超声波辅助酶解牦牛血粉提取氯化血红素的响应面工艺优化及品质表征

doi:10.3969/j.issn.1004-1524.20230926

浙江农业学报 ›› 2024, Vol. 36 ›› Issue (6): 1357-1367.DOI: 10.3969/j.issn.1004-1524.20230926

超声波辅助酶解牦牛血粉提取氯化血红素的响应面工艺优化及品质表征

张晋¹(), 吴晓丽¹^,², 田雨薇³, 赵珂¹, 李欢欢¹, 达色⁴, 次仁达杰⁴, 陈黎洪¹, 唐宏刚¹^,^*()

1.浙江省农业科学院食品科学研究所,省部共建农产品质量安全危害因子与风险防控国家重点实验室,浙江杭州 310021
2.浙江海洋大学食品与药学学院,浙江舟山 316022
3.浙江师范大学行知学院理学院,浙江金华 310018
4.那曲岗亚尔畜牧科技产业有限公司,西藏那曲 852014

收稿日期:2023-07-28 出版日期:2024-06-25 发布日期:2024-07-02
作者简介:张晋(1990—),男,山东聊城人,博士,副研究员,研究方向为动物源产品加工副产物综合利用。E-mail: zhangjin@zaas.ac.cn
通讯作者: *唐宏刚,E-mail: zaastang@163.com
基金资助:
浙江省重点研发计划(2021C04024)

Ultrasound-assisted enzymolytic extraction of chlorohemin from yak blood powder: response surface optimization and quality characterization

ZHANG Jin¹(), WU Xiaoli¹^,², TIAN Yuwei³, ZHAO Ke¹, LI Huanhuan¹, Dase ⁴, Cirendajie ⁴, CHEN Lihong¹, TANG Honggang¹^,^*()

1. State Key Laboratory for Managing Biotic and Chemical Threats to the Quality and Safety of Agro-Products, Institute of Food Science, Zhejiang Academy of Agricultural Sciences, Hangzhou 310021, China
2. College of Food and Pharmacy, Zhejiang Ocean University, Zhoushan 316022, Zhejiang, China
3. School of Science, Xingzhi College Zhejiang Normal University, Jinhua 310018, Zhejiang, China
4. Naqu Gonyal Animal Husbandry Technology and Industry Company, Naqu 852014, Xizang, China

Received:2023-07-28 Online:2024-06-25 Published:2024-07-02

摘要/Abstract

摘要：

以牦牛血粉为原料,通过超声波辅助酶解技术提取氯化血红素,优化制备工艺并进行品质评价。以超声波辅助酶解时间、酶/底物浓度和超声比功率为自变量,以纯度为响应变量,采用Box-Behnken响应面设计优化提取工艺;通过紫外-可见分光光度计、色度仪、傅立叶转换红外光谱仪表征样品的纯度、得率、色度、化学结构等理化性质。结果表明,超声波辅助酶解时间、酶/底物浓度和超声比功率对氯化血红素提取物纯度均具有显著影响(P<0.05);经响应面优化后确定的最佳提取工艺为超声波辅助酶解时间4 h、酶/底物浓度17 U·mg^-1和超声比功率800 W·L^-1,在此条件下氯化血红素提取纯度为14.89%,得率达63.30%。与普通酶解提取工艺相比,超声波辅助酶解提取得率和纯度分别提高1.29倍和1.07倍(P<0.05),提取样品的色度和化学结构与氯化血红素标准品更相近。综上所述,超声波辅助法(800 W·L^-1超声比功率下处理4 h)能在不改变理化性质的前提下显著提高从牦牛血粉中酶解提取氯化血红素的纯度和得率。

关键词: 牦牛血, 超声波辅助酶解, 氯化血红素, 响应面优化, 傅立叶转换红外光谱

Abstract:

The objective of this work was to extract, optimize, and evaluate the chlorohemin from yak blood powder using ultrasonic-assisted enzymolytic technology. The ultrasound-assisted enzymolytic time, enzyme/substrate concentration, and ultrasound specific power were the independent variables, and the purity was the response variable. The Box-Behnken response surface design was also used to optimize the extraction process. Furthermore, the purity, yield, color, and chemical structure of the extracted chlorohemin with response surface optimization were characterized by ultraviolet-visible spectrophotometer, colorimeter, and Fourier transform-infrared spectrometer, respectively. The results showed that ultrasound-assisted enzymolytic time, enzyme/substrate concentration, and ultrasound specific power had significant effects on the purity of chlorohemin extract (P<0.05). The optimal extraction conditions were determined as ultrasound-assisted enzymatic time 4 h, enzyme/substrate concentration 17 U·mg^-1, and ultrasound specific power 800 W·L^-1 after response surface optimization. The purity of extracted chlorohemin under these conditions was 14.89%, the extraction yield was 63.30%. Compared with the chlorohemin extracted through normal enzymolysis (without ultrasound-assisted treatment) under the same conditions, the yield and purity were increased by 1.29 and 1.07 times (P<0.05), respectively. Moreover, there was not significant difference in chemical structure between these two samples. However, the color of extracted chlorohemin with ultrasound-assisted enzymolysis and response surface optimization was closer to the standard substance of chlorohemin. The ultrasound-assisted treatment (4 h at 800 W·L^-1 ultrasound specific power) could significantly improve the purity and yield of chlorohemin extraction from yak blood powder without changing its physicochemical properties.

Key words: yak blood, ultrasound-assisted enzymolysis, chlorohemin, response surface optimization, Fourier transform-infrared spectroscopy

中图分类号:

TS251.93

张晋, 吴晓丽, 田雨薇, 赵珂, 李欢欢, 达色, 次仁达杰, 陈黎洪, 唐宏刚. 超声波辅助酶解牦牛血粉提取氯化血红素的响应面工艺优化及品质表征[J]. 浙江农业学报, 2024, 36(6): 1357-1367.

ZHANG Jin, WU Xiaoli, TIAN Yuwei, ZHAO Ke, LI Huanhuan, Dase , Cirendajie , CHEN Lihong, TANG Honggang. Ultrasound-assisted enzymolytic extraction of chlorohemin from yak blood powder: response surface optimization and quality characterization[J]. Acta Agriculturae Zhejiangensis, 2024, 36(6): 1357-1367.

图/表 11

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自变量 Independent variables	不同水平(编码)的设定值Setting values for different levels (codes)
自变量 Independent variables	低 Low(-1)	中 Medium(0)	高 High(1)
(A)超声波辅助酶解时间Ultrasound-assisted enzymolytic time/h	3.5	4	4.5
(B)酶/底物浓度Enzyme/substrate concentration/(U·mg^-1)	11.25	15	18.75
(C)超声比功率Ultrasound specific power/(W·L^-1)	800	1 000	1 200

自变量 Independent variables	不同水平(编码)的设定值Setting values for different levels (codes)
自变量 Independent variables	低 Low(-1)	中 Medium(0)	高 High(1)
(A)超声波辅助酶解时间Ultrasound-assisted enzymolytic time/h	3.5	4	4.5
(B)酶/底物浓度Enzyme/substrate concentration/(U·mg^-1)	11.25	15	18.75
(C)超声比功率Ultrasound specific power/(W·L^-1)	800	1 000	1 200

序号 Number	自变量组合 Combination of independent variables	纯度Purity/%
序号 Number	自变量组合 Combination of independent variables	响应值 Response value	预测值 Predicted value	相对偏差 Relative deviation
1	A₀B₁C_-1	15.91±0.31 ab	15.92	-0.02
2	A₀B₀C₀	14.78±1.37 abc	14.13	0.65
3	A₁B_-1C₀	8.22±0.11 ghi	6.89	1.33
4	A_-1B_-1C₀	6.24±0.08 i	5.78	0.46
5	A₁B₀C₁	8.79±1.22 fgh	8.87	-0.08
6	A_-1B₁C₀	9.82±0.72 efgh	10.78	-0.96
7	A₀B_-1C₁	10.69±0.54 def	11.04	-0.36
8	A₁B₀C₁	10.52±0.22 efg	11.67	-1.16
9	A₀B_-1C_-1	8.06±1.66 hi	9.50	-1.44
10	A₀B₀C₀	16.15±0.54 a	14.13	2.02
11	A₁B₁C₀	9.91±0.86 efgh	10.00	-0.09
12	A_-1B₀C_-1	13.12±0.91 bcd	12.33	0.78
13	A₀B₁C₁	13.79±2.35 abc	12.73	1.06
14	A_-1B₀C₁	7.58±0.50 hi	7.88	-0.29
15	A₀B₀C₀	12.02±1.50 cde	14.13	-2.11
16	A₀B₀C₀	14.06±1.37 abc	14.13	-0.06
17	A₀B₀C₀	14.36±0.19 abc	14.13	0.24

序号 Number	自变量组合 Combination of independent variables	纯度Purity/%
序号 Number	自变量组合 Combination of independent variables	响应值 Response value	预测值 Predicted value	相对偏差 Relative deviation
1	A₀B₁C_-1	15.91±0.31 ab	15.92	-0.02
2	A₀B₀C₀	14.78±1.37 abc	14.13	0.65
3	A₁B_-1C₀	8.22±0.11 ghi	6.89	1.33
4	A_-1B_-1C₀	6.24±0.08 i	5.78	0.46
5	A₁B₀C₁	8.79±1.22 fgh	8.87	-0.08
6	A_-1B₁C₀	9.82±0.72 efgh	10.78	-0.96
7	A₀B_-1C₁	10.69±0.54 def	11.04	-0.36
8	A₁B₀C₁	10.52±0.22 efg	11.67	-1.16
9	A₀B_-1C_-1	8.06±1.66 hi	9.50	-1.44
10	A₀B₀C₀	16.15±0.54 a	14.13	2.02
11	A₁B₁C₀	9.91±0.86 efgh	10.00	-0.09
12	A_-1B₀C_-1	13.12±0.91 bcd	12.33	0.78
13	A₀B₁C₁	13.79±2.35 abc	12.73	1.06
14	A_-1B₀C₁	7.58±0.50 hi	7.88	-0.29
15	A₀B₀C₀	12.02±1.50 cde	14.13	-2.11
16	A₀B₀C₀	14.06±1.37 abc	14.13	-0.06
17	A₀B₀C₀	14.36±0.19 abc	14.13	0.24

项目 Item	平方和 Sum of squares	自由度 Freedom	均方值 Mean square value	F值 F value	P值 P value
模型Model	129.20	4	32.30	15.35	<0.000 1
残差Residual	25.26	12	2.10	—	—
失拟项Lack of fit	16.34	8	2.04	0.916 1	0.577 5
误差Pure error	8.92	4	2.23	—	—
总和Cor total	154.45	16	—	—	—
	R²=0.836 5	修正Corrective R²=0.782 0	—	—	—

超声波辅助酶解牦牛血粉提取氯化血红素的响应面工艺优化及品质表征

Ultrasound-assisted enzymolytic extraction of chlorohemin from yak blood powder: response surface optimization and quality characterization

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项目 Item	系统系数 System coefficient	平方和 Sum of squares	自由度 Freedom	均方值 Mean square value	F值 F value	P值 P value
B	2.03	32.87	1	32.87	15.62	0.001 9
AC	1.82	13.18	1	13.18	6.62	0.027 8
A²	-3.94	65.44	1	65.44	31.09	<0.000 1
B²	-1.83	14.08	1	14.08	6.69	0.023 8

条件 Conditions	超声辅助酶解时间 Ultrasound-assisted enzymolytic time/h	酶/底物浓度 Enzyme/substrate concentration/(U·mg^-1)	超声比功率 Ultrasound specific power/(W·L^-1)	预测纯度 Predicted purity/%
最佳条件Optimized conditions	3.88	17.08	800.00	12.43
修正条件Modified conditions	4.00	17.00	800.00	12.38

理化指标 Physicochemical indexes		牦牛血粉 Yak blood powder	普通酶解 Enzymolysis	超声波辅助酶解 Ultrasound-assisted enzymolysis	氯化血红素标准品 Standard substance of chlorohemin
纯度Purity/%		3.54±0.03 d	7.21±0.02 c	14.89±0.13 b	95.50±0.80 a
得率Yield/%		—	27.61±0.11 b	63.30±1.32 a	—
粉体色度Powder chromaticity	L^*	43.29±1.46 a	42.72±0.93 a	34.96±0.23 c	39.74±0.51 b
	a^*	5.79±0.79 a	3.51±0.17 b	-0.54±0.13 c	0.04±0.03 c
	b^*	4.38±1.09 a	3.32±0.52 a	-0.72±0.13 b	-2.40±0.04 c
溶液色度Solution chromaticity	L^*	43.48±1.99 a	34.03±0.44 b	33.10±0.40 b	30.52±0.06 c
	a^*	17.17±0.30 a	11.12±1.31 b	7.55±0.98 c	0.15±0.03 d
	b^*	14.84±0.48 a	2.20±0.63 b	0.93±0.68 c	-1.08±0.05 d
红外光谱特征性官能团		—	700~720 cm^-1: Cl^-弯曲振动bending vibration
Characteristic functional groups		1 500~-1 700 cm^-1:—C=O伸缩振动stretching vibration
in FT-IR spectrum		2 360 & 2 900 cm^-1:—C—H弯曲振动bending vibration
		3 200~3500 cm^-1:—O—H伸缩振动stretching vibration