OBJECTIVE

To realize quadratic formula optimization of Renshen Jianxin Capsule (RJC) by screening Chinese herbs with major anti-myocardial ischemia effect in RJC and optimize their optimal dosages.

METHODS

By following "uniform design-pharmacodynamic experiment-mathematical modeling-formula optimization", authors employed U10(10(8)) uniform design in the experiment. Eight Chinese herbs contained in RJC were taken as observatory factors. Electrocardiograph (ECG) changes of myocardial ischemia induced by isoproterenol were taken as pharmacodynamic indices. The mathematical model between herbal factors and pharmacodynamic indices was established using stepwise regression analysis to screen Chinese herbs with major anti-myocardial ischemia effect. Their optimal dosages were optimized using the grid algorithm.

RESULTS

The regression equation was y =1. 7889 -0. 3247 Ginseng xSalvia Miltiorrhiza -0. 0663 Astragalus membranaceus xOriental Waterplantain tuber. Forecasting factors included were Ginseng, Salvia Miltiorrhiza, Astragalus membranaceus, and Oriental Waterplantain tuber. The optimal formula dosage calculated by the grid algorithm was Ginseng 1. 62 g, Astragalus membranaceus 4. 62 g, Salvia Miltiorrhiza 2. 43 g, and Oriental Waterplantain tuber 1. 66 g.

CONCLUSIONS

Uniform design combined with stepwise regression analysis and grid algorithm were able to realize quadratic formula optimization of RJC.

Background: QishenYiqi dripping pills (QSYQ), consisting of Salvia miltiorrhiza, Astragalus membranaceus, Panax notoginseng, and Dalbergia odorifera, is a widely used to treat ischemic cerebrovascular and hemorrhagic cerebrovascular in China. Objective: In this study, a rapid and sensitive ultra-HPLC coupled with a triple quadrupole MS (UHPLC-QQQ-MS) method was established for the simultaneous quantification of 19 bioactive components. Methods: Chromatographic separation was performed on an Agilent Eclipse Plus C18 Rapid Resolution High Definition (RRHD) column (2.1 × 50 mm, 1.8 μm) with a mobile phase composed of a 0.1% (v/v) formic acid aqueous solution and acetonitrile in a gradient elution manner. A QQQ-MS was used in positive and negative ionization mode with multiple reactions monitoring for quantitative analysis. Results: The established methods were validated with linearity, precision, repeatability, stability, recovery, and matrix effect. All calibration curves exhibited excellent linear relationship with the correlation coefficient (r) >0.9985 for each analyte within the test range. This method was successfully applied to quantify target compounds of different structural types in 12 batches of QSYQ. Conclusions: The results suggest that the established method is rapid, sensitive, and reliable. The developed UHPLC-QQQ-MS method can provide scientific basis and reference to quality control and consistency evaluation of QSYQ sample and other formulations. Highlights: A rapid, sensitive, and reliable UHPLC-QQQ-MS method has been developed and validated for simultaneous determination of 19 compounds including phenolic acids, saponins, isoflavonoids, and tanshinones in QSYQ for the first time.

Introduction Sequence - independent microarrays have never been used to identify and authenticate dried herbal plants.

METHODS

We report successful fingerprinting of seven species: Leonurus sibiricus, Astragalus membranaceus, Coix lachryma-jobi, Magnolia biondii, Abutilon theophrasti, Physalis alkekengi, and Salvia miltiorrhiza from dried tissues using a sequence-independent microarray, "Subtracted Diversity Array".

RESULTS

Herbal plants could be identified from tissues as they were sold at the clinic. Hierarchical cluster of these species generated using SPSS v.15.0 confirmed to their predicted taxonomical relationships as specified in the Angiosperm Phylogeny Group II classification system. A polymorphism rate of 40.7% was achieved from the 376 spots used for fingerprinting. Functional characterization of polymorphic features by sequencing revealed 27.1% of those were retroelements or genes.

CONCLUSIONS

This technique provides a new way to produce markers for authenticating dried herbal samples.

As one of the most toxic chemical carcinogens, aflatoxin B1 (AFB1) has attracted extensive attention due to its severe impairment to human health. There exists urgent demand to develop facile and sensitive method for rapid screening of AFB1. Here magnetic beads modified with mouse monoclonal antibody (McAb) were adopted for capture and enrichment of the mycotoxin in sample matrix. Then UV radiation at 365 nm was utilized to induce the enhancement of fluorescent (FL) emission of the captured AFB1 with an addition reaction. The FL signal of the derivative at 435 nm was collected to quantify AFB1. The immunoassay method for AFB1 showed a wide detection range of 1.0-1000 ng mL^-1, with a low detection limit of 0.21 ng mL^-1 (3σ). It was applied to detect AFB1 in herbal medicines including Astragalus membranaceus and Salvia Miltiorrhiza, with acceptable recovery values of 95.4-107.7%. It shows many merits including facile manipulation, low cost, high sensitivity and ideal selectivity. Due to its simple detection mechanism, the UV-induced FL derivatization-based label-free immunoassay can be furtherly extended to detection of other mycotoxins with similar chemical structures.

This paper reports the clinical treatment of 100 patients with minimal brain dysfunction (MBD), their age ranged from 7-14.2 years, and the average age was 10.5. Patients were divided into TCM and WM group randomly. The TCM group (80 cases) were treated with the therapy of subdueing hyperactivity of the Liver and invigorating the function of the Spleen by using: Bupleurum chinense, Scutellaria baicalensis, Astragalus membranaceus, Codonopsis pilosula, Ligustrum lucidum, Lophatherum gracile and thread of ivory. The WM group (20 cases) were treated with retalin 5-15 mg twice daily. One course of treatment lasted one month, and effects were evaluated after 1-3 courses of treatment. The results were as follows: In the TCM group, 23 cases were cured (clinical symptom and sign disappeared, 10 IQ units were raised in their intelligence, abnormal electroencephalogram was recovered and there was no recurrence during the first six months after recovery). 46 cases were improved (clinical symptom and sign markedly improved, 4 IQ units raised in intelligence and electroencephalogram improved), and 11 cases ineffective. The effective rate was 86.25%. In the WM group, 6 cases were cured, 12 cases improved and 2 cases ineffective, the clinical effective rate being 90.0%. There was no significant difference between the two groups in this study, but, the side-effects of the TCM group was less than the WM group, and the TCM group had more beneficial effects to improve intelligence, enuresis and the black of orbits. The study showed that the Verbal Scale in Wechsler Intelligence Scale for children is one of the indications of diagnosis and evaluating of the curative effect of MBD.

Background and aim: Astragalus membranaceus (AM) is a major Chinese herb used in the treatment of stroke. Astragaloside IV (AS)is a component of AM. This study investigated the effects of AM on the protein expression through proteomics analysis in ischemia-reperfusion injured Sprague Dawley rats.

Experimental procedure: An animal model of ischemia-reperfusion injury by occlusion of the right middle cerebral artery for 90 min followed by reperfusion for 24 h. The rats were intraperitoneally injected with AM or AS three times at 30 min, 1 day, and 2 days prior to the occlusion of the cerebral blood flow.

Results: Aldolase C was overexpressed in the cortex, and Dihydrolipoamide dehydrogenase and Triose-phosphate isomerase were overexpressed in the hippocampus.

Conclusion: Pretreatment with AM or AS can induce the overexpression of Aldolase C in the cerebral cortex and that of Dihydrolipoamide dehydrogenase and Triose-phosphate isomerase in the hippocampus, suggesting that both AM and AS may act as neuroprotectors through regulating the expression of Aldolase C, Dihydrolipoamide dehydrogenase and Triose-phosphate isomerase. However, the underlying neuroprotective mechanisms need more studies.

Keywords: Astragaloside IV; Astragalus membranaceus; Brain tissue; Ischemia-reperfusion injury; Protein biomarker.

Ethnopharmacological significance: Ginseng quinquefolium (L.), Astragalus membranaceus, and Sophora flavescens Aiton are popular folk medicines in many Asian countries and regions. These three traditional Chinese herbs and their extracts have been reported to considerably enhance the immune function. G. quinquefolium (L.) is considered the king of herbs in China. Traditionally, G. quinquefolium (L.) is believed to replenish vitality, which is considered as immune enhancement in modern Chinese pharmacy. One of the main uses of Astragalus is immunity enhancement; S. flavescens and oxymatrine obtained from its extract have been used to treat leukopenia. Considering the pharmacological properties of Ginseng, Astragalus, and oxymatrine, we evaluated the immunopotentiation effects of their combination, Ginseng-Astragalus-oxymatrine (GAO), in the present study.

Aim of the study: This study aimed to expand the clinical application of GAO and to preliminarily explore its mechanism of action by determining whether GAO injection can enhance immunity in vivo and in vitro.

Methods: Overall, 17 major chemical components in GAO were analysed using HPLC and LC-MS. The immunity-enhancing effect of GAO was studied in the cyclophosphamide (CTX)-induced immunosuppressive mouse model and RAW 264.7 cells.

Results: Quantitative analysis showed that the potential active components of GAO include at least ginsenosides, astragaloside IV, and oxymatrine. GAO could significantly improve the nonspecific immunity including the indices of the thymus and spleen, number of peripheral blood leukocytes, levels of TNF-α and IL-6, phagocytic function of macrophages, and cytotoxic activity of natural killer (NK) cells. Additionally, GAO enhanced the humoural immunity, characterised by the antibody production ability of B cells, and cellular immunity, characterised by the activity of T cells, in immunosuppressed mouse. Moreover, GAO could enhance the phagocytic and adhesion functions of RAW 264.7 cells, which may be related to the activation of reactive oxygen species and NF-κB signalling pathway.

Conclusion: GAO could dramatically ameliorate CTX-induced immunosuppression in mouse and stimulate the immune activity in RAW 264.7 cells possibly by activating the NF-κB signalling pathway.

Keywords: Ginseng–astragalus–oxymatrine injection; Immunosuppression; NF-κB; cyclophosphamide.

Astragalus membranaceus (A. membranaceus) is a widely used traditional herb in China and Korea. A. membranaceus polysaccharides (AMP), which make up a major part of the root extract, have been shown to modulate immune modulations, especially activation of bone marrow-derived dendritic cells (BMDCs) and T cells. However, the immune stimulatory effect of AMP in the mouse in vivo and human peripheral blood DCs (PBDCs) has not been well investigated. In this study, we found that intravenous (i.v.) injection of AMP in C57BL/6 mice induced remarkable elevations in co-stimulatory and MHC class I and II molecule levels in the splenic DCs and its subsets. The stimulatory effect of DCs by AMP was elevated 6 h after treatment, which rapidly decreased 18 h after injection. Furthermore, AMP promoted intracellular production of pro-inflammatory cytokines in spleen DC subsets, which contributed elevation of serum cytokine levels. Finally, the AMP promoted PBDC activation. Thus, these results demonstrate that AMP can be used as an immune stimulatory molecules in human and mouse.

Keywords: Astragalus membranaceus polysaccharide; Human peripheral blood dendritic cell; Spleen dendritic cell.

Astragalus polysaccharides (APS), which is the main effective component of Astragalus membranaceus, can inhibit tumor growth, but the underlying mechanisms are still unclear. Previous studies suggested that APS is a kind of herbal polysaccharide that can regulate the gut microenvironment, including the gut microbiota and fecal metabolites. In this work, our results showed that APS could control tumor growth in melanoma-bearing mice. It could reduce the number of myeloid-derived suppressor cells (MDSCs), as well as the expression of MDSCs related molecule Arg-1 and cytokines IL-10 and TGF-β, so that CD8⁺ T cells could kill tumor cells more effectively. However, while APS was administrated with antibiotic cocktail (ABX), MDSCs could not be reduced, and the growth rate of tumor was accelerated. Consistent with the changes in MDSCs, the serum level of IL-6, IL-1β was the lowest in the APS group. Meanwhile, we found that fecal suspension from mice in APS group could also reduce the number of MDSCs in tumor tissues. These results revealed that APS regulated the immune function in tumor-bearing mice via remodeling the gut microbiota. Next, we focused on the results of 16S rRNA, and it showed that APS significantly regulated the abundance of microorganisms such as Bifidobacterium_pseudolongum, Lactobacillus_johnsonii and Lactobacillus. According to the Spearman analysis, the changes of abundance of these microorganisms were related to the increase of metabolites like glutamate and creatine, which could control tumor growth. Taken together, this study demonstrates that APS attenuates the immunosuppressive activity of MDSCs in melanoma-bearing mice by remodeling the gut microbiota and fecal metabolites, and our findings will provide a new therapeutic strategy to control tumor growth.

Keywords: Astragalus polysaccharides; Gut microenvironment; Immunosuppressive activity; Melanoma; Myeloid-derived suppressor cells.

Spontaneous senile osteoporosis severely threatens the health of the senior population which has emerged as a severe issue for society. A SAMP6 mouse model was utilized to estimate the impact of intragastrically administered Astragalus Membranaceus (AR) on spontaneous senile osteoporosis. Bone mineral density (BMD) and bone microstructure were measured using Micro-CT; contents of calcium and phosphorus were determined with the colorimetric method; and gene and protein expressions of fibroblast growth factor 23 (FGF23), Klotho, Vitamin D receptor (VDR), CYP27B1 and CYP24A1 were detected using qPCR, Western blot and ELISA assays, respectively. The findings indicated that AR could improve the femoral BMD and bone microstructure, elevate the contents of calcium and phosphorus, and increase the expression of Klotho, VDR, and CYP27B1 whereas decreasing the expression of FGF23 and CYP24A1 in SAMP6 mice in a dose independent manner. The present study has demonstrated that AR can promote osteogenesis and alleviate osteoporosis. It is also expected to provide a new insight for the treatment of spontaneous senile osteoporosis and to serve as a research basis for AR application.

Keywords: Astragalus membranaceus; SAMP6; VD; spontaneous senile osteoporosis.

Ferroptosis, as an iron-dependent programmed cell death pathway, can induce a variety of cardiovascular diseases. Astragaloside IV (AS-IV), which is purified from Astragalus membranaceus, can protect endothelial function and promote vascular regeneration. However, the role played by AS-IV in ferroptosis remains unknown. In this study, the lipid metabolomics in HUVECs treated with/without bleomycin and/or AS-IV were explored using LC/MS. The most differential metabolite between groups was further identified via GO and pathway enrichment analyses. The effects of lysophosphatidylcholine (LPC), AS-IV, and FIN56 on cell viability were explored using the CCK-8 assay, their effects on cell senescence were examined by β-galactosidase staining, and their effects on ferroptosis were detected by a flow cytometric analysis of lipid ROS levels, transmission electron microscopy, and an assay for cellular iron levels. The related mechanisms were investigated by real-time PCR and Western blot assays. Our results showed that LPC, as the most differential metabolite, inhibited cell viability but promoted cell apoptosis and senescence as its concentration increased. Also, the decreased cell activity, increased iron ion and lipid ROS levels, and the enhanced cell senescence induced by LPC treatment were all significantly reversed by AS-IV but further enhanced by FIN56 treatment. The changes in mitochondrial morphology caused by the LPC treatment were significantly alleviated by the AS-IV treatment, while treatment with FIN56 reversed those phenomena. Moreover, AS-IV partially upregulated the levels of SLC7A11 and GPX4 expression which were reduced by LPC. However, those changes were prevented by FIN56 treatment. In conclusion, our data suggested that AS-IV could serve as a novel drug for treating ferroptosis-related diseases.

Stem cells have received attention in various diseases, such as inflammatory, cancer, and bone diseases. Mesenchymal stem cells (MSCs) are multipotent stem cells that are critical for forming and repairing bone tissues. Herein, we isolated calycosin-7-O-β-glucoside (Caly) from the roots of Astragalus membranaceus, which is one of the most famous medicinal herbs, and investigated the osteogenic activities of Caly in MSCs. Caly did not affect cytotoxicity against MSCs, whereas Caly enhanced cell migration during the osteogenesis of MSCs. Caly increased the expression and enzymatic activities of ALP and the formation of mineralized nodules during the osteogenesis of MSCs. The osteogenesis and bone-forming activities of Caly are mediated by bone morphogenetic protein 2 (BMP2), phospho-Smad1/5/8, Wnt3a, phospho-GSK3β, and phospho-AKT, inducing the expression of runt-related transcription factor 2 (RUNX2). In addition, Caly-mediated osteogenesis and RUNX2 expression were attenuated by noggin and wortmannin. Moreover, the effects were validated in pre-osteoblasts committed to the osteoblast lineages from MSCs. Overall, our results provide novel evidence that Caly stimulates osteoblast lineage commitment of MSCs by triggering RUNX2 expression, suggesting Caly as a potential anabolic drug to prevent bone diseases.

Keywords: AKT; ALP; BMP2; Caly; MSCs; RUNX2; mineralization; osteogenesis.

Dactylogyrus ctenopharyngodonid and Ichthyophthirius multifiliis are two important ectoparasites of freshwater fish. Co-infection by the two parasites leads to high fish mortality and results in heavy economic losses. This study aimed to evaluate the efficacy of medicated feed and a ginger extract bath against D. ctenopharyngodonid and I. multifiliis on grass carp and investigate the hematological response of grass carp co-infected by the two parasites. These results demonstrated that red blood cell (RBC) and thrombocyte percentage among leucocytes significantly decreased after grass carp were co-infected by D. ctenopharyngodonid and I. multifiliis. The monocyte and neutrophil percentages significantly increased with the increment of parasite mean intensities, while the lymphocyte percentage decreased. The activities of serum acid phosphatase (ACP), alkaline phosphatase (AKP), lysozyme (LZM), and superoxide dismutase (SOD) significantly increased after co-infection. When grass carp treated with medicated feed containing 4% of Astragalus membranaceus, Allium sativum, Morus alba, and Glycyrrhiza uralensis, the activities of ACP, AKP, LZM, and SOD were significantly enhanced, and the mean intensities of D. ctenopharyngodonid and I. multifiliis were significantly decreased. When grass carp was treated with medicated feed and a 4-mg/L ginger extract bath, all parasites were eliminated during 28 days. The bath of ginger extract at a concentration of 4 mg/L kept a low mean intensity of I. multifiliis and D. ctenopharyngodonid, then the two parasites were eliminated by oral administration of the medicated feed with an immunostimulant (Chinese medicine compound).

OBJECTIVE

To explore the relationship between continuous cropping obstacle and autotoxicity of Astragalus membranaceus var. mongholicus.

METHODS

Distilled water(CK), water extracts of rhizosphere soil(50, 100, 200 and 400 mg/mL) were applied to test their effect on early growth and physiological characteristics of Astragalus membranaceus var. mongholicus.

RESULTS

The water extracts from rhizospher soil of cultivated Astragalus membranaceus var. mongholicus significantly increased seedling emergence rate, root length and vigor index of Astragalus membranaceus var. mongholicus seedling when at the concentration of 100 mg/mL or below, however,there was no significant effect at 200 mg/mL or higher. The water extracts from rhizosphere soil of cultivated Astragalus membranaceus var. mongholicus significantly reduced the SOD activity in Astragalus membranaceus var. mongholicus seedling at 400 mg/mL and POD activity at 200 mg/mL and 400 mg/mL,while significantly increased the MDA content.

CONCLUSIONS

Water extracts from Astragalus membranaceus var. mongholicus rhizosphere soil significantly affected Astragalus membranaceus var. mongholicus germination and seedling growth in a concentration-dependent manner, generally, low concentrations increased the SOD and POD activity which improved seed germination and seedling growth, while high concentrations caused cell membrane damage of the seedling.

OBJECTIVE

To study the effect of astragalus membranaceus (Fisch) bge. on the lipid peroxidation and NO level of myocardium from rats with diabetic nephropathy.

METHODS

Male wistar rats were divided into five groups: (1) A normal control group, (2) Diabetes group, (3) Diabetic nephropathy group, (4) Diabetic nephropathy group complicated with acute renal function damage, (5) Astragalus membranaceus (Fisch) bge. treatment group. The activity of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), nitric oxide synthase (NOS) and content of malondialdehyde (MDA), nitric oxide (NO) was determined biochemically from myocardial homogenate among different groups.

RESULTS

The activity of GSH-Px, SOD decreased and the content of MDA, NO and NOS activity increased in each experimental group compared with control group, as was more obvious in diabetic nephropathy group with acute renal function damage, which astragalus membranaceus (Fisch) bge could improve.

CONCLUSIONS

Astragalus membranaceus (Fisch) bge. could protect the myocardium in diabetic nephropathy by inhibiting lipid peroxidation and decreasing the level of NO.

OBJECTIVE

To observe the protective effects of Rhodiola-astragalus membranaceus mixture against brain damage during hypoxia under simulated plateau environment and the mechanisms maybe involved in.

METHODS

Adult SD rats were randomly divided into 3 groups, which were normoxic control, simulated plateau hypoxia, and Rhodiola-astragalus membranaceus mixture pretreatment group. Rats in the latter two groups were exposed to simulated 8000 m altitude in a hypobaric chamber for 7 h. Water content, Na+, K(+)-ATPase activity and SOD activity in cerebral tissue, malondialdehyde (MDA) and lactic acid content in cerebral homogenate and serum were measured.

RESULTS

As compared with control group, cerebral water content was significantly higher in hypoxia group, while it was obviously lower in pretreatment group. MDA contents of hypoxia group both in cerebral homogenate and serum were higher than that of control group, while the pretreatment group they were both decreased obviously. Lactic acid content of hypoxia group in cerebral and in serum increased markedly and decreased drastically in pretreatment group compared to that of hypoxic group.

CONCLUSIONS

Rhodiola-astragalus membranaceus mixture has preventive effects on hypoxic damage induced by simulated plateau environment. This prevention may be related to the antagonistic effect on membrane lipid peroxidation and the inhibition on the accumulation of lactic acid in brain tissue and serum.

OBJECTIVE

To investigate the effect of hairy root of Astragalus membranaceus(HRA).

METHODS

HRA were given 10 g/kg per day for 50 days to aged mice treated with D-galactose, and the effect on memory and antioxidant functions were estimated. After administration of HRA 10 g/kg for four days, anti-ischemia-reperfusion kidney model of rat was prepared. The kidney function, activity of superoxide dismutase (SOD) and content of malondialdehyde(MDA) in kidney were examined. Mice with immunosuppression induced by cyclophosphamide were given orally HRA 10 g/kg for 12 days. The activity of natural killer (NK) cells was measured.

RESULTS

HRA improved the memory, raised SOD activity in brain and liver, decreased the MDA content in the liver of aged mice, reduced the MDA content in ischemia-reperfusion kidney, decreased the creatinine level in blood of rats, and promoted the activity of NK cells in immunosuppressed mice.

CONCLUSIONS

Similar to the natural A. membranaceus, HRA has senility-preventing, antioxidizing and immunomodulating functions.

In this study we investigate the effects of cadmium stress on Astragalus membranaceus seedlings and the alleviative effects of attapulgite clay in growth substrate on cadmium stress to A. membranaceus seedlings. The results showed that the Y (Ⅱ) (effective photochemical quantum yield of PSⅡ photosynthetic), qP(photochemical quenching coefficient), ETR(the rate of non-cyclic electrontransport through PSⅡ), and chlorophyll content of the leaves were significantly decreased with the increase of cadmium concentrations, while the cadmium content, non-photochemical quenching(NPQ, qN) of the leaves and cadmium content, MDA content, plasma membrane permeability, and the damage degree of root apical membrane of the roots were significantly increased. Simultaneously, the activities of APX(ascorbate peroxidase), SOD(superoxide dismutase), POD(peroxidase), CAT(catalase), soluble protein content, and soluble sugar content of roots were increased first but then decreased with the increasing cadmium concentration. Under the condition of without Cd stress, the attapulgite clay into the growth substrate did not significantly affect above physiological indexes of leaves, but significantly increased SOD activity and soluble sugar content of roots and decreased the MDA content, damage degree of root apical membrane of roots, while other physiological indexes did not significantly change. Under cadmium stress, the presence of attapulgite clay in the growth substrate significantly alleviated the cadmium-induced decreases Y (Ⅱ), qP, ETR and chlorophyll content of leaves, and the CAT activity, soluble protein content, and soluble sugar content of roots. Under condition with cadmium stress, the presence of attapulgite clay significantly alleviated the cadmium-induced increases of leaves cadmium content, qN and NPQ, and the cadmium content, MDA content, plasma membrane permeability, damage degree of root apical membrane, SOD, POD, and APX activity of the roots. And, the alleviative effects of attapulgite clay on cadmium stress to A. membranaceus roots were more obvious with the increase of cadmium stress time. The above results showed that the addition of attapulgite clay into the growth substrate has certain alleviative effect on the cadmium stress to A. membranaceus seedlings.

A 56-day feeding trial was conducted to investigate the effects of dietary supplementation of Astragalus membranaceus or/and Bupleurum chinense on the growth performance, immune enzymes, and related gene expression of Pacific white shrimp (Litopenaeus vanammei). Six experimental diets were formulated and supplemented with two levels (0.25% and 0.5%) of each herb and their combination. At the end of the trial, the specific growth rate and feed conversion ratio of shrimp were significantly (P < 0.05) improved by herbal diets. Besides, the activities of immune-related enzymes such as superoxide dismutase (SOD), alkaline phosphatase (AKP), and lysozyme in serum and hepatopancreas were significantly (P < 0.05) elevated in shrimp fed A. membranaceus or/and B. chinense. The high expression levels of immune deficiency (IMD), lysozyme, and Toll-like receptor mRNA directly or indirectly reflected the activation effect of innate immune in shrimp by dietary A. membranaceus or/and B. chinense. However, no significant difference (P > 0.05) among the herbal incorporated treatments was detected on the growth performance and immune response. In conclusion, the results suggest that A. membranaceus and B. chinense could be used as a beneficial feed additives and alternatives to antibiotics for white shrimp aquaculture.

Keywords: Astragalus membranaceus; Bupleurum chinense; Gene expression; Litopenaeus vanammei; immune response.

Introduction: Radix Astragali, the dried root of Astragalus membranaceus (Fish.) Bge. (family Fabaceae), which is known as Huangqi in China, has been proven to be an immunostimulant, diuretic, antidiabetic, analgesic, and it has also been used as a health food supplement in some Asian populations and also serves as a lead herb in many traditional Chinese medicine formulations as well as in Chinese ethnic tonifying soups.

Objective: Screening and purification of bioactive compounds from natural products is challenging work due to their complexity. We present the first report on the use of pressurised liquid extraction and on-line two-dimensional counter current chromatography as an efficient medium for scaled-up extraction and separation of six bioactive compounds from Astragalus membranaceus.

Method: We applied the established method with ultrafiltration-liquid chromatography to screen acetylcholinesterase inhibitors, which were then evaluated and confirmed for anti-Alzheimer activity using PC12 cell model.

Results: Six major compounds, namely, calycosin-7-O-β-d-glucoside, pratensein-7-O-β-d-glucoside, formononetin-7-O-β-d-glucoside, calycosin, genistein, and formononetin, with acetylcholinesterase binding affinities were identified and isolated from the raw plant materials via two sets of n-hexane/ethyl acetate/0.2% acetic acid (first-stage counter current chromatography) and n-hexane/ethyl acetate/methanol/water (second-stage counter current chromatography) solvent systems: 1.87:1.0:1.33 and 5.62:1.0:2.42:5.25, v/v/v/v, which were optimised by a mathematical model.

Conclusion: Therefore, a useful platform for the large-scale production of bioactive and nutraceutical ingredients was developed herein. With the on-line system developed here, we present a feasible, selective, and effective strategy for rapid screening and identification of enzyme inhibitors from complex mixtures.

Keywords: Alzheimer's disease; Astragalus membranaceus; PC12 cell; pressurised liquid extraction; two-dimensional counter current chromatography.

Tan sheep is an indigenous ovine breed of China known for its high meat quality and pleasing taste. Seven herbs of traditional Chinese medicine, namely, Ephedra sinica, Glycyrrhiza uralensis, Caragana korshinskii, Allium mongolicum, Thymus vulgaris, Astragalus membranaceus, and Lespedeza bicolor are commonly grazed by Tan sheep. It has been widely believed that these herbs are of high nutritive value, which may significantly contribute to the high meat quality and distinct flavor of Tan sheep. However, the nutritive values of these herbs have not been evaluated to date. In this study, samples of the seven herbs were collected from the steppe of Yanchi County of Ningxia Autonomous Region of China. The dry matter (DM), crude protein (CP), ether extract (EE), ash (Ash), calcium (Ca), phosphorus (P), neutral detergent fiber (NDF), and acid detergent fiber (ADF) of these herbs were measured using locally cultivated alfalfa as the standard forage. Digestion of the dry matter, neutral detergent fiber, acid detergent fiber, and crude protein in the rumen of Tan sheep was examined using the nylon bag method, in order to evaluate their feeding nutritional value. Our results show that all the seven herbs meet the nutritional needs of ruminants based on the standard forage alfalfa. However, Ephedra, Glycyrrhiza, Caragana, Allium, Astragalus, and Lespedeza have higher nutritive value than Thymus (P < 0.05). According to the ruminal degradation rates of dry matter, neutral detergent fiber, acid detergent fiber, and crude protein, the nutritive value of Caragana, Allium, and Lespedeza is higher than that of Ephedra, Glycyrrhiza, Astragalus, and Thymus (P < 0.05). The overall nutritive value of Allium is the highest among the seven herbs and therefore Allium is recommended to better meet the nutritional needs of Tan sheep.

Keywords: Chinese herbs; Tan sheep; degradation; nutritive value.

Ethnopharmacological relevance: Fufang Xueshuantong Capsule (FXC) is a traditional Chinese medicine (TCM) formula composed of four herbs including Panax notoginseng, Astragalus membranaceus, Salvia miltiorrhiza, and Scrophularia ningpoensis. Long-term and extensive clinical applications have confirmed that FXC could exert significant effects on fundus, cardiovascular and cerebrovascular occlusive diseases.

Aim of the review: To systematically analyze and summarize the existing researches involving quality and efficacy re-evaluation of FXC, point out the typical problems, and further propose some opinions to contribute to future study.

Materials and methods: Literatures concerning FXC were collected from online scientific databases including China National Knowledge Infrastructure, WanFang Data, PubMed, Science Direct, Scopus, Web of Science, Springer Link up to June 2020. All eligible studies are analyzed and summarized in this review.

Results: This review outlines the chemical profiles, quality control, pharmacokinetic and pharmacological properties of FXC based on reported results. Some problems are pointed out for FXC: the quality control needs further improvement, the pharmacokinetic properties have not been comprehensively investigated, and in-depth and systematic mechanism researches are scarce. Hereon we propose several directions for future study: (a) establishment of feasible HPLC or LC-MS based quantitative methods for simultaneous determination of multiple components to monitor the overall quality; (b) pharmacokinetic studies concerning humans, drug-drug interactions, and correlation with pharmacodynamics; (c) pharmacological mechanism researches integrating multi-omics technologies (gut microbiome, metabolomics, etc.).

Conclusions: This review highlights the researches on quality and efficacy re-evaluation of FXC, and points out some typical problems. Further in-depth studies should focus on the promotion of quality control, pharmacokinetic properties, and pharmacological mechanism.

Keywords: Chemical composition; Fufang xueshuantong capsule; Pharmacokinetics; Pharmacological properties; Quality control.

Astragalus membranaceus (AM) is a traditional Chinese medicine, which possesses a variety of biological activities in the cardiovascular systems. We conducted a clinical and preclinical systematic review of 28 randomized clinical control studies with 2522 participants and 16 animal studies with 634 animals to evaluate the efficacy, safety, and possible mechanisms of AM for viral myocarditis (VM). The search strategies were performed in 7 databases from inception to January 2020. Application of the Cochrane Collaboration's tool 7-item checklist, SYRCLE's tool 10-item checklist, and Rev-Man 5.3 software to analyze the risk of bias of studies and data. The results show the score of clinical study quality ranged from 3 to 7 points with an average of 3.32, and the score of animal study quality ranged from 2 to 5 points with an average of 3. In clinical study, AM significantly reduced serum myocardial enzymes and cardiac troponin I levels and improved the clinical treatment efficiency in VM patients compared with the control group (P < 0.05). There was no significant difference in the incidence of adverse reactions (P > 0.05). Significant increase of the survival rate and decrease of the cardiac cardiology score, cardiac enzymes, and cardiac troponin I were compared with the placebo group in animal studies (P < 0.05). The possible mechanisms of AM are largely through antivirus and antivirus receptors, anti-inflammatory, antioxidation, antiapoptotic, antifibrosis, and reducing cardiac calcium load. In conclusion, the findings suggested that AM is a cardioprotection candidate drug for VM.