基于UPLC-MS/MS定量检测法优化灵芝三萜热回流提取工艺的研究

基于超高效液相-三重四级杆质谱联用（UPLC-MS/MS）定量检测法优化出一种灵芝三萜热回流提取的工艺。通过本实验建立的灵芝三萜UPLC-MS/MS精确方法检测不同来源的7个栽培品种灵芝子实体中19个三萜化合物的含量,筛选出三萜含量较高的sd-2灵芝子实体作为提取原料,以三萜含量和提取物得率为指标,通过单因素和响应面实验对灵芝三萜的热回流工艺进行优化,获得的最佳条件为：乙醇浓度75%、提取时间2.5h、液料比14:1、提取次数2次。对获得的最佳提取工艺进行中试实验,灵芝子实体乙醇提取物的得率为6.10%,三萜含量为11.8591mg/g。研究结果可为灵芝三萜的工业大规模提取提供理论依据,为灵芝高附加值产品的开发利用提供技术支撑。     

Supercritical Carbon Dioxide Extraction of Allium ursinum: Impact of Temperature and Pressure on the Extracts Chemical Profile

The aim of this study was to extract Allium ursinum L. for the first time by supercritical carbon dioxide (SC−CO₂) as green sustainable method. The impact of temperature in the range from 40 to 60 °C and pressure between 150 and 400 bar on the quality of the obtained extracts and efficiency of the extraction was investigated. The highest extraction yield (3.43 %) was achieved by applying the extraction conditions of 400 bar and 60 °C. The analysis of the extracts was performed by gas chromatography and mass spectrometry (GC/MS). The most dominant sulfur-containing constituent of the extracts was allyl methyl trisulfide with the highest abundance at 350 bar and 50 °C. In addition, the presence of other pharmacologically potent sulfur compounds was recorded including S-methyl methanethiosulfinate, diallyl trisulfide, S-methyl methylthiosulfonate, and dimethyl trisulfide. Multivariate data analysis tool was utilized to investigate distributions of the identified compounds among the extracts obtained under various extraction conditions and yields. It was determined that the SC−CO₂ extraction can by efficiently used for A. ursinum.     

苦丁茶中绿原酸及其异构体的提取变化分析

以富含绿原酸类成分的苦丁茶(Ilex kaushue)为材料,使用溶剂甲醇、乙醇、丙酮和水,结合超声波提取、水浴提取、回流提取等方法对绿原酸及其异构体的提取效率及提取后各异构体的变化进行分析。应用超高效液相色谱法可使苦丁茶的6种绿原酸类成分及咖啡酸在6 min内实现分离。提取结果表明,丙酮作为提取溶剂,在采用超声波和水浴提取时能获得较高的提取效率,易获得较多总量的绿原酸类成分和高含量异绿原酸A。而最常用的溶剂乙醇并未达到理想的提取效果。在不同溶剂的回流提取中,虽然提取的绿原酸类成分总量接近,但异绿原酸A和异绿原酸C含量有较大差异。醇溶液特别是乙醇溶液的回流提取使异绿原酸C的量大幅增加,而相应的异绿原酸A的量大幅减少,表明在醇加热条件下,异绿原酸A转化为异绿原酸C,这为获得抗氧化性更强的异绿原酸C提供了新思路。     

Ultrasound-assisted aqueous extraction of total flavonoids and hydroxytyrosol from olive leaves optimized by response surface methodology

Abstract

Olive leaves were often extracted with methanol or ethanol at different proportions. In this study, ultrasound-assisted aqueous extraction was adopted for olive leaf extraction. The yields of total flavonoids (TF) and hydroxytyrosol (HT) were optimized by central composite experimental design. Two second-order polynomial equations were established to quantify the relationship between the responses and the processing parameters. Under the optimal condition of extracting at 60?°C for 60?min with the solvent-to-material ratio of 40, TF and HT amounted to 57.31?±?0.35 and 1.80?±?0.02?mg/g dry leaves (DL), respectively. The scavenging rate of all extracts against α, α-diphenyl-β-picrylhydrazyl (DPPH) and hydroxyl free radicals was screened. The integrated scores, representing both active ingredients and antioxidant capacity of the extracts, were calculated by principle component analysis (PCA). The optimal extract gained the highest score in PCA. In addition, compared to the extracts from 80% methanol to 44% ethanol, the ultrasound-assisted aqueous extract was richer in TF, HT, and polyphenols, while it also presented stronger ferric reducing antioxidant power (FRAP), but poorer strength to quench hydroxyl radicals. The study indicated that the aqueous extract of olive leaves may present broad potential opportunities in health-care sector.     

Decoupling Analysis of Four Selected Countries

We examine decoupling conditions of domestic extraction of materials, energy use, and sulfur dioxide (SO₂) emissions from gross domestic product (GDP) for two BRIC (Brazil, Russia, India and China) countries (i.e., China and Russia) and two Organisation for Economic Co‐operation and Development (OECD) countries (Japan and the United States) during 2000–2007, using a pair of decoupling indicators for resource use (D_r) and waste emissions (D_e) and the decoupling chart, which can distinguish between absolute decoupling, relative decoupling, and non‐decoupling. We find that (1) during 2000–2007, decoupling between environmental indicators and GDP was higher in the two OECD countries as compared with the two BRIC countries. The key reason is that these countries were in different development stages with different economic growth rates. (2) Changes in environmental policies can significantly influence the degree of decoupling in a country. (3) China, Japan, and the United States were more successful in decoupling SO₂ emissions from GDP than in decoupling material and energy use from GDP. The main reason is that, unlike resource use, waste emissions (e.g., SO₂ emissions) can be reduced by effective end‐of‐pipe treatment. (4) The decoupling indicator is different from the changing rate of resource use and waste emissions. If two countries have different GDP growth rates, even though they may have similar values using the decoupling indicator, they may show different rates of change for resource use and waste emissions.     

MODEL DEVELOPMENT AND PROCESS OPTIMIZATION FOR SOLVENT EXTRACTION OF POLYPHENOLS FROM RED GRAPES USING BOX–BEHNKEN DESIGN

The objective of the present study is to find out the optimum extraction conditions for extraction of polyphenols from red grapes using Box–Behnken design. Red grapes polyphenols were extracted using acid–ethanol solvent at various extraction temperature (40–60°C), extraction time (20–100 min) and different solid–liquid ratio (1:5–1:15 g:ml). The effect (main and interactive) of extraction conditions on total anthocyanin, phenolic and flavonoid content were studied using Box–Behnken design (three factors at three levels). The results showed that the contribution of the quadratic model was significant for all the responses. Second-order mathematical regression models were developed and were found to fit well with observed data. Derringer's desirability function methodology was performed to find out the optimal conditions based on both individual and combinations of all responses (extraction temperature: 57°C, time: 61 min, and solid–liquid ratio: 1:8.7 g:ml) were established. At this optimal condition, the anthocyanin yield, total phenolic and flavonoid content were 73.92 mg/100 g, 221.4 mg GAE/100 g, and 79.08 mg CE/100 g, respectively. A desirability value of 0.902 was achieved at this point.     

Comparison of Rapid DNA Extraction Methods Applied to PCR Identification of Medicinal Mushroom Ganoderma spp.

Abstract

Four different DNA extraction methods were used to extract genomic DNA of the medicinal mushroom Lingzhi from its developing stage materials, such as mycelium, dry fruiting body, or sliced and spore powder or sporoderm‐broken spore powder. The DNA samples were analyzed using agarose gel electrophoresis, UV spectrophotometer, and PCR amplification. According to the average yields and purity of DNA, high salt concentrations and low pH methods were the best for DNA extraction. The mycelia and sporoderm‐broken spore powder yielded higher and purer DNA. The method developed could effectively eliminate the influence of the secondary metabolites to DNA extraction. The DNA samples extracted from the developed method could be successfully used for PCR applications.     

Zone Precipitation Chromatography: Its Use in the Isolation of Different Collagen Types

Zone Precipitation Chromatography is useful tech-nique for the initial isolation of the different colla-gen types in their native configuration. Small quan-tities of collagen mixtures can be rapidly separated into different collagen types with relatively high degree of purity, based upon stained protein patterns on sodium dodecyl sulfate polyacrylamide gel electro-phoresis (SDS-PAGE) slab gels. Tn the commonly used bulk salt preparative method for isolating the different collagens, 50 mg of starting material was needed. Three days were required to complete the procedure. The stained protein patterns on SDS-PAGE slab gels showed about 25% contamination with the bulk purified Type III fraction and 20% contamination with the bulk purified type AB collagen. With Zone Precipitation Chromatography 5 mg of starting material was used and in less than 4 hours the mixture was separated with Types III and AB fractions showing less than 10% contamination from other collagen types. The technique is patterned after the Zone Precinitation method reported by Porath seventeen years ago and utilizes a step-wise sodium chloride gradient to precipitate and redissolve the collagens, eluting from the interbead spaces of a molecular sieve column.     

Can indicator species predict restoration outcomes early in the monitoring process? a case study with peatlands

Success in ecological restoration is rarely assessed rigorously due to insufficient planning for post-restoration monitoring programs, limited funding and, especially, lack of scientifically validated evaluation criteria and protocols. In this article, we propose the use of the Indicator Value Index technique (IndVal), which statistically determines the association of species to one or several particular site types, to obtain indicators of success at the early stages of the recovery process in restoration projects. Peat bogs extracted by the vacuum method, subsequently restored by a moss-transfer technique and regularly monitored for ~10 years were used as a model system to test this approach. We first identified 34 restored sectors of ~10 ha from 4 to 11 years old in twelve eastern-Canadian bogs. These sectors were then classified according to their degree of success in recovering a typical sphagnum moss carpet (restoration goal). Then, we retrospectively reviewed vegetation communities recorded at the third year after restoration to identify indicator species of different categories of restoration success, using the IndVal methodology. By identifying early indicator species, our method provides a tool that guides intervention soon after restoration if a site is not on a desired successional trajectory. Typical bog species, namely the bryophytes S. rubellum and Mylia anomala and the tree Picea mariana, were indicative of successful restoration; while bare peat, lichens and one species of ericaceous shrubs (Empetrum nigrum), which cope better under drier conditions, indicated sites where restoration failed. A surprising finding was that the moss Polytrichum strictum, which is known to facilitate the colonization of sphagnum in disturbed peatlands, is an early indicator of unsuccessful restoration. This finding made us question the nursing role of P. strictum at a cover threshold above ca. 30%, when P. strictum could be outcompeting sphagnum and become dominant. We conclude that the IndVal method is an effective tool to identify early indicators of restoration success when combined with a thoughtful examination of species frequency and cover within each site type.