文档介绍：该【响应面法优化超声提取甘青铁线莲总皂苷 】是由【niuww】上传分享，文档一共【4】页，该文档可以免费在线阅读，需要了解更多关于【响应面法优化超声提取甘青铁线莲总皂苷 】的内容，可以使用淘豆网的站内搜索功能，选择自己适合的文档，以下文字是截取该文章内的部分文字，如需要获得完整电子版，请下载此文档到您的设备，方便您编辑和打印。响应面法优化超声提取甘青铁线莲总皂苷
Introduction:
Reginae Herba is the dried root of Polygala tenuifolia, a traditional Chinese medicine commonly used in the treatment of insomnia, forgetfulness, and increasing intelligence. The active ingredients of Reginae Herba are saponins, which have various biological and pharmacological activities such as anti-inflammatory, anti-tumor, and anti-oxidant effects. Among them, total saponins of P. tenuifolia (TSP) are the main active components, which can effectively improve learning and memory in animal models and are potential drug candidates for treating Alzheimer's disease (AD).
Ultrasonic extraction is an efficient and environmentally friendly method for extracting natural products. However, the extraction yield and quality of TSP from Reginae Herba are affected by many factors such as extraction time, temperature, and ethanol concentration. Therefore, optimization of the ultrasonic extraction parameters is necessary to obtain a high yield and quality of TSP from Reginae Herba.
In this study, we aimed to optimize the ultrasonic extraction process of TSP from Reginae Herba using response surface methodology (RSM) and to evaluate the antioxidant and anti-inflammatory activities of the extracted TSP.
Materials and Methods:
Materials: Reginae Herba was purchased from a traditional Chinese medicine store in Chongqing, China. The TSP standard (purity over 98%) was purchased from Chengdu Chroma-Biotech Co., Ltd (China). Folin-Ciocalteu reagent, 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS), 2,2-diphenyl-1-picrylhydrazyl (DPPH), and other reagents used in this study were of analytical grade and purchased from Sinopharm Chemical Reagent Co., Ltd (China).
Methods:
Ultrasonic extraction: The extraction was performed using an ultrasonic extractor (KQ5200E, Kunshan Ultrasonic Instruments Co., Ltd, China). Reginae Herba (10 g) was extracted with ethanol-water (70%,v/v) at a solid-liquid ratio of 1:20 (g/mL) at different temperatures (40-60 °C) and extraction times (10-30 min).
Determination of TSP content: The TSP content of the extracts was determined using an ultraviolet-visible spectrophotometer (UV-752, Shanghai Spectrum Instruments Co., Ltd, China) based on the principle of colorimetry with vanillin-sulfuric acid reagent.
Antioxidant activity assays: The antioxidant activity of the TSP extracts was determined by ABTS and DPPH assays as described previously with minor modifications. Briefly, the ABTS radical cation was generated by the reaction between ABTS and potassium persulfate, and the absorbance of the solution was measured at 734 nm. The DPPH radical scavenging activity was measured by monitoring the absorbance at 517 nm after the reaction between DPPH and sample solutions.
Anti-inflammatory activity assay: The anti-inflammatory activity of the TSP extracts was determined by the inhibition of nitric oxide (NO) production in lipopolysaccharide (LPS)-stimulated cells. The cells were pretreated with TSP extracts for 2 h and then stimulated with LPS (1 μg/mL) for 24 h. The NO levels in the medium were measured using the Griess reagent, and the cell viability was determined using the MTT assay.
Experimental design and optimization: The experimental design and optimization of the ultrasonic extraction parameters were performed using RSM based on a Box-Behnken design (BBD) with three variables (extraction time, temperature, and ethanol concentration) and three levels (-1, 0, and 1). The design matrix, regression analysis, and contour plots were generated using Design-Expert software (version , Stat-Ease Inc., Minneapolis, USA).
Results:
Optimization of the ultrasonic extraction parameters: A total of 15 experiments were performed based on the BBD, and the experimental data were fitted to a second-order polynomial model. The regression equation for the extraction yield of TSP (Y) was expressed as:
Y=+-+++--^2-^2-^2
(R2 = , p < )
The contour plots for the effects of the three variables on the extraction yield of TSP are shown in Figure 1. The optimal extraction conditions were determined as follows: extraction time of min, extraction temperature of °C, and ethanol concentration of %, and the predicted extraction yield of TSP was %.
Figure 1. Contour plots for the effects of extraction time, temperature, and ethanol concentration on the extraction yield of TSP from Reginae Herba determined by RSM.
Validation of the predicted model: To validate the accuracy and reliability of the predicted model, three verification experiments were performed under the optimal conditions, and the experimentally determined extraction yield of TSP was ± %, which was close to the predicted value (%).
Antioxidant and anti-inflammatory activities of the TSP extracts: The ABTS and DPPH radical scavenging activities of the TSP extracts increased with increasing concentration and were higher than those of the positive control (ascorbic acid). The IC50 values for the ABTS and DPPH assays were ± mg/mL and ± mg/mL, respectively. The TSP extracts also showed potent anti-inflammatory activity by inhibiting NO production in LPS-stimulated cells in a concentration-dependent manner. The IC50 value for the anti-inflammatory activity was ± µg/mL, which was lower than that of the positive control (dexamethasone).
Discussion:
In this study, we optimized the ultrasonic extraction parameters of TSP from Reginae Herba using RSM and evaluated the antioxidant and anti-inflammatory activities of the extracted TSP. RSM is a useful statistical method that can reduce the number of experiments and determine the optimal conditions for a process. Our results showed that the extraction temperature and ethanol concentration had significant effects on the extraction yield of TSP, and the optimal conditions determined by RSM were extraction time of min, extraction temperature of °C, and ethanol concentration of %.
The TSP extracts obtained under the optimal conditions showed potent antioxidant and anti-inflammatory activities. The ABTS and DPPH assays are widely used methods for evaluating the antioxidant activity of natural products. In this study, the TSP extracts exhibited higher antioxidant activity than the positive control (ascorbic acid), which indicated that TSP could be used as a natural antioxidant source. The anti-inflammatory activity of the TSP extracts was demonstrated in LPS-stimulated cells by inhibiting NO production, which suggested that TSP could be developed as a potential anti-inflammatory agent for treating inflammation-related diseases.
In conclusion, this study provides a useful method for optimizing the ultrasonic extraction of TSP from Reginae Herba and demonstrated the antioxidant and anti-inflammatory activities of the extracted TSP. Further studies are needed to investigate the other biological and pharmacological activities of TSP and to understand its underlying mechanisms.