不同纯度靛蓝提取物的制备及其抑菌活性研究

doi:10.3969/j.issn.0253-2417.2023.01.013

Abstract

Abstract:

On the basis of establishing the analysis method of indigo content, a new and efficient method for the preparation of indigo with high purity was established, and the antibacterial activities of different crude indigo extracts and high purity indigo were studied. Using Baphicacanthus cusia leaves as raw material, three crude extracts samples 1-3 with different indigo mass fractions(3.06%, 20.13%, 22.16%) were obtained by water extraction, stir with alkali, acid neutralization and sodium hydroxide for impurity removal. On this basis, single factor test and Box-Behnken response surface design method were used to optimize glucose reduction process for acquiring high purity of indigo sample 4. Finally, oxford cup method was employed to compare the antibacterial activities of three kinds of crude indigo extracts and high purity indigo against Aspergillus niger, Staphylococcus aureus and Escherichia coli. Results showed that the optimal conditions of glucose reduction were glucose concentration of 25 g/L, sodium hydroxide concentration of 26.9 g/L, reaction temperature of 60 ℃ and reaction time of 30 min. Under the optimum conditions, the purity of indigo could reach 96.87%(sample 4), and the yield of indigo was 76.88%. Results of bacteriostatic experiment demonstrated that the three kinds of indigo crude extracts and high purity indigo had inhibitory effect on A. niger, S. aureus and E. coli, and the inhibitory effects on the three kinds of bacteria increased with the increase of the mass fraction of indigo in the samples. Moreover, the inhibition zone diameter, MBC values, MIC values and IC₅₀ values of these extracts were 9.00-19.69 mm, 1-512 mg/L, 8-64 mg/L(A. niger) and 0.6-448 mg/L, respectively. The high purity indigo had a good inhibitory effect on A. niger, and its MIC, MBC and IC₅₀ values were 1, 8 and 0.6 mg/L, respectively.

Key words: Baphicacanthus cusia(Nees) Bremek, indigo, purification, glucose reduction, response surface method

CLC Number:

TQ35

Yangyang YAN, Changwei ZHANG, Chuan LI, Hao ZHOU, Chengzhang WANG. Preparation of Different Purity Indigo Extracts and Study of Their Antibacterial Activities[J]. Chemistry and Industry of Forest Products, 2023, 43(1): 104-112.

Figures/Tables 6

Fig.1

Table 1

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Table 4

Table 5

References 21

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序号 No.	X₁ 葡萄糖质量浓度/(g·L^-1) glucose mass concn.	X₂ NaOH质量浓度/(g·L^-1) NaOH mass concn.	X₃ 反应温度/℃ reaction temperature	Y₁ 靛蓝质量分数/% indigo mass fraction	Y₂ 靛蓝收率/% indigo yield
1	12.5	26.9	40.00	40.20	20.89
2	25.0	26.9	60.00	96.26	75.21
3	37.5	26.9	40.00	69.32	22.34
4	25.0	26.9	60.00	96.85	76.06
5	12.5	37.5	60.00	55.31	58.77
6	25.0	37.5	40.00	60.31	25.55
7	25.0	37.5	80.00	86.27	78.65
8	12.5	16.3	60.00	55.69	47.65
9	12.5	26.9	80.00	60.11	53.70
10	25.0	16.3	80.00	72.36	56.87
11	37.5	16.3	60.00	80.98	60.75
12	37.5	26.9	80.00	84.17	79.32
13	37.5	37.5	60.00	88.12	77.98
14	25.0	16.3	40.00	64.16	29.45
15	25.0	26.9	60.00	97.01	75.56
16	25.0	26.9	60.00	97.93	75.34
17	25.0	26.9	60.00	96.34	74.98

来源 source	平方和 sum of square	自由度 degree of freedom	均方 mean square	F值 F value	P值 P value
模型model	5 332.69	9	592.52	665.93	< 0.000 1
X₁	1 547.90	1	1 547.90	1 739.69	< 0.000 1
X₂	35.36	1	35.36	39.75	0.000 4
X₃	593.75	1	593.75	667.31	< 0.000 1
X₁X₂	14.14	1	14.14	15.89	0.005 3
X₁X₃	6.40	1	6.40	7.19	0.031 4
X₂X₃	78.85	1	78.85	88.62	< 0.000 1
X₁²	1 229.62	1	1 229.62	1 381.96	< 0.000 1
X₂²	401.41	1	401.41	451.15	< 0.000 1
X₃²	1 124.05	1	1 124.05	1 263.32	< 0.000 1
残差 residual	6.23	7	0.89
失拟项 lack of fit	4.43	3	1.48	3.29	0.140 0
纯误差 pure error	1.80	4	0.45
总误差 cor total	5 338.92	16

样品 sample	样品质量浓度/(g·L^-1) sample mass concn.	靛蓝质量浓度/(g·L^-1) indigo mass concn.	抑菌圈直径 antibacterial circle diameter/mm
样品 sample	样品质量浓度/(g·L^-1) sample mass concn.	靛蓝质量浓度/(g·L^-1) indigo mass concn.	金黄色葡萄球菌 S. aureus	大肠杆菌 E. coli	黑曲霉 A. niger
DMSO	—	—	10.71±0.13	8.00±0.15	8.80±0.19
样品1 sample 1	2.0	0.06	13.57±0.16	10.67±0.17	11.19±0.09
	1.0	0.03	10.97±0.14	10.33±0.13	9.00±0.21
	0.5	0.015	11.35±0.31	10.17±0.22	9.00±0.08
样品2 sample 2	2.0	0.40	14.57±0.16	12.67±0.17	14.19±0.09
	1.0	0.20	12.47±0.14	11.36±0.13	12.50±0.21
	0.5	0.10	10.89±0.31	10.67±0.22	9.78±0.08
样品3 sample 3	2.0	0.44	14.02±0.26	12.86±0.13	14.40±0.15
	1.0	0.22	11.16±0.31	11.33±0.29	13.13±0.13
	0.5	0.11	10.85±0.15	9.96±0.08	9.89±0.06
样品4 sample 4	2.0	1.93	17.85±0.26	13.91±0.25	19.69±0.23
	1.0	0.96	17.14±0.15	12.98±0.16	15.33±0.18
	0.5	0.48	13.54±0.14	11.94±0.06	14.80±0.08
纳他霉素 natamycin	0.2	—	—	—	20.00±0.21
庆大霉素 gentamicin	0.1	—	27.08±0.28	13.71±0.14	—

样品 sample	最小抑菌浓度MIC/(mg·L^-1)			最小杀菌浓度MBC/(mg·L^-1)
样品 sample	金黄色葡萄球菌 S. aureus	大肠杆菌 E. coli	黑曲霉 A. niger	金黄色葡萄球菌 S. aureus	大肠杆菌 E. coli	黑曲霉 A. niger
1	512	512	2	—	—	64
2	512	512	1	—	—	32
3	512	512	1	—	—	32
4	256	256	1	—	—	8

样品 sample	半数抑菌浓度IC₅₀/(mg·L^-1)
样品 sample	金黄色葡萄球菌 S. aureus	大肠杆菌 E. coli	黑曲霉 A. niger
1	384	448	1.8
2	320	384	0.8
3	320	384	0.8
4	160	192	0.6

Preparation of Different Purity Indigo Extracts and Study of Their Antibacterial Activities

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