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Inhibition of Angiotensin I Converting Enzyme and Purification of Peptides from Protein Hydrolysate of Hard Clam
|Authors: ||Jia-Ling Chen|
|Contributors: ||NTOU:Department of Food Science|
Hard Clam;Angiotensin I- Converting Enzyme
|Issue Date: ||2011-06-30T07:42:12Z
|Abstract: ||將本省產生鮮文蛤肉以熱水抽出法處理，剩餘殘肉經由凍乾成粉末後，分別用不同蛋白酶Protease N( PN )、Prozyme 6( P6 )及Protamex( PX )於50℃進行一次水解5小時，或以Flavourzyme二次水解0.5小時，探討文蛤肉水解物對血管升壓素轉換酶 (ACE) 之抑制及其調節血壓的生理效果。文蛤肉水解物中只有以Protease N水解之產物具苦味，其餘皆不具苦味。胜肽含量以Prozyme 6經一次及二次水解最高，為523 mg/g。對ACE抑制能力而言，以Protamex 水解5小時者 (PX5) 最佳，IC50值為0.036 mg/ml。將不具苦味且抑制ACE能力效果佳之水解物 ( PX5)與Captopril (治療高血壓的藥物)比較，顯示文蛤水解物是屬於混合型之抑制類型，Ki值為0.027 mg/ml，而Captopril則為競爭型之抑制劑，Ki值為0.0067 μg/ml。 文蛤水解物(PX5)經腸胃道酵素作用後，IC50值由0.036升至0.060 mg/ml，對ACE之抑制活性下降。PX5以膠體層析法進行劃分，得五個主要劃分區，其中以分子量介於350至300Da間之劃分物具有最強的ACE抑制活性。PX5之劃分收集物E1及E2，經鑑定序列分別為Val-Arg-Lys及Tyr-Asn。|
Fresh hard clam meat was extracted using hot water. The meat residue was freeze dried then hydrolyzed by Protease N (PN)、Prozyme 6 (P6) or Protamex (PX) as primary hydrolysis followed by a secondary hydrolysis (Flavourzyme, F) at 50℃. The effects of hard clam hydrolysates on inhibitory activity against angiotensin I converting enzyme (ACE) and regulating blood pressure were investigated.Only the PN hydrolyzed product tasted bitter. The maximal peptide content (523 mg/g) of hydrolysate was obtained from P6 hydrolysis for 5 hr followed by F hydrolysis for 0.5 hr. However, the lowest IC50 of hydrolysate on ACE (0.036 mg/ml) was obtained by PX hydrolysis for 5 hr (PX5). The kinetics of the hydrolysate on ACE-inhibition indicated that PX5 and Captopril (drug for hypertensive) indicated a mixed-type inhibition and competitive-type inhibition, respectively. Their Ki values were 0.027 mg/ml and 0.0067 μg/ml, repectively. The hydrolysate of PX5 was digested by gastrointestinal proteases, and the IC50 of ACE was increased from 0.036 to 0.060 mg/ml. The hydrolysate of PX5 was fractionated by gel permeation chromatography with Sephadex G-25. A fraction of 350-300 Da was found with the highest inhibitory efficiency ratio being 5831.7 %/mg/ml. The E1 and E2 amino acid sequences of PX5 were Val-Arg-Lys and Tyr-Asn, respectively.
|Appears in Collections:||[食品科學系] 博碩士論文|
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