Separation of polysaccharides from Lycium barbarum L. by high-speed countercurrent chromatography with aqueous two-phase system.

The traditional method for isolation and purification of polysaccharides is time-consuming. It often involves toxic solvents that destroy the function and structure of the polysaccharides, thus limiting in-depth research on the essential active ingredient of Lycium barbarum L. Therefore, in this study, high-speed countercurrent chromatography (HSCCC) and aqueous two-phase system (ATPS) were combined for the separation of crude polysaccharides of Lycium barbarum L. (LBPs). Under the optimized HSCCC conditions of PEG1000-K2HPO4-KH2PO4-H2O (12:10:10:68, w/w), 1.0 g of LBPs-ILs was successfully divided into three fractions (126.0 mg of LBPs-ILs-1, 109.9 mg of LBPs-ILs-2, and 65.4 mg of LBPs-ILs-3). Moreover, ATPS was confirmed as an efficient alternative method of pigment removal for LBPs purification, with significantly better decolorization (97.1 %) than the traditional H2O2 method (88.5 %). Then, the different partitioning behavior of LBPs-ILs in the two-phase system of HSCCC was preliminarily explored, which may be related to the difference in monosaccharide composition of polysaccharides. LBPs-ILs-1 exhibited better hypoglycemic activities than LBPs-ILs-2 and LBPs-ILs-3 in vitro. Therefore, HSCCC, combined with aqueous two-phase system, was an efficient separation and purification method with great potential for separating and purifying active polysaccharides in biological samples.

Recovery and recycling of solvent of counter-current chromatography: The sample of isolation of zeaxanthin in the Lycium barbarum L. fruits.

An efficient method of recovering and recycling solvent for counter-current chromatography was established by which zeaxanthin was separated from Lycium barbarum L. fruits. A column with activated carbon combined with high performance counter-current chromatography formed the recovering and recycling solvent system. Using the solvent system of n-hexane-ethyl acetate-ethanol-water (8:2:7:3, v/v) from the references, five injections were performed with an almost unchanged purity of zeaxanthin (80.9, 81.2, 81.5, 81.3, and 80.2% respectively) in counter-current chromatography separation. Meanwhile, the mobile phase reduced by half than conventional counter-current chromatography. By this present method, an effective improvement of counter-current chromatography solvent utilization was achieved.

Preparative Separation of Flavonoids from Goji Berries by Mixed-Mode Macroporous Adsorption Resins and Effect on Aß-Expressing and Anti-Aging Genes.

Flavonoids are the main constituents of Goji berries and have good biological and pharmacological activities. The mixed-mode macroporous adsorption resins (MARs) for purification of flavonoids from Goji berries through computer-assisted calculation of the molecular size of flavonoids and the precise matching of MAR physical and chemical properties was firstly developed in the present study. Ten varieties of MARs with suitable molecular dimensions and polarities were used for investigating the adsorption/desorption behaviors of the flavonoids. Both AUKJ-1 and BWKX-1 showed higher separation efficiency than other MARs and then were mixed in different ratios to constitute a mixed-mode macroporous adsorption resin to obtain the optimal adsorption phase. Under optimal conditions, total flavonoid content of purified flavonoid (p-FLA) extract increased from 0.97% to 36.88% after one purification. The p-FLA extract from Goji berries significantly improved the expression of six genes with anti-aging effects and played an important role in aging-related Alzheimer's disease by down-regulating Aß expression.

Strategy for the separation of strongly polar antioxidant compounds from Lycium barbarum L. via high-speed counter-current chromatography.

In the separation of strongly polar antioxidant compounds from natural products using high-speed counter-current chromatography that is target-guided by 2,2-diphenyl-1-picrylhydrazyl high-performance liquid chromatography experimentation, low adsorption ability is encountered due to the strong polarity of the target compounds. In this study, a strategy of novel partition coefficient value calculation was proposed for overcoming this problem. The partition coefficient value was expressed as the ratio of the antioxidant activities of the upper phase and the lower phase. This strategy was used in high-speed counter-current chromatography with a hydrophilic organic/salt-containing aqueous two-phase system for bioassay-guided separation of strongly polar antioxidant compounds from Lycium barbarum L. The antioxidant activity was determined via the radical scavenging activity method using 2,2-diphenyl-1-picrylhydrazyl radicals. A hydrophilic organic/salt-containing aqueous two-phase system of 95% EtOH - sat. (NH4)2SO4 (1:1.8, v/v) was successfully used to separate Lycium barbarum L. extract. Five fractions were collected via high-speed counter-current chromatography separation. The antioxidant activity of the third fraction was the highest. Three compounds were separated via MCI gel column chromatography and Sephadex LH-20 column chromatography from the third fraction, and their antioxidant activities were determined. The antioxidant activities of the three compounds were higher than that of the third fraction. These results demonstrate that this strategy can be used to separate strongly polar antioxidant compounds from natural products.

Continuous separation of maslinic and oleanolic acids from olive pulp by high-speed countercurrent chromatography with elution-extrusion mode.

In this work, a continuous high-speed countercurrent chromatography method has been developed on the basis of elution-extrusion mode and this method was successfully applied to the separation of maslinic and oleanolic acid from the extract of olive pulp. In the process of 'elution', the sample solution was continuously loaded into the column and the maslinic acid was steadily eluted out in this step while highly retained oleanlic acid always stayed in the column. In the process of 'extrusion', the oleanlic acid was pushed out of the column with the stationary phase. In this way, we achieved a large sample loading. A total of 120 mL sample solution (about 89.55% of the column volume) which contains 600 mg olive pulp extract was pumped in the apparatus by a constant-flow pump and the maslinic and oleanolic acids were largely separated within 120 min. Both of these two compounds presented high yields and high purities (271.6 mg for maslinic acid with 86.7% and 83.9 mg oleanolic acids with 83.4%).

[Research progress on biological activities of Olea europaea leaf extract].

Olea europaea oil is one of the most important part of the "Mediterranean dietary pattern", and a lot of epidemiological evidences showed that people with the Mediterranean diet having a lower morbidity of the cardiovascular system diseases, skin cancer and colon cancer. The health benefits of a Mediterranean diet not only attributed to monounsaturated fatty acids and vitamins and other nutrients in O. europaea oil, but also the phenolic compounds named as antioxidant effect. Studies have shown that O. europaea leaf contains much more antioxidant activity composition than the fruit, and oleuropein, flavonoids such polyphenols are the main active ingredients in O. europaea leaf. A small amount of O. europaea was introduced into China in 1956, after widely cultivated in Fujian, Guangdong, Taiwan, Sichuan, Shaanxi, Yunnan, and Longnan in Gansu province is the biggest O. europaea planting area in the country. In every winter pruning O. europaea will produce a large number of the leaves, which could be a high added value products (phenolic compounds) of rich source. This article through consulting the literature at home and abroad, classified and summarized the biological activity research status of O. europaea leaf extract and the possible mechanisms, including antimicrobial, antitumor, antioxidation, and on the function of brain, cardiovascular system, anti-diabetes, anti-inflammatory and analgesia and so on. At the same time looked ahead to its development prospects of O. europaea leaf extract, it has variety and high content of active ingredients, and antioxidant synergy, which provide a theoretical basis for the further development and utilization of O. europaea leaf. And O. europaea leaf extract has a rich cheap source and good bioavailability, which provided a broad space in the application of medical and health care.

DPMA, a deoxypodophyllotoxin derivative, induces apoptosis and anti-angiogenesis in non-small cell lung cancer A549 cells.

We found that the deoxypodophyllotoxin derivative, 2,6-dimethoxy-4-(6-oxo-(5R,5aR,6,8,8aR,9-hexahydrofuro[3',4':6,7]naphtho[2,3-d][1,3]dioxol-5-yl)phenyl ((R)-1-amino-4-(methylthio)-1-oxobutan-2-yl)carbamate (DPMA), exhibited superior cytotoxicity compared with etoposide. In this study, we investigated the mechanism of action of DPMA. DPMA exhibited anti-proliferative activity and induced apoptosis in A549 cells in a dose- and time-dependant manner. DPMA inhibited microtubule formation and induced expression of Bax, cleaved caspase-3, p53 and ROS, and inhibited Bcl-2 expression. DPMA also affected cyclinB1, cdc2 and p-cdc2 expression, inducing cell cycle arrest. DPMA also inhibited tube formation of VEGF-induced human umbilical vein endothelial cells. These studies demonstrate that DPMA inhibits p53/cdc2/Bax signaling, thereby inhibiting cell growth/angiogenesis and inducing apoptosis.

[Effects of applying pig manure on lettuce yield and nitrate content and soil nutrients].

A pot experiment with two representative soils (purple soil and yellow soil) in Southwest China was conducted to study the effects of applying pig manure on the lettuce (Lactuca sativa L. var. capitata L. ) yield and nitrate content and the soil nutrients. Applying pig manure increased the lettuce yield significantly, and the increment was higher for yellow soil than for purple soil. The nitrate and total nitrogen, phosphorus, and potassium contents in lettuce plants were closely related to soil type and pig manure application rate. According to the evaluation standards of the nitrate pollution level of vegetables formulated by the Chinese Academy of Agricultural Sciences, the nitrate content in lettuce plants growing on purple soil was lower than the grade I (< or = 432 mg x kg(-1), slight pollution) in treatments CK (no pig manure application) and M1 ( applying 200 kg N x hm(-2) of pig manure), but generally higher than the grade II (< or = 758 mg x kg(-1), moderate pollution) while not exceeded the grade III (< or = 1440 mg x kg(-1), heavy pollution) in other treatments. The nitrate content in lettuce plants growing on yellow soil was lower than the grade I, except that in the treatments of chemical fertilizations and of M8 (applying 1600 kg N x hm(-2) of pig manure) where the plant nitrate content was exceeded the grade II. The critical value of Olsen-P characterizing the apparent leaching risk level of phosphorous in yellow soil and purple soil was 96.3 and 107.7 mg x kg(-1), respectively. The environmental safety capacity of pig manure was higher for yellow soil than for purple soil. Applying pig manure increased the organic carbon and total nitrogen contents of the two soils significantly.

Synthesis and evaluation of the cell cycle arrest and CT DNA interaction properties of 4ß-amino-4'-O-demethyl-4-deoxypodophyllotoxins.

A series of 4ß-amino-4'-O-demethyl-4-deoxypodophyllotoxin derivatives were synthesized, and their cytotoxicities against several human cancer cell lines, including HepG2, A549, HeLa and HCT-8 cells, evaluated. Some of these compounds exhibited higher levels of cytotoxicity than the anticancer drug etoposide. 4ß-N-(4-Nitrophenyl piperazinyl)-4'-O-demethyl-4-deoxypodophyllotoxin (11) was found to be the most potent synthesized compound in the current study, and induced cell cycle arrest in the G2/M phase in HeLa cells, which was accompanied by apoptosis. Furthermore, this compound activated the expression of cdc2, cyclin B1, p53 and caspase-3 in HeLa cells, leading to changes in the conformation of calf thymus DNA from the B-form to a more compact C-form.

Synthesis and biological evaluation of conjugates of deoxypodophyllotoxin and 5-FU as inducer of caspase-3 and -7.

In order to generate compounds with superior antitumor activity and reduced toxicity, a series of conjugates of deoxypodophyllotoxin and 5-FU were synthesized by coupling 4'-demethyl-4-dexoypodophyllotoxin with N-(5-fluorouracil-N(1)-ly acetic)- amino acids (or 5-fluorouracil-N(1)-ly acetic acid). The cytotoxic activity of these compounds against four human cancer cell lines (HL-60, A-549, HeLa and SiHa) were evaluated, and results indicated that these compounds were more potent in terms of cytotoxicity than either parent compound DPT or anticancer drug VP-16 and 5-FU. In addition, we found that 14d induced cell cycle arrest in the G2/M phase accompanied by apoptosis in A-549 cells, and 14d activated caspase-3 and -7. These results suggested that caspase-mediated pathways are involved in 14d induced apoptosis.