• Archives of Pharmacal Research
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• 제27권9호
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• pp.968-972
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• 2004

A 1, 2-diglyceride-based multi-step colorimetric assay to measure the pancreatic lipase activ-ity was applied for the determination of the kinetic profiles of the lipase inhibition with a slight modification and the validity verification. With this assay method, our study revealed that platy-codin D, one of major constituents of Platycodi Radix, inhibits the pancreatic lipase activity in a competitive type, with the value of $K_1$ being 0.18${\pm}$0.02 mM. In addition, PO has affected the val-ues of $K_{m}$, app/ and $K_{cat}$/$K_{m}$ in a dose-dependent manner. The results shed a meaningful light on how PO mediates lipid metabolism in the intestinal tracts. On the other hand, since the revised assay is sensitive, rapid, and does not affect the accuracy to the kinetic properties, it is applica-ble not only to evaluation of the kinetic properties of the pancreatic lipase, but also to high-throughput screening of pancreatic lipase activity.

• Archives of Pharmacal Research
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• 제27권10호
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• pp.1048-1052
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• 2004

A 1, 2-diglyceride-based multi-step colorimetric assay to measure the pancreatic lipase activity was applied for the determination of the kinetic profiles of the lipase inhibition with a slight modification and the validity verification. With this assay method, our study revealed that platycodin D, one of major constituents of Platycodi Radix, inhibits the pancreatic lipase activity in a competitive type, with the value of $K_I$ being 0.18${\pm}$0.02 mM. In addition, PD has affected the values of $K_{m,app}\;and\;K_{cat}/K_m$ in a dose- dependent manner. The results shed a meaningful light on how PD mediates lipid metabolism in the intestinal tracts. On the other hand, since the revised assay is sensitive, rapid, and does not affect the accuracy to the kinetic properties, it is applicable not only to evaluation of the kinetic properties of the pancreatic lipase, but also to highthroughput screening of pancreatic lipase activity.

• 생명과학회지
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• 제18권8호
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• pp.1159-1163
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• 2008

이 연구의 목적은 물-오일 계면에서 특이적인 lipase 활성을 측정할 수 있는 high-throughput assay 방법을 확립하는 것이다. 이 방법은 pH에 따라 형광의 세기가 변하는 fluorescein의 특성을 이용하여 lipase의 작용으로 방출되는 지방산으로 인한 pH 변화를 fluorescein의 형광 변화로 측정하도록 되어 있다. 활성의 측정은 오일 에멀션과 fluorescein 그리고 효소를 포함하는 반응 용액을 반응시키면서 일정한 간격으로 형광을 측정함으로써 이루어진다. 이 방법을 통해 형광의 세기가 효소의 양에 비례하는 속도로 감소하는 것을 관찰할 수 있었으며 시간에 따른 형광 변화 그래프로부터 계산한 반응 속도가 효소의 양에 선형으로 비례한다는 것을 확인할 수 있었다. 또 한 가지 중요한 사실은 assay를 하는데 있어서 pH 6.0-8.0의 범위에서 다른 pH 조건을 사용할 수 있었다는 점이다.

• BMB Reports
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• 제36권4호
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• pp.417-420
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• 2003

The present work describes a colorimetric microplate assay for lipase activity based on the reaction between 5,5'-dithiobis(2-nitro benzoic acid) (DTNB) and the hydrolysis product of 2,3-dimercapto-1-propanol tributyrate (DMPTB). Reaction mixtures containing DTNB, DMPTB, and lipase were prepared in microplate wells, and the absorbance at 405nm was recorded after incubation at $37^{\circ}C$ for 30 min. A linear relationship was obtained in the range of 0.1-1 U of lipase activity by this method. The reaction conditions were also optimized for the range of 0.01-0.1 U or 1-10 U. When assaying crude tissue extracts, the reaction of DTNB with non-specific reducing agents created a major source of error. However, this error was corrected by the use of blank samples that did not contain DMPTB.

• 한국식품영양과학회지
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• 제42권4호
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• pp.556-562
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• 2013

본 연구에서는 기존의 유기용매를 이용한 추출을 지양하고 환경 친화적이고 간단한 추출방법을 고안하여 톳의 다당류 성분(WSP-A)을 추출해내었고, WSP-A가 지질의 흡수에 미치는 효과를 알아보기 위하여 in vitro 상으로 lipase inhibition assay와 in vivo로 고지방 식이로 비만을 유도한 rat을 이용한 동물실험을 수행하였다. 그 결과 톳 유래 다당류는 alginate와 유사한 성분이지만 성분당 분석을 통하여 fucose가 주 당성분인 polysaccharide로 밝혀졌고, 이는 alginate보다 뛰어나고 안정한 lipase inhibitor로써 작용하였다. 또한 동물실험에서는 WSP-A를 포함하는 음용수로 제공한 것과 제공하지 않은 대조군은 무게 변화량에 유의적인 차이를 보일 뿐 아니라 비만 예방군과 비만 치료군 사이에서도 유의적인 차이를 나타내었다. 식이섬유를 이용한 연구와 비교해보았을 때, 톳 유래 다당류는 식이섬유 이상의 효과뿐만 아니라 뛰어난 lipase inhibitor로 체중감량을 나타내었고 이는 인간췌장유래 리파아제(human pancreatic lipase)의 저해활성과 더불어 인체 내에서의 작용 가능성을 짐작하고 있다. 따라서 톳 유래 다당류 성분은 식이성 섬유의 공급원 뿐만 아니라 추후에 효소활성 저해 효과에 대한 보다 명확한 작용기전을 밝혀낸다면 단순한 건강 기능성 식품 소재보다 lipase inhibitor로써 항비만 약물로 고부가가치를 창출해낼 수 있을 것으로 사료된다.

• 한국염색가공학회지
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• 제8권4호
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• pp.25-30
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• 1996

This study was carried out to examine the effect of lipase on the removal of tripalmitin in the various conditions of washing. The relations between the removal and the hydrolysis of tripalmitin by lipase were discussed. The hydrolysis characteristics of lipase were examined by a colorimetric determination of liberated fatty acids as a new assay of lipase in reverse micelies. The hydrolysis of tripalmitin by lipase was increased with the increase of reaction time and reaction above lipase concentration 150mg/l pH at reaction temperature 4$0^{\circ}C$.

• 한국자원식물학회:학술대회논문집
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• 한국자원식물학회 2018년도 춘계학술발표회
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• pp.81-81
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• 2018

The present study was investigated on in vitro anti-obesity effect of 4-hydroxybenzyl alcohol from Cudrania tricuspidata. We isolated various compounds from Cudrania tricuspidata. Among these compounds, anti-obesity effects of 4-hydroxybenzyl alcohol was examined by lipase activity assay, cyclic adenosine monophosphate (cAMP)-specific phosphodiesterase type IV (PDE4) activity assay, and citrate synthase activity assay. 4-hydroxybenzyl alcohol and Cudrania tricuspidata extracts inhibited the enzymatic activities of lipase, PDE4, and citrate synthase. Lipase is known to mediate the hydrolysis of triacylglycerol in adipose tissue and cholesterol esters in other tissue or cells. Also, PDE4 hydrolyses cAMP, a crucial secondary messenger for in metabolic pathways including glucose and lipid metabolism, lipolysis, and thermogenic function. 4-hydroxybenzyl alcohol and Cudrania tricuspidata extracts induced the inhibitory effect against each enzymatic activity on several specific substrates as observed by detection at 405 or 412 nm. These findings might be attributable to the inhibition of adipogenesis, and partial prevention of obesity. In conclusion, these results show that 4-hydroxybenzyl alcohol and Cudrania tricuspidata may be a critical candidate as a natural anti-obesity source.

• 한국미생물·생명공학회지
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• 제48권4호
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• pp.491-505
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• 2020

A newly isolated strain, Proteus vulgaris OR34, from olive mill waste was found to secrete an alkaline extracellular lipase at 11 U·ml-1 when cultivated on an optimized liquid medium. This lipase was purified 94.64-fold with a total yield of 9.11% and its maximal specific activity was shown to be 3232.58 and 1777.92 U·mg-1 when evaluated using the pH-stat technique at 55℃ and pH 9 and Tributyrin TC4 or olive oil as the substrate. The molecular mass of the pure OR34 lipase was estimated to be around 31 kDa, as revealed by SDS-PAGE and its substrate specificity was investigated using a variety of triglycerides. This assay revealed that OR34 lipase preferred short and medium chain fatty acids. In addition, this lipase was stable in the presence of high concentrations of bile salt (NaDC) and calcium ions appear not to be necessary for its activity. This lipase was inhibited by THL (Orlistat) which confirmed its identity as a serine enzyme. In addition, the immobilization of OR34 lipase by adsorption onto calcium carbonate increased its stability at higher temperatures and within a larger pH range. The immobilized lipase exhibited a high tolerance to organic solvents and retained 60% of its activity after 10 months of storage at 4℃. Finally, the OR34 lipase was applied in biodiesel synthesis via oleic acid mediated esterification of methanol when using hexane as solvent. The best conversion yield (67%) was obtained at 12 h and 40℃ using the immobilized enzyme and this enzyme could be reused for six cycles with the same efficiency.

• 한국식품영양학회지
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• 제32권1호
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• pp.27-32
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• 2019

The objective of this study was to evaluate novel usability as natural anti-obesity supplement of Selaginella tamariscina extract. The total phenol contents and total flavonoid contents were $60.29{\pm}3.11GAE\;mg/g$ and $14.90{\pm}0.34QE\;mg/g$, respectively. To evaluate anti-obesity activity of Selaginella tamariscina extract, pancreatic lipase inhibition activity as well as its inhibition effect of lipid accumulation in adipocytes were conducted by Oil Red O staining and lipolysis assay. The result of pancreatic lipase inhibition activity of S. tamariscina extract showed a wide range between 40 and 73% dose dependently. While the incubation of 3T3-L1 cells with S. tamariscina extract inhibited differentiation of preadipocytes and reduced lipid accumulation, the level of released free glycerol into culturing medium was increased in multiple concentrations. These results showed that S. tamariscina extract inhibit adipogenesis and pancreatic lipase activity. Thus, S. tamariscina extract can be a candidate for regulating lipid accumulation in obesity.

• 한국식품저장유통학회지
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• 제18권1호
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• pp.53-58
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• 2011

신선한 울릉도 민들레 뿌리를 70% ethanol로 침지 추출하여 얻어진 추출물을n-hexane, EtOAc, n-BuOH로 순차용매 분획하였다. 이중 상대적으로 높은 pancreatic lipase 저해활성을 나타낸 EtOAc 분획에 대해 Toyopearl 및 $C_{18}$겔을 활용한 column chromatography를 수행하여 5종의 페놀성 화합물을 분리하였다. 각 화합물의 화학구조는 NMR 스펙트럼 데이터 해석 및 표품과의 HPLC 직접 비교를 통하여 3,5-di-O-caffeoylquinic acid, chicoric acid, caffeic acid, protocatechuic aldehyde, luteolin 으로 동정하였다. 이들 화합물중 3,5-di-O-caffeoylquinic acid 는 $IC_{50}$ 값이 $65.1\;{\pm}0.7\; {\mu}M$로 가장 강한 효능을 나타내었으며, 다음으로 tartaric acid의 수산기에 2분자의 caffeic acid가 결합한 chicoric acid 의 IC50 값이 $126.3{\pm}2.3{\mu}M$의 저해능을 나타내었다. 민들레 뿌리의 EtOAc 가용부에에 존재하는 caffeic acid 유도체가 pancreatic lipase 저해활성 물질임을 확인하였으며, 이들 활성은 caffeic acid가 결합한 quinic acid 및 tatrtaric acid 등의 구성화합물에 따라 다름이 시사되었다. 향후 이들 활성물질의 활성 기작에 대한 연구가 필요하며 본 연구결과는 보다 우수한 pancreatic lipase 저해능을 가지는 새로운 선도화합물 발굴을 위한 기초자료로 이용될 수 있을 뿐만 아니라 울릉도에 자생하는 민들레 뿌리의 식물 화학적 성분에 대한 기초자료로 이용될 수 있을 것으로 사료된다.