N-acetyl-d-glucosamine-based oligosaccharides from chitin: Enzymatic production, characterization and biological activities.

Chitin, the second most abundant biopolymer, possesses diverse applications in the food, agricultural, and pharmaceutical industries due to its functional properties. However, the potential applications of chitin are limited owing to its high crystallinity and low solubility. N-acetyl chitooligosaccharides and lacto-N-triose II, the two types of GlcNAc-based oligosaccharides, can be obtained from chitin by enzymatic methods. With their lower molecular weights and improved solubility, these two types of GlcNAc-based oligosaccharides display more various beneficial health effects when compared to chitin. Among their abilities, they have exhibited antioxidant, anti-inflammatory, anti-tumor, antimicrobial, and plant elicitor activities as well as immunomodulatory and prebiotic effects, which suggests they have the potential to be utilized as food additives, functional daily supplements, drug precursors, elicitors for plants, and prebiotics. This review comprehensively covers the enzymatic methods used for the two types of GlcNAc-based oligosaccharides production from chitin by chitinolytic enzymes. Moreover, current advances in the structural characterization and biological activities of these two types of GlcNAc-based oligosaccharides are summarized in the review. We also highlight current problems in the production of these oligosaccharides and trends in their development, aiming to offer some directions for producing functional oligosaccharides from chitin.

Combination treatment of Danggui Buxue Decoction and endothelial progenitor cells can enhance angiogenesis in rats with focal cerebral ischemia and hyperlipidemia.

ETHNOPHARMACOLOGICAL RELEVANCE: Danggui Buxue Decoction (DBD) is a classic prescription of traditional Chinese medicine that is mainly used for treating clinical anemia for more than 800 years. This prescription has been utilized for nourishing "Qi" and enriching "Blood" for women suffering from menopausal symptoms. Meanwhile, DBD has the role of improving angiogenesis and promoting the neuroprotective functions. Bone marrow-derived endothelial progenitor cells (EPCs) was suboptimal to treat the focal cerebral ischemia (FCI). Thus, it's may be a novel strategy of DBD combined with EPCs transplantation for the FCI. AIM OF THE STUDY: To investigate the mechanistic effects of DBD in combination with EPCs transplantation to improve behavioral function of the FCI and hyperlipidemia. MATERIALS AND METHODS: We used rats with hyperlipidemia to develop a FCI model using photo-thrombosis, and treated the DBD in combination with EPCs transplantation. We adopted the Modified Neurological Severity Score to evaluate the neurological deficit, undertook the 2,3,5-triphenyltetrazolium chloride staining to calculate the total infarct volume. We carried out the RT-qPCR, Immunohistochemical analyses, TUNEL, ELISA, and Western blotting to measure the gene and protein levels which related to anti-apoptosis mechanisms and angiogenesis. RESULTS: Administration of DBD in combination with EPCs transplantation was found to improve behavioral function, reducing the infarct volume and decrease the level of total-cholesterole (TC) and low-density lipoprotein-cholesterol (LDL-C). Treatment of DBD plus EPCs increased the mRNA and protein expression of vascular endothelial growth factor A, fibroblastic growth factor-2, and angiopoietin-1 and decreased the apoptosis of endothelial cells by activating the phosphoinositide 3-kinase/protein kinase B/Bcl-xL/Bcl-2 associated death promoter (PI3K/Akt/BAD) pathway and promoting activation of the extracellular signal-regulated kinase (ERK) pathway, which induced angiogenesis directly. CONCLUSIONS: Our findings provided that DBD administration combined with EPCs transplantation promoted reconstruction of nervous function. This was achieved by enhancing expression of the growth factors related to anti-apoptosis mechanisms and angiogenesis thanks to regulation of the PI3K/Akt/BAD and ERK signaling pathways, and might be relate to the lowering of TC and LDL-C levels.

The Treatment of Complementary and Alternative Medicine on Female Infertility Caused by Endometrial Factors.

Recently, with the development of the social economy, the incidence of infertility has increased year by year. With its complex etiology and diversified syndromes, infertility has become one of the most important diseases that plague the physical and mental health of women of childbearing age worldwide. Endometrial factors as an important part affecting female reproductive capacity, due to which induced repeated abortion and multiple uterine cavity operations occur, can destruct endometrium, failing to provide a normal implantation environment for zygote, thus resulting in infertility. Many patients failed to achieve expected results after receiving conventional treatments such as hormone therapy, assisted reproductive technology (ART), granulocyte colony-stimulating factor (G-CSF) therapy, and cell therapy, then turn to complementary and alternative medicine (CAM) therapies for help. Aiming at clarifying the effectiveness and mechanisms of CAM therapy in the treatment of infertility caused by endometrial factors, our paper systematically searched and studied present related literature on the PubMed, CNKI, and other databases, focusing on the aspects of clinical application and mechanism explorations and highlighting the therapeutic effects of Chinese herbal medicine (CHM), acupuncture, and moxibustion on such diseases. Moreover, this paper also introduces the CAM treatments of traditional Chinese medicine (TCM) retention enema, neuromuscular electrical stimulation (NMES), photobiomodulation therapy, dietary intervention, and other measures for infertility caused by endometrial factors, in order to provide a reference for subsequent basic research and clinical work.

Two-color fluorescent proteins reporting survivin regulation in breast cancer cells for high throughput drug screening.

Reporter gene assay is widely used for high throughput drug screening and drug action mechanism evaluation. In this study, we developed a robust dual-fluorescent reporter assay to detect drugs repressing the transcription of survivin, a cancer biomarker from the inhibitor of apoptosis family, in breast cancer cells cultured in three-dimensional (3D) microbioreactors. Survivin is overexpressed in numerous malignancies but almost silent in normal tissue cells and is considered a lead target for cancer therapy. Breast cancer MCF-7 cells were engineered to express enhanced green fluorescent protein driven by a survivin promoter and red fluorescent protein driven by a cytomegalovirus promoter as internal control to detect changes in survivin expression in cells as affected by drugs. This 3D dual-fluorescent reporter assay was validated with YM155 and doxorubicin, which were known to downregulate survivin in cancer cells, and further evaluated with two widely used anticancer compounds, cisplatin, and epigallocatechin gallate, to evaluate their effects on survivin expression. The results showed that the 3D dual-fluorescent reporter assay was robust for high throughput screening of drugs targeting survivin in breast cancer cells.

The Treatment with Complementary and Alternative Traditional Chinese Medicine for Menstrual Disorders with Polycystic Ovary Syndrome.

Polycystic ovary syndrome (PCOS) is a frequent gynecological female endocrinopathy, characterized by chronic anovulation, hyperandrogenism, and insulin resistance (IR). Menstrual disorders are one of the main clinical manifestations of PCOS. Other symptoms include hirsutism and/acne. At present, the treatment of PCOS with irregular menstruation is mainly based on oral contraceptives, but there are some side effects and adverse reactions. In recent years, more and more attention has been paid to the complementary and alternative medicine (CAM), which has been widely used in clinical practice. Modern Western medicine is called "conventional medicine" or "orthodox medicine," and the complementary and alternative medicine is called "unconventional medicine" or "unorthodox medicine." CAM includes traditional medicine and folk therapy around the world. Around 65-80% of world health management business is classified into traditional medicine by the World Health Organization, which is used as alternative medicine in Western countries. In our country, Chinese medicine, acupuncture, and other therapies are commonly used due to their significant efficacy and higher safety. Therefore, this review aims to summarize and evaluate the mechanisms and the effect of current complementary replacement therapy in the treatment of menstrual disorders caused by PCOS, so as to provide guidance for the following basic and clinical research.

The Treatment of Complementary and Alternative Medicine on Premature Ovarian Failure.

It has been confirmed by growing evidence that common hormone replacement therapy is associated with an increasing risk of causing cardiovascular disease and cancer, while complementary and alternative medicine (CAM) is gaining popularity and application in more and more patients with premature ovarian failure (POF). Although there is little data concerning the clinical safety and efficacy of CAM, the literature includes application studies on the phytoestrogen-rich herbal, acupuncture treatment and intervention therapy. This article reviews recent literature on CAM therapy for POF, aiming to provide theoretical support for clinical application.

Correlation study on non-volatile ingredients of Menthae Haplocalycis Herba formula granules and traditional herbal pieces by UPLC-MS/MS / 中国中药杂志

To establish the method for determining non-volatile ingredients of neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid, caffeic acid, isochlorogenic acid A, rosmarinic acid, ferulic acid, rutin, luteoloside, isoquercitrin, hesperidin, diosmin, diosmetin, luteolin, acacetin and linarin in Menthae Haplocalycis Herba formula granules and traditional herbal pieces by UPLC-MS/MS, and analyze the correlation of non-volatile ingredients in Menthae Haplocalycis Herba formula granules and traditional herbal pieces. Shim-pack GIST C_(18) column(2.1 mm×100 mm, 2 μm) was adopted with acetonitrile-0.1% formic acid aqueous solution as the mobile phase for gradient elution at the flow rate of 0.4 mL·min~(-1). The column temperature was set at 35 ℃. The quantitative analysis was performed using the electrospray ionization source and the multiple reaction monitoring. The linear relationship, resolution, repeatability and recovery of the 16 chemical components all met the requirements. The 16 non-volatile ingredients in traditional herbal pieces of Menthae Haplocalycis Herba could be tracked in formula granules. There were certain differences of the 16 chemical components among Menthae Haplocalycis Herba formula granules of different manufacturers and traditional herbal pieces of different producing areas. The UPLC-MS/MS method was simple, rapid and accurate, and could be used for the quality control of non-volatile ingredients in Menthae Haplocalycis Herba formula granules and traditional herbal pieces.

Erythromycin has therapeutic efficacy on muscle fatigue acting specifically on orosomucoid to increase muscle bioenergetics and physiological parameters of endurance.

At present, there are still no official or semi-official recommendations for the treatment of muscle fatigue. We previously reported that acute phase protein orosomucoid (ORM) can enhance muscle endurance and exert anti-fatigue effect. In attempting to seek anti-fatigue drugs that target ORM, we found macrolide antibiotics, particularly erythromycin, were effective. Erythromycin can significantly prolong the time of mice forced-swimming and treadmill running, increase muscle fatigue index, alleviate fatigue-induced tissue damage, and elevate glycogen content, mitochondria function and ATP level in the muscle. Also, erythromycin increases ORM protein expression in a dose- and time- dependent manner both in vitro and in vivo. Further studies found that erythromycin could increase the activity of ORM promoter and the stability of ORM mRNA, which might both be responsible for the ORM up-regulation. ORM knockdown or knockout could abolish the promoting effect of erythromycin in mice forced-swimming time, muscle fatigue index and glycogen level. Furthermore, those effects were also abolished in mice with C-C motif chemokine receptor 5 (CCR5) antagonist administration or AMPKα2 deficiency. Therefore, erythromycin could enhance muscle glycogen and endurance via up-regulating the level of ORM and activating CCR5-AMPK pathway, indicating it might act as a potential drug to treat muscle fatigue.

Activation of the Cholinergic Anti-Inflammatory Pathway as a Novel Therapeutic Strategy for COVID-19.

The outbreak of coronavirus disease 2019 (COVID-19) underlined the urgent need for alleviating cytokine storm. We propose here that activating the cholinergic anti-inflammatory pathway (CAP) is a potential therapeutic strategy. However, there is currently no approved drugs targeting the regulatory pathway. It is evident that nicotine, anisodamine and some herb medicine, activate the CAP and exert anti-inflammation action in vitro and in vivo. As the vagus nerve affects both inflammation and specific immune response, we propose that vagus nerve stimulation by invasive or non-invasive devices and acupuncture at ST36, PC6, or GV20, are also feasible approaches to activate the CAP and control COVID-19. It is worth to investigate the efficacy and safety of the strategy in patients with COVID-19.

Inhibitory effect of chitooligosaccharides on retinol metabolism and bioavailability in mice.

This study investigated the intervention effects of chitooligosaccharides (COS) on retinol metabolism and included comparisons of the retinol level, retinol binding protein 4 (RBP4) content, key genes, and protein expression between mice on a COS-enriched diet and a normal diet. The results showed that COS markedly decreased the retinol and RBP4 concentrations in the serum and liver. Furthermore, COS suppressed the mRNA and protein expression of RBP4, cellular retinol binding protein 1 (CRBP1), lecithin: retinol acyltransferase (LRAT) and cytochrome P45026A1 (CYP26A1). In addition, COS inhibited the mRNA expression of stimulated by retinoic acid 6 (STRA6). However, the protein expression of STRA6 was not significantly decreased. Thus, COS reduced the retinol concentration in the serum and disrupted the metabolism of retinol. The intervention mechanism of COS on retinol metabolism may be attributed to the modulation of RBP4, CRBP1, LRAT, STRA6, and CYP26A1 expression at the mRNA and protein levels. PRACTICAL APPLICATIONS: Chitooligosaccharides (COS), known to be the degradation products of chitosan, have been found to induce pinkeye in industrial workers who participate in the manufacturing of COS. Meanwhile, 5% population with COS dietary supplement also have similar phenomenon. The aim of this study is to explore the possible mechanism underlay of this potential risk. The results of this study showed that high exposure to COS during manufacture influences retinol metabolism and leads to a decrease in retinol content, ultimately causing pinkeye. These findings provide new evidence for understanding COS-induced retinol metabolism alteration and drawing attention toward the prevention of potential risk in high-exposure populations.

The Evolution and Biocatalysis of FAD2 Indicate Its Correlation to the Content of Seed Oil in Plants.

Unsaturated fatty acids are the main components of vegetable oils. Fatty acid desaturase 2 (FAD2) catalyzes oleic acid (OA) into linoleic acid (LA) transformations, which are essential to the profile of FAs in seeds. To further understand the roles of FAD2s in the synthesis of oil, the evolution and biocatalysis of FAD2s were comprehensively analyzed. The evolution history of the FAD2 gene family showed that most of the FAD2 genes formed monophyletic clades except in eudicots. The FAD2 genes in some eudicots diverged into constitutive and seed-specific expression clades. Notably, the biocatalysis of seed-specific or -abundant expression FAD2s in soybean, perilla, rice, and spruce revealed that their catalytic activity was strongly correlated with the total oil content of their seeds in nature. Additionally, it was found that I and Y in site 143 of GmaFAD2-1 were strictly conserved in the seed-specific and constitutive expression clades of Fabaceae, respectively. Furthermore, the site-directed mutation demonstrated that I and Y are vital to improving and reducing the activity of GmaFAD2s. Therefore, the results indicate that the activity of FAD2s in seeds might be a reference to the total oil content of seeds, and site 143 might have been specifically evolved to be required for the activity of FAD2s in some expression-diverged eudicots, especially in legumes.

Comparison of eleven active components in Lonicerae Japonicae Flos,Lonicerae Japonicae Caulis and leaves of Lonicero japonica by UPLC / 中国中药杂志

This present study aims to establish a UPLC method for simultaneously determining eleven components such as new chlorogenic acid,chlorogenic acid,caffeic acid,cryptochlorogenic acid,artichoke,isochlorogenic acid A,isochlorogenic acid B,isochlorogenic acid C,rutin,hibisin and loganin in Lonicerae Japonicae Flos,Lonicerae Japonicae Caulis and leaves of Lonicera japonica and comparing the differences in the contents of phenolic acids,flavonoids and iridoid glycosides of Lonicerae Japonicae Flos,Lonicerae Japonicae Caulis and leaves of Lonicera japonica.The method was carried out on an ACQUITY UPLC BEH C18column(2.1 mm×100 mm,1.7 μm) by a gradient elution using acetonitrile and 0.1% phosphoric acid.The flow rate was 0.3 mL·min-1.The column temperature was maintained at 30 ℃.The sample room temperature was 8 ℃.The wavelength was set at 326 nm for new chlorogenic acid,chlorogenic acid,caffeic acid,cryptochlorogenic acid,artichoke,isochlorogenic acid A,isochlorogenic acid B and isochlorogenic acid C,352 nm for rutin and lignin,and 238 nm for loganin.The injection volume was 1 μL.The eleven components has good resolution and was separated to baseline.Each component had a wide linear range and a good linear relationship(r≥0.999 6),the average recovery rate(n=9) was 98.96%,100.7%,97.24%,97.06%,99.53%,96.78%,98.12%,95.20%,95.12%,100.2%,98.61%and with RSD was 2.5%,1.4%,1.9%,2.1%,1.7%,1.9%,1.6%,2.0%,1.4%,2.2%,2.0%,respectively.Based on the results of the content determination,the chemometric methods such as cluster analysis and principal component analysis were used to compare the Lonicerae Japonicae Flos,Lonicerae Japonicae Caulis and leaves of Lonicera japonica.The results showed that Lonicerae Japonicae Flos and leaves of Lonicera japonica were similar in the chemical constituents,but both showed chemical constituents difference compored to Lonicerae Japonicae Caulis.The established multi-component quantitative analysis method can provide a reference for the quality control of Lonicerae Japonicae Flos,Lonicerae Japonicae Caulis and leaves of Lonicera japonica.

Effect of intralipid on myocardial injury during valve replacement surgery with concomitant radiofrequency ablation: A randomized controlled trial.

BACKGROUND: This study aimed to evaluate the effect of intralipid postconditioning (ILPC) on myocardial damage in patients undergoing valve replacement surgery with concomitant radiofrequency ablation (RFA) for atrial fibrillation (AF). METHODS: Randomized patient and assessor-blind controlled trial conducted in adult patients undergoing valve replacement surgery with concomitant RFA. Sixty-nine patients were randomly assigned to ILPC group (n = 34) or control group (n = 35): ILPC group received an intravenous infusion of 20% intralipid (2 mL/kg) just 10 minutes before aortic cross-unclamping, and control group received an equivalent volume of normal saline. Serum cardiac troponin-T (cTnT) and creatine kinase-MB (CK-MB) was measured before surgery and at 4, 12, 24, 48, and 72 hours after surgery. The primary endpoints were the 72-hour area under the curve (AUC) for cTnT and CK-MB. RESULTS: The total 72-hour AUC of cTnT (P = .33) and CK-MB (P = .52) were comparable between 2 groups. The left ventricle ejection fraction at discharge (P = .011) was higher in the ILPC group than that in the control group, while the AF recurrence did not differ significantly between 2 groups. CONCLUSIONS: There was no observed beneficial effect of ILPC on myocardial injury documented by the cardiac biomarkers in patients undergoing valve replacement surgery with concomitant RFA, and the effect of intralipid against myocardial I/R injury is undetectable within the background of massive biomarker release following ablation owing to localized myocardial necrosis. Besides, there are no other published data about the cardioprotective role of intralipid in patients undergoing this procedure and benefits of this protection need further studies to validate.

Oral Chinese Herbal Medicine as an Adjuvant Treatment for Chemotherapy, or Radiotherapy, Induced Myelosuppression: A Systematic Review and Meta-Analysis of Randomized Controlled Trials.

OBJECTIVE: Myelosuppression is a common side effect in cancer patients receiving chemotherapy or radiotherapy. Chinese herbal medicine (CHM) has shown promise in alleviating myelosuppression. METHOD: We searched for randomized controlled trials (RCTs) from seven databases without language restriction. We included RCTs in adults, in which hematological toxicity was measured according to WHO criteria and control group underwent chemotherapy and/or radiotherapy and the treatment group was given oral CHM. RESULTS: We searched 1021 articles from the date of databases inception to October 7, 2016. We selected 14 articles for the final analysis. Pooled data showed that CHM significantly decreased the suppression rate of leukocytes, neutrophils, hemoglobin, and platelets compared with the control group, particularly in grade III-IV toxicity (leukocytes: RR = 0.43, 95% CI = 0.33-0.56; neutrophils: RR = 0.39, 95% CI = 0.27-0.58; hemoglobin: RR = 0.33, 95% CI = 0.18-0.61; platelets: RR = 0.61, 95% CI = 0.39-0.95). CONCLUSIONS: CHM as an adjuvant can alleviate myelosuppression induced by chemotherapy or radiotherapy, reduce grade III-IV toxicity, and maintain therapeutic dose and treatment cycle. However, due to heterogeneity and publication bias, the results should be interpreted with caution and validated by conducting strictly designed multicenter RCTs of high quality and large scale.

Just noticeable differences for pitch direction, height, and slope for Mandarin and English listeners.

Previous studies on tones suggest that Mandarin listeners are more sensitive to pitch direction and slope while English listeners primarily attend to pitch height. In this study, just noticeable differences were established for pitch discrimination using a three-interval, forced-choice procedure with a two-down, one-up staircase design. A high rising and a high falling Mandarin tone were manipulated in terms of pitch direction, height, and slope. Results indicate that, overall, Mandarin listeners are more sensitive to pitch slope and English listeners to pitch height. However, these effects are modulated by both the direction (falling/rising) and slope of the pitch contours.

Effect of intralipid postconditioning on myocardial injury in patients undergoing valve replacement surgery: a randomised controlled trial.

OBJECTIVE: This study was conducted to determine whether the administration of intralipid just before aortic cross-unclamping would reduce myocardial injury in patients undergoing valve replacement surgery. METHODS: Seventy-three adult patients, scheduled for elective aortic or mitral valve surgery without significant coronary stenosis (>70%), were randomly assigned to the intralipid postconditioning (ILPC) group (n=37) or control group (n=36): the ILPC group received an intravenous infusion of 20% intralipid (2 mL/kg) just 10 min before aortic cross-unclamping, and the control group received an equivalent volume of normal saline. Serum cardiac troponin T (cTnT) and creatine kinase-MB (CK-MB) was measured before surgery and at 4, 12, 24, 48 and 72 hours after surgery. The primary end points were the 72-hour area under the curve (AUC) for cTnT and CK-MB. RESULTS: No significant difference between the ILPC and control arm was observed, including the age, sex or number of aortic versus mitral valves or left ventricular ejection fraction at baseline. The total 72-hour AUC of cTnT and CK-MB in patients assigned to ILPC were significantly reduced by 32.3% (p=0.004) and 26.4% (p=0.0185) compared with control, respectively. None of the treated patients had abnormal blood lipid metabolism, abnormal renal or hepatic function or significant related complications. CONCLUSION: The protective effect of postischaemic administration of intralipid prior to aortic cross-unclamping on reperfusion injury was found when determined by biomarkers of myocardial injury but not by cardiac function or other clinical outcomes in patients undergoing valve replacement surgery. Hence, clinical benefits of this protection need larger clinical trials to confirm. TRIAL REGISTRATION NUMBER: ClinicalTrials.gov ID: ChiCTR-IOR-14005318.

The Acute-Phase Protein Orosomucoid Regulates Food Intake and Energy Homeostasis via Leptin Receptor Signaling Pathway.

The acute-phase protein orosomucoid (ORM) exhibits a variety of activities in vitro and in vivo, notably modulation of immunity and transportation of drugs. We found in this study that mice lacking ORM1 displayed aberrant energy homeostasis characterized by increased body weight and fat mass. Further investigation found that ORM, predominantly ORM1, is significantly elevated in sera, liver, and adipose tissues from the mice with high-fat diet (HFD)-induced obesity and db/db mice that develop obesity spontaneously due to mutation in the leptin receptor (LepR). Intravenous or intraperitoneal administration of exogenous ORM decreased food intake in C57BL/6, HFD, and leptin-deficient ob/ob mice, which was absent in db/db mice and was significantly reduced in mice with arcuate nucleus (ARC) LepR knockdown, whereas enforced expression of ORM1 in ARC significantly decreased food intake, body weight, and serum insulin level. Furthermore, we found that ORM is able to bind directly to LepR and activate the receptor-mediated JAK2-STAT3 signaling in hypothalamus tissue and GT1-7 cells, which was derived from hypothalamic tumor. These data indicated that ORM could function through LepR to regulate food intake and energy homeostasis in response to nutrition status. Modulating the expression of ORM is a novel strategy for the management of obesity and related metabolic disorders.

Evaluation of safety of modified-Danggui Buxue Tang in rodents:immunological, toxicity and hormonal aspects.

ETHNOPHARMACOLOGICAL RELEVANCE: Radix Astragali (RA), Radix Angelicae Sinensis (RAS) and Folium Epimedii (FE) are three of the extensively applied herbs among traditional Chinese medicines for gynecological disorders and osteoporosis. A derivative herbal formula-RRF, consisting of the three medicines with a weight ratio of 5:1:5, is derived from a famous Chinese herbal formula-Danggui Buxue Tang (DBT). RRF has shown noteworthy perimenopause ameliorating effect in both ovariectomized rats and natural aging female rats, which might represent a promising candidate for the treatment of perimenopausal disorders. The aim of this study was to evaluate its immunological potential, chronic toxicity and reproductive effects by 26-week repeated daily administration in female rats, in order to optimize its safe use. MATERIALS AND METHODS: The effect of RRF on immunological function was studied by macrophage phagocytosis, immune organ index, serum immunoglobulin level as well as delayed type hypersensitivity (DTH) in mice. For toxicity assessment, acute toxicity study was performed according to fixed dose procedure with a single oral administration of RRF to mice. In the oral chronic toxicity, 120 female rats were administrated RRF orally in 0, 1100, 4400, or 8800mg/kg/day doses for 26 weeks. Clinical signs, mortality, body weights, feed consumption, haemato-biochemical parameters, organ weights, histopathology and reproductive hormone profiles were examined at the end of the 13- and 26-week dosing period, as well as after the 4-week recovery period. RESULTS: Oral administration of RRF at three doses (282, 564 and 1128mg/kg) significantly increased the indices of phagocytosis K, as compared with prednisone acetate (PR) group (p<0.05 or 0.01). Exposure of RRF dose-dependently boosted circulating serum IgM level (all p<0.01) in response to CRBC in PR-induced mice. Furthermore, RRF treatment elicited a significant increment (all p<0.01) in DNFB-induced DTH response and the immune organ indices in a dose-dependent manner in mice, in parellel to DNFB-induced group. In the single dose acute toxicity and repeated dose 90-day chronic toxicity investigations, no toxic signs/mortality were observed. RRF treatment did not cause any toxicologically significant changes in clinical signs, food consumption, body weight, relative organ weight, hematological parameters, clinical chemistry, gross pathology and histopathology between treatment and control groups. No treatment related gross/histopathological lesions were observed and no target organ was identified. Long-term repeated administration of RRF exerted a significant promotion on serum level of steroid hormone estradiol, progesterone and testosterone release, along with decrease of circulating pituitary follicle stimulating hormone, luteinizing hormone, and prolactin levels in female rats. The No Observed Adverse Effect Level (NOAEL) of RRF was determined to be over 8800mg/kg/day for elderly female rats, a dose that was equivalent to 50 times of human dose. CONCLUSION: The present investigation demonstrated that RRF possessed appreciable immunopotentiating activity and had a relatively wide margin of safety. Long-term treatment of RRF exhibited estrogenic properties, and retarded certain age-associated degenerations. RRF might have the potential for further development as a safe and effective alternative/complementary to conventional medication in relieving perimenopausal symptoms.

[Rapid analysis of compounds in leaves of Chinese seabuckthorn and Tibetan seabuckthorn by UPLC/Q-TOF-MS].

The study is aimed to analyze the chemical components in leaves of Chinese seabuckthorn and Tibetan seabuckthorn qualitatively and compare the differences between them by using ultra performance liquid chromatography coupled with quadrupole-time-of-flight mass spectrometry (UPLC/Q-TOF-MS).The chromatographic separation of the components was achieved ona Waters ACQUITY UPLC-T3 column (2.1 mm×100 mm, 1.7 µm）using gradient mobile phase consisting of acetonitrile (A) and aqueous solution (B). The identification of the separated compounds was performed on atandem mass spectrometry (MS/MS)by fragmentation patterns under the negative electrospray ionization. The parameters of ion source were as follows：capillary voltage, 2 000 V; Cone voltage, 40 V. The ion source temperature, 100 ℃; collision gas argon; sheath gas flow rate, 900 L•h⁻¹; sheath gas temperature, 450 ℃. Through the analysis of mass spectrometry data and with the help of literature data, a total of 35 compounds were detected and most of them were flavonoids. Among these compounds, 29 were common components for the two species, two components were unique to Chinese seabuckthorn and 4 were characteristic components of Tibetan seabuckthorn. The results indicated that the compositions of the two kinds of seabuckthorn leaves were quite similar. It is also demonstrated that UPLC/Q-TOF-MS method could be applied to rapidly and effectively analyze and speculate the compounds in leaves of Chinese seabuckthorn and Tibetan seabuckthorn.

[Effect of Rhizoma Polygonati on the Function of Endothelial Progenitor Cells and Telomerase Ac- tivity in Nature Senescent Rats].

Objective To observe the effect of Rhizoma Polygonati (RP) on the function of spi- nal endothelial progenitor cells (EPCs) and telomerase activity in nature senescent rats. Methods To- tally 48 natural senescent rats (20 months old) were randomly divided into 4 groups according to their body weight, i.e., the blank group, high, medium, low dose RP groups, 12 in each group. Meanwhile, 12 young rats (5 months old) were recruited as a normal control group. RP decoction at 4, 2, 1 g/kg was in- tragastrically administered to rats in high, medium, low dose RP groups, once per day for 12 successive weeks. Equal volume of normal saline was intragastrically administered to rats in the normal control group and the blank group, once per day for 12 successive weeks. Contents of antioxidant indices were detec- ted. Meanwhile, adherence cells were identified after in- vitro culture of mononuclear cells of bone marrow in each group. The proliferation was detected using MTT assay. The migration was detected by Transwell assay. The tubule function was detected using in vitro angiogenesis kit. And the telomerase activity was detected by real time fluorescent quantitative PCR. Results Compared with the normal control group, serum total-antioxidant capacity (T-AOC) , contents of superoxide dismutase (SOD) and glutathione-per- oxidase (GSH-Px) were obviously reduced, malondialdehyde (MDA) content obviously increased, the function of bone marrow EPCs was obviously damaged in the blank group (P <0. 01). Compared with the blank group, serum contents of T-AOC, SOD, and GSH-Px obviously increased and MDA content decreased in high and medium dose RP groups (P <0. 05,P <0. 01). The proliferation and migration of EPCs, the tubule function, and the telomerase activity could be promoted in high and medium dose RP groups (P <0. 05,P <0. 01). Conclusion RP could protect damaged function of EPCs in senescent rats by promoting serum antioxidant level and telomerase activity.