福建柏叶总二萜提取工艺优化及抑菌活性研究

doi:10.13304/j.nykjdb.2024.0391

摘要/Abstract

摘要：

为优化福建柏叶总二萜提取工艺并研究其抑菌活性，通过响应面法对福建柏叶总二萜提取工艺进行优化，并采用菌丝生长速率法探究福建柏叶总二萜对6种植物病原真菌的抑制活性，以胶孢炭疽菌作为供试菌，进一步对菌丝生理生化指标的变化及保护酶活性进行测定。结果表明，福建柏叶总二萜的最佳工艺条件如下：使用无水乙醇作为提取溶剂，提取温度为79 ℃，提取时间为53 min，液料比为14.5 mL∶1 g。在此条件下，福建柏叶总二萜得率为36.8%。福建柏叶总二萜对胶孢炭疽菌抑制效果较好，EC₅₀值为1.466 mg·mL^-1。经福建柏叶总二萜处理后的胶孢炭疽菌菌丝干重显著降低、电导率升高、还原糖含量显著增加、抗氧化酶活性先升高后降低。以上研究结果为福建柏叶的资源利用奠定基础。

关键词: 福建柏, 抑菌活性, EC₅₀, 生理生化, 胶孢炭疽, 响应面

Abstract:

To optimize the extraction process of total diterpene from Fokienia hodginsii leaves and antifungal activity， the response surface method was used， and the antifungal activity of total diterpenoids of Fokienia hodginsii leaves against 6 plant pathogenic fungi was investigated by mycelium growth rate method. Colletotrichum gloeosporioides Panz. was used as the test bacterium， and the changes of physiological and biochemical indexes of mycelia and the activity of protective enzymes were further determined to explore the antifungal activity of total diterpene extracts. The results showed that the optimal process conditions were as follows： anhydrous ethanol was used as the extraction solvent， extraction temperature was 79 ℃， extraction time was 53 min， liquid-solid ratio was 14.5 mL∶1 g. Under this condition， the total diterpene yield was 36.8%. The total diterpenoids of Fokienia hodginsii leaves had a good inhibitory effect on Colletotrichum gloeosporioides Panz.， and theEC₅₀ value was 1.466 mg·mL^-1. The dry weight of mycelia of Colletotrichum gloeosporioides Panz. treated with total diterpenoids was significantly decreased， the conductivity was increased， the content of reducing sugar was significantly increased， and the activities of protective enzymes in the fungi increased first and then decreased. Above results laid a foundation for the resource utilization of Fokienia hodginsii leaves.

Key words: Fokienia hodginsii, antifungal activity, EC₅₀, physiology and biochemistry, Colletotrichum gloeosporioides Penz., response surface

中图分类号:

S791.43

付硕硕, 龚伟勇, 林逸轩, 李渊顺, 刘澄灿, 倪林. 福建柏叶总二萜提取工艺优化及抑菌活性研究[J]. 中国农业科技导报, 2025, 27(5): 146-155.

Shuoshuo FU, Weiyong GONG, Yixuan LIN, Yuanshun LI, Chengcan LIU, Lin NI. Optimization of Extraction Technology and Antifungal Activity of Total Diterpene from Fokienia Hodginsii[J]. Journal of Agricultural Science and Technology, 2025, 27(5): 146-155.

图/表 9

表1 响应面试验因素水平

Table 1 Factor and level of response surface

水平 Level	因素Factor
水平 Level	A：时间 Time/min	B：温度 Temperature/℃	C：液料比 Liquid-solid ratio/（mL·g^-1）
-1	50	75	10∶1
0	60	80	15∶1
1	70	85	20∶1

图1 对照品反式樱柏酸的标准曲线

Fig. 1 Standard curve of trans-communic acid

图2 不同条件下福建柏叶总二萜得率A：提取时间；B：提取温度；C：液料比

Fig. 2 Extraction rate of total diterpene from Fokienia hodginsii under different conditionsA： Extraction time； B： Extraction time； C： Liquid-solid ratio

表2 响应面试验结果

Table 2 Experimental results of response surface test

编号 Serial number	A：时间 Time/min	B：温度 Temperature/℃	C：液料比 Liquid-material/（mL·g^-1）	得率 Yield/%
1	70	85	15∶1	26.28
2	60	80	15∶1	37.20
3	60	80	15∶1	36.66
4	60	80	15∶1	36.18
5	60	75	20∶1	25.20
6	60	75	10∶1	30.72
7	60	85	20∶1	24.60
8	50	75	15∶1	29.82
9	50	80	10∶1	28.68
10	70	75	15∶1	31.08
11	60	80	15∶1	35.94
12	50	80	20∶1	28.68
13	50	85	15∶1	28.56
14	60	80	15∶1	36.72
15	70	80	20∶1	24.78
16	60	85	10∶1	24.00
17	70	80	10∶1	27.90

表3 回归模型与方差分析

Table 3 Regression model and variance analysis

方差来源 Source of variation	平方和 Sum of squares	自由度 df	均方 Mean square	F值 F value	P值 P value	显著性 Significance
模型Model	352.02	9	39.11	84.25	<0.000 1	**
A	4.06	1	4.06	8.75	0.021 2	*
B	22.38	1	22.38	48.20	0.000 2	**
C	8.08	1	8.08	17.40	0.004 2	*
AB	3.13	1	3.13	6.75	0.035 5	*
AC	2.43	1	2.43	5.24	0.055 8
BC	9.36	1	9.36	20.17	0.002 8	*
A²	40.79	1	40.79	87.86	<0.000 1	**
B²	84.98	1	84.98	183.04	<0.000 1	**
C²	147.44	1	147.44	317.57	<0.000 1	**
残差Residual	3.25	7	0.46
失拟Lack of fit	2.28	3	0.76	3.12	0.149 9
纯误差Pure error	0.97	4	0.24
总计Total	355.27	16

图3 各因素间交互作用对福建柏叶总二萜得率的影响

Fig. 3 Effect of interaction among factors on yield of total diterpene from Fokienia hodginsii

图4 福建柏叶总二萜对胶孢炭疽菌的毒力测定

Fig. 4 Determination of the toxicity of total diterpenoids of Fokienia hodginsii to Colletotrichum gloeosporioides Penz.

图5 不同总二萜处理下对胶孢炭疽菌的菌丝干重注：不同小写字母表示不同处理间在P<0.05水平差异显著。

Fig. 5 Dry weight of Colletotrichum gloeosporioides Penz. of different total diterpenoids treatmentsNote：Different lowercase letters indicate significant differences between different treatments at P<0.05 level.

图6 福建柏叶总二萜处理下胶孢炭疽菌的生理生化指标注：不同小写字母表示同一时间不同处理间在P<0.05水平差异显著。

Fig 6 Physiological and biochemical indexs of Colletotrichum gloeosporioides Penz.treated with total diterpene from Fokienia hodginsiiNote：Different lowercase letters indicate significant differences between different treatments of same time at P<0.05 level.

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