• Nutritional Sciences
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• 제6권3호
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• pp.148-154
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• 2003

Although various raw plant materials have been demonstrated to exert anti-obesity effects to a greater or lesser extent in both humans and animals when they are used to supplement the diet, it has not been shown extensively that they influence adipocyte cell differentiation involving lipid metabolic gene expressions. Using a well-established 3T3-L1 preadipocyte differentiation system, we decided to look into molecular and cellular event occurring during adipocyte differentiation when raw plant materials aye included in the process, in an effort to demonstrate the potential use of a screening system to define the functions of traditionally well-known materials. To these ends, the effects of ethanol (EtOH) or EtOH/distilled water (DW) extracts of Wax Gourd were examined using cytochemical and molecular analyses to determine whether components of the extracts modulate adipocyte differentiation of 3T3-Ll preadipocytes in vitro. The cytochemical results demonstrated that EtOH or EtOH/DW extracts did not affect lipid accumulation and cell proliferation, although the degree of lipid accumulation was influenced slightly depending on the extract. EtOH extract was highly effective in apoptotic induction during differentiation of 3T3-Ll preadipocytes (p<0.05). Reverse transcription-polymerase chain reaction (RT-PCR) analysis of lipoprotein lipase (LPL), Uncoupling protein (Ucp) 2, 3 and 4 also showed that while LPL expression was not influenced, Ucp2, 3 and 4 were up regulated in the EtOH extract-treated group and down regulated in the EtOH/DW extract-treated group. These changes in gene expressions suggest that the components in different fractions of Wax Gourd extracts may modulate lipid metabolism by either direct or indirect action. Taking these results together, it was concluded that molecular and cellular analyses of adipocyte differentiation involving lipid metabolic genes should facilitate understanding of cellular events occurring during adipocyte differentiation. Furthermore, the experimental scheme and analytical methods used in this study should provide a screening system for the functional study of raw plant materials in obesity research.

• 한방비만학회지
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• 제22권1호
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• pp.30-37
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• 2022

Objectives: Adipogenesis is the process by which pre-adipocytes are differentiated into adipocytes. It also plays an important role in adipocyte formation and lipid accumulation. Ranunculus sceleratus (R. sceleratus) extracts are used for the treatment of various diseases such as hepatitis, jaundice, and tuberous lymphadenitis in oriental medicine. However, its effect on adipogenesis has not yet been studied. In this study, we investigated the effects of R. sceleratus on adipogenesis in 3T3-L1 cells. Methods: Cells were treated with 50, 100, and 200 ㎍/ml of R. sceleratus and cell viability was evaluated. To differentiate the 3T3-L1 preadipocytes, a 3-isobutyl-1-methylxanthine, dexamethasone, and insulin (MDI) solution were used. The accumulation of lipid droplets was determined by Oil Red O staining. The expression levels of adipogenesis-related proteins were also determined. Results: MDI solution differentiated the preadipocytes into adipocytes and accumulation of lipids was observed in the differentiated 3T3-L1 cells. Interestingly, the amount of lipid droplets was reduced after R. sceleratus treatment. In addition, the expression levels of key adipogenic transcription factors, such as CCAAT/enhancer-binding proteins-𝛼 (C/EBP-𝛼) and peroxisome proliferator-activated receptors-𝛾 (PPAR-𝛾) were also reduced after R. sceleratus treatment. Furthermore, R. sceleratus increased AMP-activated kinase (AMPK) phosphorylation and decreased sterol regulatory element-binding protein-1 expression. Conclusions: Our results showed that R. sceleratus reduced preadipocyte differentiation by inhibiting C/EBP-𝛼 and PPAR-𝛾 levels via the AMPK pathway. Therefore, we suggest that R. sceleratus may be potentially used as an anti-adipogenic agent.

• 생명과학회지
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• 제18권4호
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• pp.537-541
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• 2008

백색지방조직(white adipose tissue)은 에너지 저장뿐만 아니라 다양한 adipokines을 분비하는 중요한 내분비 기관이다. 급성기반응 단백질로 알려져 있는 합토글로빈(haptoglobin)도 adipokine의 한 종류로서 지방세포에서 합성되고 분비된다. 그러나 adipokine으로서의 기능과 지방조직에서의 역할은 아직까지 규명되지 않았다. 본 연구에서는 3T3-L1 지방전구세포를 합토글로빈 유전자로 transfection 시켜 합토글로빈을 과발현하는 세포를 만들고 세포증식, 염증관련 인자들의 발현조절 및 단구세포의 유인성을 조사하였다. 그 결과, 합토글로빈은 3T3-L1 세포의 성장에는 별 영향을 미치지 않았으나 IL-6와 COX-2 발현을 저해하고 HO-1 합성을 증가하였다. 또한 THP-1 단구세포를 이용한 invasion assay에서는 합토글로빈이 단구세포의 이동을 저해하였다. 이러한 결과들은 합토글로빈이 지방조직에서 항염증 반응에 관여함을 시사한다. 만성적 염증상태(chronic low-grade inflammatory state)로 인식되고 있는 비만은 염증관련 인자들에 의한 인슐린저항성이 유도되는 바, 합토글로빈은 비만 관련 인슐린저항성을 방지하고 인슐린민감성을 향상시키는 데에도 기여할 것으로 생각된다.

• 대한본초학회지
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• 제30권4호
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• pp.121-128
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• 2015

Objectives : Ojeok-san (OJS), an oriental herbal formula, has been used in Asian countries including Korea, China and Japan to treat the common cold and illnesses including fatigue and gastrointestinal disorders. The purpose of this study was to examine the anti-obesity effect and molecular mechanism of OJS, on adipogenesis in 3T3-L1 cells. Also, the effects of OJS in obese mice fed a high-fat diet on adiposity were examined.Methods : Preferentially, we analyzed the component of OJS and measured the stability of its component in OJS according to study periods using high performance liquid chromatography (HPLC). In vitro, 3T3-L1 cells were treated with OJS (50 to 200 μg/mL) during differentiation for 8 days. The accumulation of lipid droplets was determined by Oil Red O staining. The expressions of genes related to adipogenesis were measured by RT-PCR and Western blot analyses. For anti-obesty effect in vivo, we experimented for 8 weeks with four group (normal diet (CON), high-fat diet (HF), high-fat diet with OJS (HF+OJS) and high-fat diet with Bang-pung-tong-sung-san (HF+BTS) in comparison group HF+OJS).Results : OJS showed inhibitory activity on adipocyte differentiation at 3T3-L1 preadipocytes without affect cell toxicity as assessed by measuring fat accumulation and adipogenesis. In addition, OJS significantly reduced the expression levels of several adipocyte marker genes including proliferator activated receptor-γ(PPAR-γ) and CCAAT/enhancer-binding protein-α(C/EBP-α). Also OJS-administered mice showed significant inhibitory of body weights and abdominal adipose tissue weights.Conclusions : This study showed that traditional medicine OJS has an anti-obesity effect in vitro and in vivo. Thus, OJS could be developed as a supplement for reduction of body weight gain induced by an obesity.

• 대한약침학회지
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• 제11권1호
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• pp.149-162
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• 2008

Objectives : The purpose of this study is to investigate the effects of hot pepper extract and capsaicin on the adipogenesis in 3T3-L1 cells, lipolysis in rat epididymal adipocytes and histological changes in porcine adipose tissue. Methods : Inhibiton of preadipocyte differentiation and/or stimulation of lipolysis play important roles in reducing obesity. 3T3-L1 preadipocytes were differentiated with adipogenic reagents by incubating for 3 days in the absence or presence of hot pepper extract or capsaicin ranging from 0.01 to $1mg/m{\ell}$. The effects of hot pepper extract and capsaicin on adipogenesis were examined by measuring GPDH activity and by Oil Red O staining. Mature adipocytes from rat epididymal fat pad was incubated with hot pepper extract or capsaicin ranging from 0.01 to $1mg/m{\ell}$ for 3 hrs. The effects of hot pepper extract and capsaicin on lipolysis were examined by measuring free glycerol released. Fat tissue from pig skin was injected with hot pepper extract or capsaicinCFP ranging from 0.1 to $10mg/m{\ell}$ to examine the effects of hot pepper extract and capsaicin on histological changes under light microscopy. Results : The following results were obtained from present study on adipogenesis of preadipocytes, lipolysis of adipocytes and histological changes in fat tissue. 1. Hot pepper extract and capsaicin inhibited adipogenic differentiation at the concentration of 0.1 and $0.01mg/m{\ell}$, respectively, indicating that capsaicin was more effective in inhibiting adipogenesis than hot pepper extract. 2. Hot pepper extract and capsaicin decreased the activity of glycerol-3-phosphate dehydrogenase(GPDH) at the concentration of 0.1 and $0.01mg/m{\ell}$, respectively, indicating that capsaicin was more effective in inhibiting adipogenic differentiation than hot pepper extract. 3. Hot pepper extract and capsaicin increased glycerol release at the concentration of $0.1mg/m{\ell}$. There was no difference in lipolytic activity between hot pepper extract and capsaicin at the corresponding concentration. 4. Hot pepper extract and capsaicin caused shrinkage of fat cells, resulting in cell death at the concentration of $1.0mg/m{\ell}$, although capsaicin exerted this action over wide area than hot pepper extract. Conclusions : These results suggest that hot pepper extract and capsaicin efficiently inhibited adipogenesis, increased lipolysis of adipocytes and caused to shrink fat cells. Future studies are needed to make use of hot pepper extract pharmacopuncture for the treatment of obesity.

• Journal of Ginseng Research
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• 제41권3호
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• pp.257-267
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• 2017

Background: Heat-processed ginseng, sun ginseng (SG), has been reported to have improved therapeutic properties compared with raw forms, such as increased antidiabetic, anti-inflammatory, and antihyperglycemic effects. The aim of this study was to investigate the antiobesity effects of SG through the suppression of cell differentiation and proliferation of mouse 3T3-L1 preadipocyte cells and the lipid accumulation in Caenorhabditis elegans. Methods: To investigate the effect of SG on adipocyte differentiation, levels of stained intracellular lipid droplets were quantified by measuring the oil red O signal in the lipid extracts of cells on differentiation Day 7. To study the effect of SG on fat accumulation in C. elegans, L4 stage worms were cultured on an Escherichia coli OP50 diet supplemented with $10{\mu}g/mL$ of SG, followed by Nile red staining. To determine the effect of SG on gene expression of lipid and glucose metabolism-regulation molecules, messenger RNA (mRNA) levels of genes were analyzed by real-time reverse transcription-polymerase chain reaction analysis. In addition, the phosphorylation of Akt was examined by Western blotting. Results: SG suppressed the differentiation of 3T3-L1 cells stimulated by a mixture of 3-isobutyl-1-methylxanthine, dexamethasone, and insulin (MDI), and inhibited the proliferation of adipocytes during differentiation. Treatment of C. elegans with SG showed reductions in lipid accumulation by Nile red staining, thus directly demonstrating an antiobesity effect for SG. Furthermore, SG treatment down-regulated mRNA and protein expression levels of peroxisome proliferator-activated receptor subtype ${\gamma}$ ($PPAR{\gamma}$) and CCAAT/enhancer-binding protein-alpha ($C/EBP{\alpha}$) and decreased the mRNA level of sterol regulatory element-binding protein 1c in MDI-treated adipocytes in a dose-dependent manner. In differentiated 3T3-L1 cells, mRNA expression levels of lipid metabolism-regulating factors, such as amplifying mouse fatty acid-binding protein 2, leptin, lipoprotein lipase, fatty acid transporter protein 1, fatty acid synthase, and 3-hydroxy-3-methylglutaryl coenzyme A reductase, were increased, whereas that of the lipolytic enzyme carnitine palmitoyltransferase-1 was decreased. Our data demonstrate that SG inversely regulated the expression of these genes in differentiated adipocytes. SG induced increases in the mRNA expression of glycolytic enzymes such as glucokinase and pyruvate kinase, and a decrease in the mRNA level of the glycogenic enzyme phosphoenol pyruvate carboxylase. In addition, mRNA levels of the glucose transporters GLUT1, GLUT4, and insulin receptor substrate-1 were elevated by MDI stimulation, whereas SG dose-dependently inhibited the expression of these genes in differentiated adipocytes. SG also inhibited the phosphorylation of Akt (Ser473) at an early phase of MDI stimulation. Intracellular nitric oxide (NO) production and endothelial nitric oxide synthase mRNA levels were markedly decreased by MDI stimulation and recovered by SG treatment of adipocytes. Conclusion: Our results suggest that SG effectively inhibits adipocyte proliferation and differentiation through the downregulation of $PPAR{\gamma}$ and $C/EBP{\alpha}$, by suppressing Akt (Ser473) phosphorylation and enhancing NO production. These results provide strong evidence to support the development of SG for antiobesity treatment.

• 생명과학회지
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• 제24권5호
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• pp.505-514
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• 2014

항 비만 활성소재를 탐색하기 위하여 국내산 8종의 잡곡으로부터 80% 에탄올 추출물을 확보하여 마우스 3T3-L1 전지방세포에 대한 세포독성을 조사한 결과, 기장이 가장 강한 세포독성을 보였다. 따라서 본 연구에서는 기장 에탄올 추출물을 다시 메칠렌 크로라이드, 에칠아세테이트 및 부탄올로 단계 추출하여 각 유기용매 추출물을 확보하고 각 추출물의 항 비만 활성소재를 3T3-L1전지방세포의 지방세포로의 분화 억제능으로 조사하였다. 그 결과 기장의 부탄올 추출물은 지방세포 분화 유도기간(6일) 동안 처리하였을 때, 대조구보다 중성지방 축적을 약 50.3% 억제하는 것으로 나타나 가장 강한 항비만 활성 분획임을 확인하였다. 이와 같은 기장 부탄올 추출물의 항비만 활성은 지방세포 분화유도인자($(C/EBP{\alpha}$, $PPAR{\gamma}$)의 초기 발현을 억제하여 전지방세포의 지방세포로의 분화를 억제하고, 나아가 지방대사 관련 유전자(aP2 와LPL)의 발현까지도 저해함을 알았다. 그리고 기장의 부탄올 추출물은 25 ${\mu}g/ml$ 농도에서는 3T3-L1 전지방세포의 지방세포로의 분화를 저해하지만 50 ${\mu}g/ml$농도에서는 3T3-L1 전지방세포의 미토콘드리아 세포막 전위(${\Delta}{\psi}m$)를 변화시키고, caspase-3 활성화와 PARP 분해를 유도하여 세포사멸을 유도하였다. 이와 같은 결과는 기장이 항비만 소재를 가진 우수한 잡곡으로서 비만 관련 질환 예방에 우수한 식소재로 활용할 수 있음을 시사한다.

• 한국미생물·생명공학회지
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• 제42권3호
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• pp.249-257
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• 2014

본 연구에서는 백두구(A. cardamomum L.) 메탄올 추출물(ACME)의 항산화 및 항비만 활성을 DPPH radical 소거능과 췌장 lipase 효소 활성 억제능, 그리고 세포실험계를 이용하여 분석하였다. 그 결과 ACME는 DPPH radical을 농도 의존적으로 소거하였으며 DPPH radical 소거능의 50% 저해농도($IC_{50}$)는 $25.15{\mu}g/ml$로 나타났다. 또한 ACME는 농도 의존적으로 lipase 효소 활성을 유의적으로 억제시켰으며, 3T3-L1 preadipocyte를 이용하여 지방세포 분화 및 지방생성, 생성된 지방의 분해에 미치는 영향을 분석한 결과 ACME는 지방세포 분화, 세포 내 지방 축적, TG 함량 등을 독성 없이 농도의존적으로 억제하였으며 지방세포 내 중성지방을 유의적으로 분해시키는 것으로 나타났다. 이러한 백두구의 지방세포 분화 억제능은 핵심 작용 인자인 $C/EBP{\alpha}$, $C/EBP{\beta}$, 그리고 $PPAR{\gamma}$의 유전자 및 단백질 발현조절에서 기인함을 확인하였다. 이러한 결과는 백두구가 보유한 항산화능과 췌장 lipase 활성 저해능, 지방세포 분화 억제능, 지방세포 내 지방 분해능을 통한 항비만 활성을 처음으로 밝혀낸 것이며 추후 계속적인 연구를 통해 활성 물질의 규명이 필요할 것으로 판단된다.

• 고려인삼학회:학술대회논문집
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• 고려인삼학회 2007년도 춘계 학술대회
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• pp.23-24
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• 2007

Purpose: 인삼이 항당뇨 활성을 가진다는 연구가 많은 연구자들에 의해 진행되었고, 이는 인삼의 구성 성분 중 ginsenoside에 기인한다는 보고가 있다. 본 연구는 ginsenoside의 항당뇨 작용기전을 in vitro에서 알아보고자 3T3-L1 지방세포에서 glucose uptake와 췌장 베타세포인 HIT-T15 세포에서 insulin 분비 효과를 확인하였다. 이를 위하여 인삼을 식초로 처리한 긴삼의 70% MeOH 분획으로부터 protopanaxadiol 계인 ginsenoside $Rb_2$, $Rg_3$ 그리고 protopanaxtriol 계인 $Rg_2$를 분리하여 본 실험에 사용하였다. Method: Ginsenoside $Rb_2$, $Rg_2$, $Rg_3$가 지방 세포에서 glucose uptake에 미치는 효과를 확인하기 위하여 3T3-L1 세포를 DMEM (Dulbecco's Modified Eagle's Medium) 배지에서 분화 유도시켰으며 3T3-L1 preadipocyte가 80% 정도 자라면 분화 유도 배지 (5% fetal bovine serum (FBS), 0.5 mM isobutylmethylxanthine (IBMX), 1 mM dexamethasone 그리고 $10{\mu}g/ml$ insulin가 포함된 DMEM)로 4일, $10{\mu}g/ml$ insulin가 포함된 DMEM으로 2일, FBS만 포함된 DMEM으로 2일 배양하여 총 8일 동안 분화를 유도하였다. 분화 유도된 3T3-L1 adipocytes 에 각각 $Rb_2$, $Rg_2$, $Rg_3$를 $20{\mu}M$로 처리하여 16시간 배양하여 low glucose DMEM에서 3시간 배양한 후에 $37^{\circ}C$에서 insulin 10 ng/ml 과 각각 $Rb_2$, $Rg_2$, $Rg_3$가 포함된 Krebs Ringer Hepes buffer(KRP buffer)에서 20분간 배양하였다. 2-deoxy-D-[$^3H$]-glucose를 넣고 10분 후에 차가운 PBS로 반응을 종결시켜 lysis buffer로 cell을 모은 후 scintillation counter를 이용하여 glucose를 측정하였다. Insulin 분비 효과는 HIT-T15 세포와 일차 배양한 흰쥐 소도세포(islets)를 사용하여 확인하였다. HIT-T15 세포는 24 well plate에 well 당 $2{\times}10^5$ 개씩 분주하여 24시간 동안 배양한 후 시료를 처리하였으며 소도 세포는 Sprague-Dawley rat의 췌장에 collagenase가 포함된 Hanks' Balanced Salt Solution(HBSS)을 주입하여 분리하고 islets을 얻었다. 분리한 소도세포를 $1{\sim}2$일 동안 배양하여 $Rb_2$, $Rg_2$, $Rg_3$가 각각 $20{\mu}M$의 농도로 첨가된 insulin 측정용 buffer인 Krebs-Ringer buffer (KRB+0.3% BSA, KRBB)에 $37^{\circ}C$에서 1시간 incubation 시킨 후 배양액으로 분비된 인슐린의 양을 측정하였다. 한편 ginsenoside의 인슐린 분비 촉진 기전을 알아보기 위한 실험에서는 ATP-sensitive $K^+$ channel opener인 diazoxide (0.5 mM)가 ginsenoside에 의해 촉진된 인슐린 분비를 억제하는지 살펴보았다. Result: glucose uptake assay 에서는 $Rg_2$가 가장 크게 glucose uptake를 증가시켰고 $Rb_2$, $Rg_3$는 그 활성이 크지 않았다. 한편 Insulin 분비 효과는 diol계인 $Rg_3$에서 용량 의존적으로 인슐린의 분비를 촉진시켰으며 $20{\mu}M$ 농도에서 대조군과 비교해 1.5배 이상의 분비 촉진 효과를 보였고 triol계인 $Rg_2$ 에서는 이러한 효과가 나타나지 않았다. $Rg_3$의 인슐린 분비 촉진 기전을 0.5 mM 의 diazoxide를 이용하여 확인한 결과 $Rg_3$에 의해 촉진된 인슐린 분비를 감소시켰다. 이로 미루어보아 $Rg_3$의 인슐린 분비 촉진 기전은 ATP-sensitive $K^+$ 채널의 봉쇄에 의한 것임을 확인할 수 있었다.

• 생명과학회지
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• 제27권8호
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• pp.888-895
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• 2017

솔장다리(S. collina)는 건조한 대지에 널리 분포되어 있는 한해살이 식물로 한방에서는 고혈압의 치료에 사용되어왔으며 선행연구를 통해 솔장다리가 보유한 항산화 및 항암 활성을 밝힌바 있다. 본 연구에서는 솔장다리 에탄올 추출물(SCEE)의 항비만 활성을 췌장 lipase 효소 활성 억제능과 세포실험계를 이용하여 분석하였다. 그 결과 SCEE는 농도 의존적으로 lipase 효소 활성을 유의적으로 억제시켰으며, 3T3-L1 preadipocyte를 이용하여 지방세포 분화 및 지방생성, 생성된 지방의 분해에 미치는 영향을 분석한 결과 지방세포 분화, 세포 내 지방 축적, TG 함량 등을 독성 없이 농도의존적으로 억제하였고, 지방세포 내 중성지방을 유의적으로 분해시키는 것으로 나타났다. 이러한 솔장다리의 지방세포 분화 억제능은 핵심 작용 인자인 $C/EBP{\alpha}$, $C/EBP{\beta}$, 그리고 $PPAR{\gamma}$의 유전자 및 단백질 발현조절에서 기인함을 확인하였다. 이러한 결과는 솔장다리가 보유한 췌장 lipase 활성 저해능, 지방세포 분화 억제능, 지방세포 내 지방 분해능, 지방분화관련 인자 신호전달기전 조절을 통한 항비만 활성을 처음으로 밝혀낸 것이며 추후 계속적인 연구를 통해 활성 물질의 규명이 필요할 것으로 판단된다.