• 대한본초학회지
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• 제33권6호
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• pp.29-34
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• 2018

Objectives : It is necessary to manage herbal medicines based on effectiveness by comparing the efficacy of herbal medicines by region. In this study, we compared anti-oxidative activity and marker compounds of Eucommiae Cortex by regional groups. Methods : Eucommiae Cortex grown and harvested in Gangjin, Sancheong, Yeongwol, Jangsu, and Jecheon were used. Eucommiae Cortex was extracted in distilled water at $100^{\circ}C$ for 3 hours, and filtered. filtered. Extract samples were freeze-dried at $-80^{\circ}C$. Comparison of anti-oxidant activity in Eucommiae Cortex samples from regional groups was measured in 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging, 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) free radical scavenging, and ferric reducing antioxidant power (FRAP) between regional groups of Eucommiae Cortex. In addition, high-performance liquid chromatography (HPLC) analysis was conducted to compare pinoresinol diglucoside concentration by regional groups. Results : HPLC analysis found that pinoresinol diglucoside concentration, widely known as the marker compound of Eucommiae Cortex, was the highest in Gangwon Yeongwol. There was a significant difference in anti-oxidative activity of Eucommiae Cortex between regional groups as discovered in DPPH, ABTS and FRAP assays. DPPH free radical scavenging was the highest in Jeonbuk Jangsu. ABTS free radical scavenging was the highest in Jeonbuk Jangsu and Chungbuck Jecheon. FRAP was the highest in Jeonbuk Jangsu. Conclusions : Although pinoresinol diglucoside concentration was high, anti-oxidative activity was not proportionately high. Pinoresinol diglucoside concentration was the highest in Gangwon Yeongwol. Anti-oxidative activity was the highest in Jeonbuk Jangsu.

• 한방재활의학과학회지
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• 제30권2호
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• pp.67-75
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• 2020

Objectives SJ004 is a natural herbal medicine that contains Acyranthes japonica Nakai and Eucommia ulmoides Oliver traditionally used for joint and spinal diseases. This study aimed to establish an efficient method of extracting SJ004 to standardize using the yield, high-performance liquid chromatography (HPLC), and antioxidant assay. Methods SJ004 was extracted with distilled water, 70% and 100% of ethyl alcohol (EtOH). The method validation of 20-hydroxyecdysone and pinoresinol diglucoside was determined by HPLC-photo diode array and the content of SJ004 was calculated. The antioxidant activity of each extract was compared and measured using total flavonoids, total phenolic compounds, 2,2-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid), and ferric reducing antioxidant power according to the standard protocol. Results The yield was highest in pure water extract and lowest in 100% EtOH. But, the content of marker compounds indicating 20-hydroxyecdysone and pinoresinol diglucoside was highest in 100% EtOH extract. In the physiological activity measurement using antioxidant activity, 100% ethanol extract was highest. The limit of detection indicating 20-hydroxyecdysone and pinoresinol diglucoside were analyzed 0.33 ㎍/mL, 0.1616 ㎍/mL, and the limit of quantification were analyzed 1.01 ㎍/mL and 0.49 ㎍/mL respectively. Conclusions The experimental results showed that the extraction conditions have a significant effect on content of marker compounds and antioxidant activity. As a result of method validation, SJ004 was standardized by 20-hydroxyecdysone and pinoresinol diglucoside.

• 생약학회지
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• 제46권2호
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• pp.123-132
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• 2015

In this study, we carried out quantification analysis of the six marker components, geniposidic acid, chlorogenic acid, geniposide, pinoresinol diglucoside, liriodendrin, and genipin in the 70% ethanol extracts of non-processed Eucommiae Cortex and processed Eucommiae Cortex using a high-performance liquid chromatography coupled with photodiode array detector. The six components were separated on Gemini C₁₈ column (5 μm, 4.6×250 mm) by the gradient elution with 1.0% (v/v) acetic acid in water and 1.0% (v/v) acetic acid in acetonitrile as mobile phase. The flow rate was 1.0 mL/min and the injection volume was 10 mL. The amount of geniposidic acid, chlorogenic acid, geniposide, pinoresinol diglucoside, liriodendrin, and genipin in non-processed Eucommiae Cortex were 1.31, 0.31, 0.66, 0.46, 0.46, and 0.03%, respectively, while the amount of the six compounds in non-processed Eucommiae Cortex were 0.04-0.78, 0.01-0.14%, 0.05-0.63%, 0.01-0.37%, 0.15-0.42%, and not detected, respectively. After processing treatment, the contents of three iridoids, two lingnan, and one phenylpropanoid decreased in Eucommiae Cortex.

• Journal of Microbiology and Biotechnology
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• 제27권8호
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• pp.1428-1440
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• 2017

Phomopsis sp. XP-8 (an endophytic fungus) was previously found to produce pinoresinol diglucoside (PDG), a major antihypertensive compound of Tu-Chung (the bark of Eucommia ulmoides Oliv.), which is widely used in Chinese traditional medicines. In the present study, two bioconversion systems were developed for the production of PDG in Tris-HCl buffer containing glucose and Phomopsis sp. XP-8 cells (both resting and freeze-dried). When other factors remained unchanged, the bioconversion time, glucose concentration, cell ages, cell dosage, pH, temperature, and stirring speed influenced PDG production in a similar and decreasing manner after an initial increase with increasing levels for each factor. Considering the simultaneous change of various factors, the optimal conditions for PDG production were established as 70 g/l cells (8-day-old), 14 g/l glucose, $28^{\circ}C$, pH 7.5, and 180 rpm for systems employing resting cells, and 3.87 g/l cells, 14.67 g/l glucose, $28^{\circ}C$, pH 7.5, and 180 rpm for systems employing freeze-dried cells. The systems employing freeze-dried cells showed lower peak PDG production ($110.28{\mu}g/l$), but at a much shorter time (12.65 h) compared with resting cells (23.62 mg/l, 91.5 h). The specific PDG production levels were 1.92 and $24{\mu}g$ per gram cells per gram glucose for freeze-dried cells and resting cells, respectively. Both systems indicated a new and potentially efficient way to produce PDG independent of microbial cell growth.

• Natural Product Sciences
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• 제19권3호
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• pp.236-241
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• 2013

A high-performance liquid chromatography-diode array detector (HPLC-DAD) method was established for quantitative evaluation of nine constituents of Fraxinus rhynchophylla such as four coumarins, esculin (1), fraxin (2), esculetin (3), fraxetin (4), three lignans, syringaresinol 4,4'-O-${\beta}$-diglucoside (5), pinoresinol 4-O-${\beta}$-glucoside (6), pinoresinol (9), one secoiridoid, oleuropein (7), and one coumarinolignan, cleomiscosin C (8). The preferred chromatographic condition was obtained on Phenomenex Gemini-NX (3 ${\mu}m$, C18 110A, $150{\times}4.60$ mm) and the mobile phase was composed of water and acetonitrile using a gradient elution. The wavelength was set at 220 nm. Extraction condition of these constituents in F. rhynchophylla was also optimized through extraction time, extraction solvent and extraction method using established method. From this study, extraction at $70^{\circ}C$ with the mixture of ethanol and water for more than 12 h was suggested to be good extraction condition for these constituents. Quantitation of nine constituents in different F. rhynchophylla samples was also successfully accomplished with the newly established method.

• 한방재활의학과학회지
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• 제27권1호
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• pp.19-25
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• 2017

Objectives Eucommiae cortex is one of the frequently used herbs for musculoskeletal disorders, with well-known anti-inflammatory effect. Meanwhile, powdered form of herbal drugs is more advantageous than decoction form, in that storing and taking is more convenient. Thus, the authors aimed to investigate whether the drying method affects the index compound level and anti-inflammatory effect of eucommiae cortex water extract. Methods Eucommiae cortex was extracted in distilled water at $100^{\circ}C$ for 3 hours, filtered, and then evaporated under vacuum. One half of the sample was freeze-dried at $-80^{\circ}C$. Another half of the sample was added with dextrin and then spray-dried at $180^{\circ}C$. To assess the possible change in index compound content, pinoresinol diglucoside was selected as the index marker. The content level of the index compound in various extract sample was quantified through high performance liquid chromatography. In addition, iNOS assay in LPS-induced RAW 264.7 cells was adopted to determine the anti-inflammatory effect of various extract samples. Furthermore, MTT assay was performed to confirm that the result of iNOS assay was not due to cytotoxicity. Results There was no significance difference in index compound content between extract samples obtained through two different drying methods. Anti-inflammatory activity of extract samples were similar at the matching concentration, regardless of the drying methods. Extract samples did not show any significant cytotoxicity. Conclusions Extract samples of eucommiae cortex were obtained through freeze-drying and spray-drying. Neither change in index compound content nor difference in anti-inflammatory activity was observed between drying methods.

• 대한임상검사과학회지
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• 제48권3호
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• pp.176-182
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• 2016

대식세포에서 염증반응이 진행되면, interleukin-6 (IL-6), tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$) 등의 cytokine들이 발현되어 inducible nitric oxide synthase (iNOS), prostaglandin E2(PGE2) 등의 염증유발인자가 생성된다. 오가피, 우슬, 두충 각각의 추출물이 어느 정도의 항 염증 효능을 보이며 어떤 pro-inflammatory cytokine의 발현을 억제하는지에 대한 연구를 진행하였다. 오가피, 우슬, 두충은 물 추출하고 동결 건조시켰다. 각각의 추출물의 구성 성분들이 잘 추출되었는지 확인하기 위하여 지표물질인 acanthoside D, 20-hydroxyecdysone, pinoresinol diglucoside를 HPLC로 분석하였다. 항 염증 효능을 확인하기 위하여 lipopolysaccharide (LPS)로 RAW 264.7 세포주를 자극하여 염증 반응을 일으킨 상태에서 각각의 추출물을 농도 별로 처리하고 NO assay를 통해 항 염증 효능을 확인하였으며 real time PCR로 pro-inflammatory cytokine들의 발현량을 측정하였다. 결과적으로 각각의 추출물은 지표성분들이 검출되었으며 오가피와 우슬이 두충보다 NO assay에서 높은 활성을 보였다. Cytokine 발현량 측정에서는 오가피와 우슬은 iNOS와 IL-6의 발현을 억제하였고, 우슬은 TNF-${\alpha}$의 발현을 억제하였다. 우리나라는 전통적으로 약재를 조합하여 처방하여 왔다. 본 연구는 관절염에 전통적으로 사용해 오던 약재들이 어떤 기전에 의하여 항 염증 반응을 보이는지 확인하고 이들을 조합하여 사용하였을 때 어떤 근거에 의하여 시너지 효능을 보이는지 확인하였다.

• 한방재활의학과학회지
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• 제27권2호
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• pp.29-38
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• 2017

Objectives The present study aimed to investigate the change in marker compounds, anti-oxidative and anti-inflammatory effects of salt-water processed Cortex Eucommiae. Methods To evaluate the influence of processing on anti-oxidant effect of Cortex Eucommiae, changes in total phenol, total flavonoid, 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging, 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) free radical scavenging, and ferric reducing antioxidant power (FRAP) between processed and raw Cortex Eucommiae were assessed. In addition, nitrite assay was conducted to determine the influence of processing on anti-inflammatory effect of Cortex Eucommiae. Cell viability was also examined as to elucidate whether processing affects cytotoxicity of Cortex Eucommiae. Finally, high-performance liquid chromatography (HPLC) analysis was conducted to monitor changes in pinoresinol diglucoside amount of processed and raw Cortex Eucommiae. Results Salt-water processed Cortex Eucommiae showed higher total phenol and flavonoid amount, compared to raw Cortex Eucommiae. Furthermore, anti-oxidative activity of processed Cortex Eucommiae was improved as discovered in DPPH, ABTS, and FRAP assays. Anti-inflammatory effect of Cortex Eucommiae was also enhanced following salt-water processing, as evidenced in nitrite assay. HPLC analysis found that the amount of pinoresinol diglucoside, widely known as the marker compound of Cortex Eucommiae, increases through salt-water processing. All experiments were performed with non-toxic concentration of Cortex Eucommiae; processing did not affect the cytotoxicity of Cortex Eucommiae up to the currently adopted concentration. Conclusions The present results support that salt-water processing of Cortex Eucommiae is beneficial in terms of marker compound amount, anti-oxidative, and anti-inflammatory activities. Additional investigations are needed to standardize the processing method of Cortex Eucommiae.

• Natural Product Sciences
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• 제22권2호
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• pp.111-116
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• 2016

Phytochemical investigation of the stems of Pueraria lobata (Wild) Ohwi (Leguminosae), led to the isolation of eighteen known compounds: ${\beta}$-amyrone (1), (+)-pinoresinol (2), (+)-syringaresinol (3) $(+)-syringaresinol-O-{\beta}-{\small{D}}-glucoside$ (4), (+)-lariciresinol (5), (-)-tuberosin (6), naringenin (7), liquiritigenin (8), isoliquiritigenin (9) genistein (10), daidzein (11) daidzin (12) daidzein 4',7-diglucoside (13) 2,4,4'-trihydroxy deoxybenzoin (14), S-(+)-1-hydroxy-3-(4-hydroxyphenyl)-1-(4-hydroxy-2-methoxy-phenyl)propan-2-one (15), methyl $2-O-{\beta}-{\small{D}}-glucopyranosylbenzoate$ (16), pyromeconic acid $3-O-{\beta}-{\small{D}}-glucopyranoside$ 6'- (O-4''-hydroxy-3-methoxybenzoate) (17), and allantion (18). The chemical structures of these compounds were elucidated from spectroscopic data and by comparison of those data with previously published results. The effects of isolated compounds on mushroom tyrosinase enzymatic activity were screened. The results indicated that, chloroform extract of P. lobata stems turned out to be having tyrosinase inhibitory effect, and only compounds 5, 8, 9, and 11 showed enzyme inhibitory activity, with $IC_{50}$ values of $21.49{\pm}4.44$, $25.24{\pm}6.79$, $4.85{\pm}2.29$, and $17.50{\pm}1.29{\mu}M$, respectively, in comparison with these of positive control, kojic acid ($IC_{50}\;12.28{\pm}2.72{\mu}M$). The results suggest that P. lobata stems extract as well as its chemical components may represent as potential candidates for tyrosinase inhibitors.