A novel angiotensin-converting enzyme (ACE) inhibitory peptide from tilapia skin: Preparation, identification and its potential antihypertensive mechanism

3.0 科研～小助 2025-09-01 4 4 3.09MB 11 页 1知币

侵权投诉

Food Chemistry 430 (2024) 137074

Available online 2 August 2023

A novel angiotensin-converting enzyme (ACE) inhibitory peptide from

tilapia skin: Preparation, identication and its potential

antihypertensive mechanism

Ye Dong , Wen Yan , Yi-Qi Zhang

, Zhi-Yuan Dai

Collaborative Innovation Center of Seafood Deep Processing, Key Laboratory of Aquatic Products Processing of Zhejiang Province, Institute of Seafood, Zhejiang

Gongshang University, Hangzhou 310035, China

ARTICLE INFO

Keywords:

Fish skin

Steam explosion

Purication

ACE inhibitory peptides

Molecular docking

ABSTRACT

To obtain food-derived peptides with high ACE inhibitory activity, tilapia skin was pretreated with steam ex-

plosion prior to enzymatic hydrolysis. The results showed that steam explosion pretreatment improved the hy-

drolysis efciency and ACE inhibitory activity of sh skin hydrolysates. A novel ACE inhibitory peptide

VGLFPSRSF (1009.17 Da) was obtained from steam-exploded sh skin hydrolysates. VGLFPSRSF had an IC

value of 61.43

M for ACE inhibitory activity, showing a non-competitive binding mode and gastrointestinal

enzyme hydrolysis resistance. Molecular docking results showed that VGLFPSRSF interacted with ACE receptor

protein through hydrogen bonding and hydrophobic interactions. Based on the results of network pharmaco-

logical analysis and molecular docking, VGLFPSRSF might regulate blood pressure through interaction with

hypertensive targets such as AKT1, ACE, CD4, REN, and MMP9. Steam-exploded tilapia skin peptides had po-

tential antihypertension activity and might be promising to achieve high-value utilization of sh skin by-

products.

1. . Introduction

Hypertension, “the silent killer”, is a common chronic disease and

the predominant risk factor for cardiovascular and cerebrovascular

disease, with reports indicating that more than 1.6 billion people will

have hypertension by 2025 (Chen, Wang, Shu, & Li, 2021). Conven-

tional pharmacological treatment of hypertension has focused on the

inhibition of renin and ACE activity or inhibition of angiotensin II

binding to receptors (Khurana & Goswami, 2022). The side effects of

taking chemical drugs for hypertension cannot be ignored (Feng et al.,

2022). It is crucial to explore natural and healthy anti-hypertensive

products. Food-derived ACE inhibitory peptide has attracted wide-

spread interest (Piovesana et al., 2018). Studies have demonstrated that

the ACE inhibitory activity of peptides depends on the sequence and

molecular weight (Li et al., 2018). Higher ACE inhibitory activity was

observed when the C-terminal were aromatic amino acids, proline, and

hydroxyproline (Zhang, Olsen, Grossi, & Otte, 2013). In addition, pep-

tides with hydrophobic or basic amino acids at the N-terminal exhibited

higher ACE inhibitory activity (Xiang, Qiu, Zhao, Zheng, & Qiao, 2023).

The pyrrole ring present in proline plays an important role in the

inhibition of ACE, and due to the presence of proline, peptides exhibit

greater stability in gastrointestinal digestion (Chen, Sun, Li, & Liu, 2021;

Murray & FitzGerald, 2007).

Tilapia (Oreochromis niloticus) is an important freshwater farmed sh

in the global market. In 2018, the total production of tilapia in China

was 1.625 million tons, accounting for about 76% of total global tilapia

exports (Cao et al., 2022), primarily in the form of sh llets. Inevitably,

numerous sh by-products are generated, which are usually considered

as waste for disposal, leading to ecological and economic issues.

Particularly, tilapia skin is a good source of collagen, with proper pro-

cessing, these raw materials can nd effective applications in numerous

elds (Munawaroh et al., 2022). Collagen accounts for about 30% of the

total protein content and is the predominant brous protein in the

extracellular matrix (Abdollahi, Rezaei, Jafarpour, & Undeland, 2018),

rich in proline, hydroxyproline and hydrophobic amino acids, with po-

tential ACE inhibitory activity and better stability (Chen et al., 2021a).

However, the triple helix structure of collagen, which is stabilized by

covalent cross-linking and hydrogen bonding, is difcult to be cleaved

by common commercial proteases except matrix metalloprotease family,

substantially limits its application (Zhang et al., 2013). In this

* Corresponding author.

E-mail address: zhangyq@zjgsu.edu.cn (Y.-Q. Zhang).

Contents lists available at ScienceDirect

Food Chemistry

journal homepage: www.elsevier.com/locate/foodchem

https://doi.org/10.1016/j.foodchem.2023.137074

Received 27 May 2023; Received in revised form 27 July 2023; Accepted 31 July 2023

Food Chemistry 430 (2024) 137074

perspective, pretreatment is required before enzymatic hydrolysis to

break the triple helix structure in natural collagenous materials to

ensure maximize the release of ACE active peptides.

Recently, application of hydrothermal pretreatment in the macro-

molecule substances hydrolysis has been explored. Under hydrothermal

conditions, water has unique features, such as increased ionic and

structural changes in the hydrogen bond. The increase in ionic product

drives the formation of H

and OH

−

ions, which facilitates water to

appear as an acid or base catalyst (Ahmed & Chun, 2018). Steam ex-

plosion pretreatment is a typical hydrothermal process, involving a

short-time treatment of the biomass under saturated vapor pressure,

followed by explosive decompression within 0.1 s. Natural collagenous

materials contain a large number of intermolecular interaction forces

such as hydrogen bonds, which are easy to break under high tempera-

ture conditions (Zhang et al., 2023), thereby exposing internal sites that

can be hydrolyzed by commercial proteases. Thus, we speculate that

steam explosion may effectively break the triple helix structure in nat-

ural sh skin raw materials and promote the efcient release of ACE

inhibitory peptides.

The objective of this study was to prepare sh skin hydrolysates with

high ACE inhibitory activity by combining steam explosion pretreat-

ment and enzymatic hydrolysis, to obtain ACE inhibiting peptides

through separation, purication, identication and in silico screening.

The inhibition pattern and gastrointestinal digestive stability of the

peptide were further investigated. Finally, the interaction of ACE

inhibitory peptides with core targets in hypertension were explored

through network pharmacology and molecular docking. This strategy

could be benecial for the utilization of sh processing by-products

more efciently.

2. . Materials and methods

2.1. Materials

Fresh tilapia skin was provided by Zhanjiang Universal Seafood

Corp. (Zhanjiang, Guangdong, China). Alcalase 3.0 T was purchased

from the Chinese branch of the Danish company Novozyme (Tianjin,

China), the enzyme activity was approximately 200 000 U/g. Angio-

tensin I-converting enzyme (ACE) from rabbit lungs, hippuric acid (HA),

Hippuryl-histidyl-leucine (HHL), L-isoleucine, bacitracin (1422.69 Da),

aprotinin (6511.40 Da), cytochrome C (12 500 Da), carbonic anhydrase

(29 000 Da) and o-pthaldialdehyde (OPA) were obtained from Sigma-

Aldrich (Milwaukee, WI, USA). All other reagents with an analytical

grade were purchased from Sinopharm Chemical Reagent Co., Ltd.

(Shanghai, China).

2.2. Fish skin preparation

The frozen sh skins were thawed, cut into small pieces (about 5 ×5

cm) and then washed with tap water. The pieces were immersed in 0.1%

NaOH (1:10, w/v) at 4 ◦C for 2 h to remove the fat and non-collagenous

proteins. The samples were then washed with tap water for 3 h.

2.3. Steam explosion pretreatment

The pretreatment was carried out on a QBS-200B steam explosion

device with a 5 L chamber from Gentle Science & Technology Co. Ltd.,

China. The apparatus consists of a steam generator, material vessel,

receiver, and rapid-opening piston valve. About 100 g of skin were

placed inside the vessel, treated at 0.2–0.9 MPa for 1 min with saturated

steam and then terminated by explosive decompression within 0.1 s. The

exploded materials were collected and dried at 40 ◦C for 6 h and then

ground with an IKA A11 basic analytical mill (IKA, Staufen, Germany) to

get sh skin protein powder.

2.4. Hydrolysis of samples

Fish skin protein powder (10 g) was resuspended in 1 L of distilled

water. The powder was hydrolyzed by Alcalase with an enzyme-to-

substrate ratio (E:S) of 1:200 (w/w) at the condition of 55 ◦C, pH 8.0

and 125 rpm for 3 h in a rotary thermostatic oscillator. During the hy-

drolysis reaction, the pH was maintained by the periodic addition of 0.1

M NaOH solution. The hydrolysis was ended by heating immediately at

100 ◦C for 10 min to inactivate the protease activity. After cooling, the

solution was centrifuged at 8000 ×g for 10 min. The supernatant was

collected and freeze-dried for further analysis.

2.5. Determination of degree of hydrolysis (DH)

DH was determined by the o-phenylene formaldehyde (OPA) method

(Nielsen, Petersen, & Dambmann, 2001). Briey, 400 µL of hydrolysate

was added to 3 mL of OPA reagent. After 2 min, the UV absorbance of the

mixture at 340 nm was measured. Calculation of free amino acid content

of hydrolysate by L-isoleucine standard curve.

2.6. Determination of peptide content

TCA-soluble peptide content as an indicator for protein degradation

was determined according to Fonkwe and Singh (1996) with some

modications. The sh skin hydrolysates were mixed with 10% (w/v)

TCA (volume ratio of 1:1). After the mixture was reacted at 25 ◦C for 30

min, it was centrifuged at 12000 ×g for 10 min. The content of TCA

soluble peptides in the hydrolysate was determined by the Kjeldahl

method.

2.7. Determination of ACE inhibitory activity

The ACE inhibitory activity was determined according to Wu, Aluko,

and Muir (2002) with some modications. Briey, 40

L sample was

mixed with 25

L ACE solution (100 U/L) in a 1.5 mL centrifuge tube.

After incubation in a 37 ◦C water bath for 10 min, 40

L of the substrate

(6.5 mM HHL in 0.1 M borate buffer, containing 0.3 M NaCl, pH 8.3) was

added into the mixture to initiate the reaction. The reaction was

terminated by the addition of 85

L HCl (1 M) after incubation for 30

min. After ltration through a 0.22

m membrane lter, hippuric acid

(HA) was separated by a Waters e2695 HPLC system (Waters Corp.,

Milford, MA, USA) equipped with a SunFire C18 column (4.6 mm ×250

mm, 5

m). The elution was carried out with 30% (v/v) acetonitrile

containing 0.1% (v/v) TFA at 0.8 mL/min and its absorbance was

monitored at 228 nm. IC

value was dened as the concentration of

peptide required to reduce ACE activity by 50%. ACE inhibitory activity

(%) was calculated as follows:

ACE inhibitory activity (%) = (A−B)/A×100

where A is the HA content of the control (without the sample) and B

is the HA content of the reaction with the sample.

2.8. Determination of molecular weight distribution

Molecular weight distribution of hydrolysates was analyzed on a

TSK-Gel G2000 SW

column (7.8 mm ×300 mm, Tosoh, Tokyo, Japan)

using a Waters e2695 HPLC system. The eluent was 45% acetonitrile

solution containing 0.1% triuoroacetic acid (TFA). The ow rate was

0.5 mL/min. Absorbance was monitored at 220 nm. The standards were

shown as follows: HHL (429 Da), bacitracin (1 422 Da), aprotinin (6 500

Da), and cytochrome C (12 400 Da).

2.9. Gel ltration chromatography

The hydrolysate (20 mg/mL) was ltered through a 0.45

m mem-

brane and then slowly loaded onto an equilibrated Bio-Gel P2 column

Y. Dong et al.

文档加载中……请稍候！
如果长时间未打开，您也可以点击刷新试试。

下载文档到电脑，查找使用更方便

1 知币 4人已下载

立即下载

A novel angiotensin-converting enzyme (ACE) inhibitory peptide from tilapia skin: Preparation, identification and its potential antihypertensive mechanism.pdf

共11页,预览4页

还剩页未读，继续阅读

A novel angiotensin-converting enzyme (ACE) inhibitory peptide from tilapia skin: Preparation, identification and its potential antihypertensive mechanism

相关推荐

开通VIP享超值会员特权

作者详情

相关内容

推荐作者

热门标签

举报选择: