Spectrum-effect relationship between fingerprints and hemopoietic effects of small molecular fraction of Polygoni Multiflori radix praeparata.

Polygoni multiflori Radix Praeparata (PMRP) is a traditional medicine used for nourishing essence and blood in China. However, it is unclear which PMRP compounds are responsible for its hematopoietic effect. In this study, spectrum-effect relationship was used to discovery potential hematopoietic compounds. The fingerprints of 20 PMRP batches were established by HPLC and the hematopoietic effect was determined using red blood cell, hemoglobin, hematocrit, and platelet indexes in aplastic anemia model mice. The spectrum-effect relationship between common peaks and hematopoietic efficacy values was established using gray relational analysis and partial least squares analysis. Spectrum-effect relationship results showed that peaks 21 (emodin-8-O-(6´-O-acetyl)-ß-D-glucoside), 15 (2, 3, 5, 4'-tetrahydroxystilbene-2-O-di-glucoside), 16 (cis-2,3,5,4'-tetrahydroxy-stilbene-2-O-ß-D-glucoside), 11 (unknown), 20(unknown, 12 (epicatechin), 29 (carboxyl emodin), and 31 (emodin) in the fingerprints were closely related to the hematopoietic effect. This work successfully established the spectrum-effect relationship between PMRP hematopoietic effect and its fingerprints, which can be used to explain the material basis for the PMRP hematopoietic effect.

Extraction and identification of collagen-derived peptides with hematopoietic activity from Colla Corii Asini.

ETHNOPHARMACOLOGICAL RELEVANCE: Colla Corii Asini is a widely used traditional Chinese medicine to treat anemia with a long history due to its stimulating effect in hematopoiesis, but the components contributing to this effect are still unknown. In this study, we aimed to establish a methodology to isolate the bioactive components and provide pharmacological basis for its usage in treating anemia. METHODS: 5-FU and γ-ray radiation induced anemic mice models were generated by treating with 5-FU at 150mg/kg body weight and γ-rays by a 4MV linear accelerator by total body irradiation using female ICR mice respectively. Oral administration of fraction A was performed by gastric lavage at 1g/kg and 2g/kg body weight for 12 days and 25 days and peripheral blood sample was collected from ocular sinus red blood cell (RBC) and white blood cell (WBC) counts every 3 days and 5 days for 5-FU and radiation induced models, respectively. Next, fraction A was separated to A1 and A2 using cation exchange chromatography (IEC) based on ionic strength. Fraction A1 was further separated using reverse phase chromatography (RPC) based on the hydrophobicity first with 0-10% linear gradient, then 20%, 30%, 50% constant gradient of 60% acetonitrile in neutral Na2HPO4 buffer. Peak fractions were pooled, evaporatively dried, and dissolved in ultrapure water. Finally, fraction A11 was analyzed combining tandem mass spectrometry and proteomic tools and two peptides (peptide 11 and 16) were identified. The hematopoietic effects of multiple fractions and the two peptides were measured using colony-forming units-erythroid (CFU-E), an indication of late erythroid progenitor cells and colony-forming units granulocyte-monocyte (CFU-GM), an indication of granulocyte and monocyte progenitor cells respectively on hematopoietic progenitor cells prepared from bone marrow (Till and Mcculloch 1961). RESULTS: Fraction A at 1g/kg and 2g/kg could increase RBC and WBC counts in 5-FU and radiation induced anemic mice models. Fraction A1 at 0.1mg/ml and 0.5mg/ml, exhibited stronger hematopoietic activity than fraction A2, both of which were subfractions from fraction A using IEX, by elevated CFU-E and CFU-GM of mouse bone marrow cells. Furthermore, fraction A11 at 0.1mg/ml showed stronger CFU-E and CFU-GM than fractions A12 to A14 from RPC separation. Finally, peptide 11 and peptide 16 were identified from tandem mass spectrometry and peptide 11 increased CFU-E and CFU-GM in a dose dependent manner. CONCLUSIONS: We combined multiple approaches including chromatography, mass spectrometry, cell-based assays, as well as animal studies to identify and demonstrate that the hematopoietic effect of Colla Corii Asini is at least in part from the peptidic components identified using our methodology. This is the first time to isolate peptidic components from Colla Corii Asini, and to provide molecular basis for its usage in treating anemia, which may particularly have the potential to benefit cancer patients suffering from myelosuppression due to radiotherapy or chemotherapy.

Detecting doping use: more than an analytical problem.

The recent Armstrong case, where more than 250 negative doping tests are confronted with the athlete's confession of erythropoietin use, blood doping, steroid, and growth hormone abuse, illustrates the limitations of current laboratory tests in detecting doping in sport. Despite numerous doping controls and simultaneous indications of common doping abuse among professional athletes in the last two decades, the number of positive urine tests for recombinant human erythropoietin (rHuEPO) remains remarkably low. Athletes are using various masking strategies, among them protease inhibitors, intravenous injections of rHuEPO and alternative erythropoiesis stimulating agents. As one of the countermeasures, the Athlete's Biological Passport has been introduced. The sensitivity of the Athlete's Biological Passport is limited if the effect of a low-dose doping remains within the intra-individual reference range. A possible solution could be the use of a novel Epo test (MAIIA Diagnostics). Another performance-enhancing strategy is the return to 'old' doping techniques, such as autologous blood transfusions. Several indirect methods to detect autologous blood transfusions have been proposed with the majority relying on changes in erythropoiesis-sensitive blood markers. Currently, an algorithm based on the haemoglobin (Hb) level concentration and the percentage of reticulocytes (OFF-hr model; Hb(g/l)-60·âˆš%ret) is approved by the World Anti-Doping Agency. Genetic factors have been identified which may interfere with test interpretation. A large inter- and intra-ethnic variation in testosterone glucuronidation and excretion has been described. Consideration of genetic variation should improve performance of the testosterone doping test. Taking into account the pre-analytical care and better tailoring of the threshold values could increase test sensitivity. Anti-doping laboratories should routinely adjust for multiple testing as failure of doping control to detect cheaters could lead to more frequent controls. Finally, despite the huge technological progress, there is a need for increased collaboration between physiologists, analytical chemists, biostatisticians, and ethicists to reduce doping in sport.

Short-term stability of a new generic sodium ferric gluconate in complex with sucrose.

BACKGROUND: Sodium ferric gluconate in complex (SFG) is used to treat iron deficiency anemia in patients aged ≥6 years undergoing chronic hemodialysis and receiving supplemental epoetin therapy. Both the branded product (Ferrlecit, branded SFG) and a new generic version of sodium ferric gluconate in complex (Nulecit; generic SFG) are provided in 5 mL vials. SFG may be administered by slow intravenous (IV) injection of the undiluted product or by 1 h IV infusion after dilution in 100 mL 0.9% sodium chloride. This study evaluated the short-term stability of undiluted and diluted generic SFG at room temperature and under refrigeration. METHODS: Samples of generic SFG undiluted in 10 mL syringes or diluted in IV infusion bags containing 0.9% sodium chloride solution were stored at room temperature or under refrigerated conditions (2-8°C). Samples at room temperature were stored for ≤48 h if undiluted and for ≤24 h if diluted. All refrigerated samples were stored for ≤7 days. Parameters evaluated were elemental iron (Fe) concentration and SFG apparent molecular weight. All tests were performed on two lots of the generic product. RESULTS: Fe concentrations were identical in both lots and did not vary substantially over time under different conditions of storage or dilution. SFG apparent molecular weight varied across all samples from 306,000 to 354,000 Daltons, well within the range of 289,000 to 440,000 Daltons specified as the molecular weight in the FDA-approved prescribing information. CONCLUSION: Iron content and SFG apparent molecular weight were stable under all experimental conditions. Undiluted generic SFG was stable for ≥2 days at room temperature and ≥7 days under refrigerated conditions, and generic SFG diluted in IV infusion bags containing 0.9% sodium chloride solution was stable for ≥1 day at room temperature and ≥7 days under refrigerated conditions.

Colonic mucosal concentrations of folate are accurately predicted by blood measurements of folate status among individuals ingesting physiologic quantities of folate.

Folate status is inversely related to the risk of colorectal cancer. Whether conventional blood measurements of folate status accurately reflect folate concentrations in the colorectal mucosa has been a controversial topic. This is an important issue because accurate measures of folate status in the colorectal mucosa are important for ascertaining the risk of colorectal cancer in epidemiological studies and for determining the effects of folate supplementation in clinical trials. We examined whether conventional blood measurements of folate and a more sensitive, inverse indicator of systemic folate status, serum homocysteine, accurately reflect folate concentrations in human colonic mucosa obtained by endoscopic biopsy. Study subjects (n = 20) were participants in a randomized trial that investigated the effect of folate supplementation (5 mg daily for 1 year) on provisional molecular markers of colon cancer. Blood samples and biopsies of normal rectosigmoid mucosa were obtained at baseline, at 6 months, and at 1 year. Serum, RBC, and colonic mucosal folate and serum homocysteine concentrations were determined. Colonic mucosal folate concentrations correlated directly with serum folate concentrators at each time point (r = 0.572-0.845; P < 0.015) and with RBC folate concentrations at 6 months and 1 year (r = 0.747-0.771; P < 0.001). Colonic mucosal folate concentrations correlated inversely with serum homocysteine concentrations at each time point (r = -0.622-0.666; P < 0.008). Systemic measures of folate status did not correlate with colonic mucosal folate concentrations among individuals receiving supplemental folate. Our observations indicate that colonic mucosal concentrations of folate may be predicted accurately by blood measurements of folate status only among individuals not ingesting supraphysiological quantities of folate.