Rapid Isolation and Hypoglycemic Activity of Secondary Metabolites of Eurotium cristatum by High-Speed Countercurrent Chromatography.

In this study, secondary metabolites of Eurotium cristatum were isolated and purified by high-speed counter-current chromatography (HSCCC), and their hypoglycemic activities were studied. The general-useful estimate of solvent systems (GUESS) for counter-current chromatography was employed to select the appropriate solvent systems of n-hexane-ethyl acetate-methanol-water (HEMW, 4:6:5:5, v/v/v/v) for HSCCC practice, and three compounds were separated from the crude ethyl acetate extract of E. cristatum in one single step; 6.1 mg of Compounds 1, 5.6 mg of Compound 2 and 3.8 mg of Compound 3 were obtained from 100 mg of crude extract with a stationary phase retention of 75%. The compounds were then identified as emodin methyl ether, chrysophanol and emodin, respectively. The activity of the target compounds in the secondary metabolites of E. cristatum was verified by testing their inhibition on α-glucosidase activity and molecular docking simulation. The results showed that emodin, chrysophanol and emodin methyl ether had significant inhibitory effects on the α-glucosidase activity. This work confirmed the effectiveness of HSCCC in the separation of compounds in complex extracts and provided reference for further research and application of E. cristatum.

A simple and practical solvent system selection strategy for high-speed countercurrent chromatography based on the HPLC polarity parameter model.

The selection of an appropriate solvent system is the most crucial step in high-speed countercurrent chromatography (HSCCC) separation. The compound polarity plays an important role in HPLC analysis and HSCCC separation, and it can be calculated by the HPLC polarity parameter model and the average polarity of the HSCCC solvent system, respectively. However, flow rates, columns and methanol concentrations of the HPLC experiment can influence the calculation of the compound polarity. Therefore, the applicability and accuracy of the HPLC polarity parameter model still needed to be extensively validated. We chose 14 compounds to conduct the shake-flask experiments and HPLC analysis on, such as apigenin, honokiol, phloridzin and dihydromyricetin. The HPLC analysis results showed that different flow rates and columns have negligible effects on the calculated compound polarities. However, there was a certain variation trend in the calculated polarities with different methanol concentrations. Although the polarity values of some compounds showed a difference between the HPLC analysis and shake-flask experiments, their partition coefficients (K) in the HSCCC solvent systems were still located in the range of 0.5 < K < 2.0. Guided by the HPLC polarity parameter model, the appropriate HSCCC solvent systems for mangosteen peel and Hypericum sampsonii Hance were selected, and the two main components (mangostin and quercetin) were isolated from their extracts, respectively. The separation results showed that the predicted compound polarities were sufficient to meet the HSCCC separation requirements. Meanwhile, this method required only 1 to 2 HPLC analyses with reference compounds, greatly improved the efficiency of the HSCCC solvent system selection, and shortened the experimental time. The polarity parameter model was a fast and efficient analysis method for the selection of an appropriate HSCCC solvent system.

[Association of neonatal blood calcium levels with perinatal factors and neonatal urinary calcium levels measured by an intelligent urine test system].

OBJECTIVE: To study the association of neonatal blood calcium levels with perinatal factors and neonatal urinary calcium levels measured by an intelligent urine test system. METHODS: The medical data of 96 full-term singleton neonates with mild diseases were collected by a cross-sectional survey, who were hospitalized in the Department of Neonatology, The First Affiliated Hospital of Xi'an Jiaotong University, from June to August 2018. Urinary calcium levels measured by an intelligent urine test system, total blood calcium levels, ionized calcium levels, and the mother's calcium and vitamin D supplementation during pregnancy were recorded. RESULTS: Compared with the group without vitamin D supplementation for the mother (17 neonates), the group with vitamin D supplementation for the mother (79 neonates) had significantly higher levels of total blood calcium and ionized calcium (P < 0.05).The group with both vitamin D and calcium supplementation for the mother (68 neonates) had significantly higher levels of ionized calcium than controls (28 neonate) (P=0.05). There was no significant difference in the levels of total blood calcium and ionized calcium between the group with calcium supplementation for the mother (74 neonates) and the group without calcium supplementation for the mother (22 neonates) (P > 0.05). The hypothermia group (5 neonates) had a significantly lower level of total blood calcium than the normal body temperature group (91 neonates) (P < 0.05). There was a significantly positive correlation between the maternal blood total calcium level and the neonatal blood total calcium and ionized calcium levels (r=0.881 and 0.703 respectively; P < 0.05). The neonatal urinary calcium level measured by the intelligent urine test system was significantly correlated with the blood ionized calcium level (r=0.526, P=0.025). CONCLUSIONS: Vitamin D supplementation during pregnancy can increase the blood levels of total calcium and ionized calcium in neonates, and calcium supplementation alone cannot increase the blood levels of total calcium or ionized calcium in neonates. Hypothermia in neonates might cause the reduction in blood calcium levels. The urinary calcium level measured by the intelligent urine test system is positively correlated with the blood level of ionized calcium.

Ultrasonic-Assisted Aqueous Two-Phase Extraction and Properties of Water-Soluble Polysaccharides from Malus hupehensis.

Malus hupehensis (M. hupehensis), an edible and medicinal plant with significant antioxidant and hypoglycemic activity, has been applied to new resource foods. However, the structural characterization and biological effects of its polysaccharides (MHP) are less known. The optimum extraction parameters to achieve the highest extraction efficiency (47.63%), the yield (1.68%) and purity of MHP (89.6%) by ultrasonic-assisted aqueous two-phase system (ATPS) were obtained under the liquid-to-solid ratio of 23 g/mL, ultrasonic power of 65 W, and ultrasonic time of 33 min. According to the analysis results, MHP was composed of Man, GlcA, Rha, GalA, Glc, Gal, Xyl, Ara, and Fuc, in which Ara and Gal were the main components, and the content of GlcA was the lowest. In in vitro activity analysis, MHP showed a significant antioxidant capacity, and an inhibition activity of α-glucosidase and the advanced glycation end products (AGEs) formation in the BSA/Glc reaction model. MHP interacted with α-glucosidase and changed the internal microenvironment of the enzyme, and inhibited the AGEs formation, which provides more evidence for the antihyperglycemic mechanism of MHP. The results suggest that ATPS is an efficient and environmentally friendly solvent system, and M. hupehensis has broad application prospects in functional foods, healthcare products, and pharmaceuticals.

Extraction and separation of flavonoids from Malus hupehensis using high-speed countercurrent chromatography based on deep eutectic solvent.

In the present experiment, a green and highly efficient extraction method for flavonoids established on deep eutectic solvents (DESs) was investigated by using the response surface methodology. The DES-based high-speed countercurrent chromatography (HSCCC) solvent systems were developed for the separation of high purity compounds from the DES extract of Malus hupehensis for the first time. Under the optimal conditions (liquid-to-solid ratio of 26.3 mL/g, water content of 25.5%, and extraction temperature of 77.5°C), the yield of flavonoids was 15.3 ± 0.1%, which was superior to that of the methanol extraction method. In accordance with the physical property of DES-based HSCCC solvent systems and K values of target compounds, DES-based HSCCC solvent systems composed of choline chloride/glucose-water-ethyl acetate (ChCl/Glu-H2O-EAC, 1:1:2, v/v) was selected for the HSCCC separation. Thus, five flavonoids (two novel compounds 1-2, 6´´-O-coumaroyl-2´-O-glucopyranosylphloretin and 3´´´-methoxy-6´´-O-feruloy-2´-O-glucopyranosylphloretin; three know compounds 3-5, namely, avicularin, phloridzin, and sieboldin) were efficiently separated from Malus hupehensis. DESs are the environment friendly and highly efficient solvents as the components of extraction solvent and HSCCC solvent system, and can be re-utilized many times. However, ethyl acetate can be soluble with a few hydrogen bond donors, such as urea, carboxylic acid and polyol, through the shake flask test. It is the great difficulty for the efficient and rapid separation of target compounds from the DESs extract because of the DESs residual in the HSCCC fractions. ChCl and Glu are the great choices of DESs without this problem. In addition, K values increased with the increase of the molar ratio of ChCl/Glu and the content of water, which could effectively guide us to choose the suitable DES-based HSCCC solvent system. The twice HSCCC separation results indicated that DES was the valuable and green solvent for the HSCCC separation of pure compounds from the extract for the first time, and showed the recycle superiority of DES-based HSCCC solvent system.

Separation of six antioxidants from Hypsizygus marmoreus by high-speed countercurrent chromatography utilizing an approach based upon the polarity parameter model.

For a successful high-speed countercurrent chromatography (HSCCC) separation of multiple components, the suitable solvent system selection is a critical operation. Despite the difference in separation mechanism between HSCCC and HPLC, K values of compounds in the solvent system and the retention factor of compounds in HPLC were connected with polarity, and the polarity mainly depended on the structure and chemical properties of compounds. On the basis of the concept of "like dissolves like", the average polarity of the solvent system is equal to the average polarity of multiple components at the "sweet point" log K = 0. The result of theoretical deduction showed a negative linear correlation between the polarity of each component and the logarithm of the gradient range of the organic phase. Therefore, the average polarity of unknown multiple components could be obtained through the polarity parameter model established in the HPLC analysis. The suitable solvent system composed of n-hexane-ethyl acetate-methanol-water (8:3:8:1, v/v) was selected through the average polarity of multi-components combining K values of the enriched samples. In the context of bioassay, an efficient HSCCC separation procedure was established, and two groups of analogue benzaldehyde derivatives (four main antioxidants, namely, isodihydroauroglaucin, isoaspergin, isotetrahydro-auroglaucin, and flavoglaucin; two minor antioxidants, namely, 6'-oxo-chaetopyranin and chaetopyranin) were obtained from Hypsizygus marmoreus. The predicted polarity values of multi-components were sufficient to meet the HSCCC experimental requirements. The HPLC analysis of reference compounds and multi-components showed a significant consistency with different chromatographic columns. Therefore, the polarity parameter model established in the HPLC analysis was a simple, rapid, and helpful tool for looking an appropriate solvent system, which was a forwarding step for the HSCCC separation of multiple components.

Involvement of abscisic acid and salicylic acid in signal cascade regulating bacterial endophyte-induced volatile oil biosynthesis in plantlets of Atractylodes lancea.

The enormous biological diversity of endophytes, coupled with their potential to enhance the production of bioactive metabolites in plants, has driven research efforts focusing on endophytes. However, limited information is available on the impacts of bacterial endophytes on plant secondary metabolism and signaling pathways involved. This work showed that an endophytic Acinetobacter sp. ALEB16, capable of activating accumulation of plant volatile oils, also induced abscisic acid (ABA) and salicylic acid (SA) production in Atractylodes lancea. Pre-treatment of plantlets with biosynthetic inhibitors of ABA or SA blocked the bacterium-induced volatile production. ABA inhibitors suppressed not only the bacterium-induced volatile accumulation but also the induced ABA and SA generation; nevertheless, SA inhibitors did not significantly inhibit the induced ABA biosynthesis, implying that SA acted downstream of ABA production. These results were confirmed by observations that exogenous ABA and SA reversed the inhibition of bacterium-induced volatile accumulation by inhibitors. Transcriptional activities of genes in sesquiterpenoid biosynthesis also increased significantly with bacterium, ABA and SA treatments. Mevalonate pathway proved to be the main source of isopentenyldiphosphate for bacterium-induced sesquiterpenoids, as assessed in experiments using specific terpene biosynthesis inhibitors. These results suggest that Acinetobacter sp. acts as an endophytic elicitor to stimulate volatile biosynthesis of A. lancea via an ABA/SA-dependent pathway, thereby yielding additional insight into the interconnection between ABA and SA in biosynthesis-related signaling pathways.

[Meta-analysis on acupuncture for treatment of dementia].

OBJECTIVE: To assess the therapeutic effect of acupuncture on dementia. METHODS: The literatures of acupuncture for treatment of dementia are comprehensively searched in according with the demands of the evidence-based medicine (EBM), which are collected from relevant domestic medical literature databases in the last ten years. Meta-analysis is conducted on the literatures enrolled. RESULTS: Twenty-two randomized controlled trials are included, among them, 19 trials are carried out by Meta-analysis. The total OR is 3.72 [2.73, 5.07], and the funnel plot is approximately symmetry. It is indicated that the curative effect of acupuncture groups is better than the control groups (Z = 8.32, P < 0.00001). CONCLUSION: Acupuncture therapy is effective on dementia according to the domestic clinical literatures. However, the quality of the studies needs further improving and increasing.

Meta-analysis on acupuncture for treatment of dementia / 中国针灸

<p><b>OBJECTIVE</b>To assess the therapeutic effect of acupuncture on dementia.</p><p><b>METHODS</b>The literatures of acupuncture for treatment of dementia are comprehensively searched in according with the demands of the evidence-based medicine (EBM), which are collected from relevant domestic medical literature databases in the last ten years. Meta-analysis is conducted on the literatures enrolled.</p><p><b>RESULTS</b>Twenty-two randomized controlled trials are included, among them, 19 trials are carried out by Meta-analysis. The total OR is 3.72 [2.73, 5.07], and the funnel plot is approximately symmetry. It is indicated that the curative effect of acupuncture groups is better than the control groups (Z = 8.32, P < 0.00001).</p><p><b>CONCLUSION</b>Acupuncture therapy is effective on dementia according to the domestic clinical literatures. However, the quality of the studies needs further improving and increasing.</p>