regression (r2 > 0.9996). The limits of detection (LOD) and limits of quantification (LOQ) for analytes were ranged from 0.04 to 0.16 μg/mL and 0.12 to 0.47 μg/mL, respectively. The relative standard deviations (RSD%) of the precision test, intra- and inter-day, were less than 0.32% and 0.10%, respectively. All recoveries of the six marker compounds were ranged from 100.05% to 102.25% with RSD less than 0.63%. These results have shown that this developed method was efficient for the quality evaluation of Soshiho-tang.

Neuroprotective and cognitive enhancing activity of the fermented Bozhougyiqi-Tang

Background: Alzheimers disease is a neurodegenerative disease related to memory impairments and neuronal cell death. Bozhougyiqi-Tang (BZYQT), a traditional herbal medicine, has been therapeutically used for the treatment of pulmonary tuberculosis. Objective: The aim of this study is to evaluated the neuroprotective effect of the fermented BZYQT and compared with unfermented BZYQT in HT22 cells by MTT assay and tested the beneficial effect on memory impairments induced by scopolamine (1 mg/kg, i.p.) using the passive avoidance and Morris water maze tests. Results: Compared with unfermented BZYQT, the neuroprotective effect of fermented BZYQT on glutamate induced neurotoxicity in HT22 cells increased at a concentration of 100 μg/mL. Fermented BZYQT increased the step-through latency of the passive avoidance response. Furthermore, in Morris water maze test for evaluation of spatial learning and memory, escape latency time was significantly reduced by fermented BZYQT. Conclusion: These results suggest that the fermentation process of BZYQT led to improve neuroprotective and cognitive enhancing effect.

SIMULTANEOUS QUANTIFICATION OF ELEVEN CHEMICAL COMPONENTS IN TRADITIONAL HERBAL MEDICINAL FORMULA SOCHEONGRYONGTANG BY HPLC-DAD AND LC-MS

Sochengryongtang (SCRT), a traditional herbal medicine, has been used for the treatment of cold for thousands of years. In this study, high performance liquid chromatography with diode array detector (HPLC-DAD) and electrospray ionization mass spectrometry (ESI-MS) was applied for the reliable and sensitive simultaneous analysis of 11 major components (homogentisic acid, ephedrine HCl, paeoniflorin, cinnamic acid, cinnamaldehyde, glycyrrhizin, 6-gingerol, schisandrin, methyl engenol, gomisin A, and gomisin N) in SCRT. Components were separated on a C18 column using gradient elution with 0.1% TFA water and acetonitrile and characterized based on the retention time in HPLC chromatogram, UV spectrum, and MS data. All calibration curves of major components proved to have good linearity (r2 > 0.9999). The LOQ and LOD ranged from 7.7 to 47.8 ng/mL and 23.5 to 131.2 ng/mL, respectively. This developed method showed good precision and accuracy with intra-day and inter-day (relative standard deviation, R.S.D.) values were less than 1.93% and 1.74%, respectively. The overall recoveries ranged from 92.42% to 107.87% with the R.S.D. ranging from 0.01% to 1.87%. The established method was successfully applied for the simultaneous determination of 11 components in SCRT.

Simultaneous determination of nine bioactive compounds in Yijin-tang via high-performance liquid chromatography and liquid chromatography-electrospray ionization-mass spectrometry

Background Yijin-tang (YJ) has been used traditionally for the treatment of cardiovascular conditions, nausea, vomiting, gastroduodenal ulcers, and chronic gastritis. In this study, a simple and sensitive high-performance liquid chromatography (HPLC) method was developed for the quantitation of nine bioactive compounds in YJ: homogentisic acid, liquiritin, naringin, hesperidin, neohesperidin, liquiritigenin, glycyrrhizin, 6-gingerol, and pachymic acid. Methods Chromatographic separation of the analytes was achieved on an RS Tech C18 column (4.6 mm × 250 mm, 5 μm) using a mobile phase composed of water containing 0.1% (v/v) trifluoroacetic acid (TFA) and acetonitrile with a gradient elution at a flow rate of 1.0 mL/min. Results Calibration curves for all analytes showed good linearity (R2 ≥ 0.9995). Lower limits of detection and lower limits of quantification were in the ranges of 0.03–0.17 μg/mL and 0.09–0.43 μg/mL, respectively. Relative standard deviations (RSDs; %) for intra- and interday assays were < 3%. The recovery of components ranged from 98.09% to 103.78%, with RSDs (%) values ranging from 0.10% to 2.59%. Conclusion This validated HPLC method was applied to qualitative and quantitative analyses of nine bioactive compounds in YJ and fermented YJ, and may be a useful tool for the quality control of YJ.

Quantitative analysis of the eight major compounds in the Samsoeum using a high-performance liquid chromatography coupled with diode array detection and electrospray ionization mass spectrometer

Background: Samsoeum was traditionally used for treatment of a respiratory disease. Objective: The simultaneous determination of eight major compounds, ginsenoside Rg3, caffeic acid, puerarin, costunolide, hesperidin, naringin, glycyrrhizin, and 6-gingerol in the Samsoeum using a high-performance liquid chromatography (HPLC) coupled with diode array detection (DAD) and an electrospray ionization mass spectrometer was developed for an accurate and reliable quality assessment. Materials and Methods: Eight compounds were qualitative identified based on their mass spectra and by comparing with standard compounds and quantitative analyzed by HPLC-DAD. Separation of eight compounds was carried out on a LUNA C18 column (S-5 μm, 4.6 mm i.d. ×250 mm) with gradient elution composed of acetonitrile and 0.1% trifluoroacetic acid. Results: The data showed good linearity (R2 > 0.9996). The limits of detection and the limits of quantification were <0.53 μg and 1.62 μg, respectively. Inter- and Intra-day precisions (expressed as relative standard deviation values) were within 1.94% and 1.91%, respectively. The recovery of the method was in the range of 94.24–107.90%. Conclusion: The established method is effective and could be applied to quality control of Samsoeum.

Neuroprotective effects of 4,5-dimethoxypyrocatechol isolated from Cynanchum paniculatum on HT22 cells.

Background: Cynanchum paniculatum, belongs to the family Asclepiadaceae and is used to treat various diseases, such as invigorate blood, alleviate edema and to relieve pain and toxicity for a long time. Materials and Methods: 4,5-Dimethoxypyrocatechol was isolated from the 80% methanol extract of C. paniculatum and its neuroprotective effect was evaluated by MTT assay. Results: 4,5-Dimethoxypyrocatechol had neuroprotective effect on the glutamate-induced cellular oxidative death in HT22 cells. Conclusion: Furthermore, we found that reactive oxygen species (ROS) accumulation and calcium concentration by oxidative stress were reduced by 4,5-dimethoxypyrocatechol in HT22 cells.

Simultaneous determination of ten bioactive compaounds from the roots of Cynanchum paniculatum by using high performance liquid chromatography coupled-diode array detector

Background: Cynanchum paniculatum Kitagawa belongs to Asclepiadaceae and was used in traditional medicine to invigorate blood, alleviate edema, relieve pain, and relieve toxicity for a long time. Objective: A novel and reliable high performance liquid chromatography coupled with diode array detector method has been established for simultaneous determination of 10 bioactive compounds isolated from Cynanchum paniculatum Kitagawa, one of the herbal medicines. Materials and Methods: The chromatography analysis was performed on a SHISEIDO C18 column (S-5 μm, 4.6 mm I.D. × 250 mm) at 35°C with a gradient elution of acetonitrile and water at a flow rate of 1ml/min and UV detection at 210, 230, and 280 nm. Results: The method was validated for linearity, precision, and accuracy. All calibration curves showed good linear regression (r2 > 0.9996). Limits of detection (LOD) and limits of quantification (LOQ) fell in the ranges 0.01 - 0.28 μg/ml and 0.04 - 0.83 μg/ml, respectively. The relative standard deviation (RSD) of the intra- and inter- day test, precision test were within 1.92% and 2.43%, respectively. The mean recovery of all ranged from 92.82 to 107.96% with RSD values 0.12 - 2.18%. Conclusion: The results of validation appeared that this established method was very accurate and stabilized for the quantification of 10 bioactive compounds isolated from C. paniculatum.