The role of chemical fingerprinting: application to Ephedra

Ephedra sinica, known as Ma Huang, is one of the oldest medicinal herbs in Traditional Chinese Medicine (TCM). Preparations, namely teas, of E. sinica have been used for over 5000 years as a stimulant and as an antiasthmatic. In the West, extracts of E. sinica, E. intermedia or E. equisetina are most commonly used in dietary supplements as a stimulant and to promote weight loss. More than 50 species of Ephedra are native to both hemispheres, but the detection of ephedrine alkaloids has been limited to species in Eurasia. Currently, methods exist to quantitate the ephedrine alkaloids in extracts of plant material or dietary supplements, but the methods are not able to verify the extract is of an Ephedra species. Reverse phase high performance liquid chromatography with photodiode array detection was applied for the chemical fingerprinting of the Ephedra species. Two regions of comparison were determined in the chromatograms at 320 nm. The series of peaks between 52 and 64 min confirms an Ephedra species is being analyzed. The aforementioned peaks also could distinguish between Ephedra species from Eurasia, North America and South America. Peaks at ca. 57 and 59 min were isolated and determined to be two new compounds, 4-(2-eicosyloxycarbonyl-vinyl)-benzoic acid and 4-(2-docosyloxycarbonyl-vinyl)-benzoic acid respectively. Authentication of ground plant material as Ephedra can be achieved by this chemical fingerprinting method.     

The prevalence of androstenone anosmia

It has been estimated that approximately 30% of the population is unable to detect the odor of androstenone. These estimates, however, were made using tests and criteria optimized for identifying detection. Such criteria favor Type II over Type I errors--that is, they are excellent at identifying true detectors at the cost of erroneously labeling some detectors as non-detectors. Because these criteria were used to identify non-detectors, it is possible that the rate of non-detection may have been overestimated. To test this we screened 55 subjects for non-detection employing previously used methods. This screen yielded nine putative non-detectors, a 16.3% putative non-detection rate. We then retested these putative non-detectors using a forced choice (yes-no) paradigm to obtain a precise measure of their sensitivity. We found that this group of putative non-detectors was significantly above chance at detecting androstenone (P < 0.001), despite very low self-confidence in their performance. Based on the results of the signal detection analysis in this sample, we estimate the rate of actual androstenone non-detection in young healthy adults is between 1.8 and 5.96%, which is significantly lower than previously estimated. This finding is significant considering the implications of specific anosmias on the understanding of odor discrimination.     

The effect of plant growth regulators, nitric oxide, nitrate, nitrite and light on the germination of dimorphic seeds of Suaeda salsa under saline conditions

Suaeda salsa, a leaf succulent shrub in the family Chenopodiaceae, is one of the most important halophytes in China. Suaeda salsa produces dimorphic seeds (soft brown seeds and hard black seeds). Seeds of S. salsa were collected from the coastal salt flats near Huanghua City, China. Experiments were conducted to determine the salinity-alleviating effect of plant growth regulators, nitric oxide, nitrate, nitrite and light on the germination of dimorphic seeds of S. salsa. Brown seeds had a higher germination rate than black seeds in all experiments. Black seeds were more sensitive to salt in the absence of light in comparison to brown seeds. Brown seeds absorbed water more quickly in comparison to black seeds and were found to be more tolerant of salt stress. Our results showed that 1-aminocyclopropane-1-carboxylate (ACC, the immediate precursor of ethylene), nitrite, GA₄ and BA improved seed germination in the presence of salt. However, nitrate, GA₁, GA₃ failed to alleviate salt stress. ABA inhibited seed germination and seedling growth. Possible mechanisms involved in the alleviation of salt stress in S. salsa seeds and the ecological adaptation of the seeds to the environment are discussed.     

Critical factors influencing the occurrence of Vibrio cholerae in the environment of Bangladesh

The occurrence of outbreaks of cholera in Africa in 1970 and in Latin America in 1991, mainly in coastal communities, and the appearance of the new serotype Vibrio cholerae O139 in India and subsequently in Bangladesh have stimulated efforts to understand environmental factors influencing the growth and geographic distribution of epidemic Vibrio cholerae serotypes. Because of the severity of recent epidemics, cholera is now being considered by some infectious disease investigators as a "reemerging" disease, prompting new work on the ecology of vibrios. Epidemiological and ecological surveillance for cholera has been under way in four rural, geographically separated locations in Bangladesh for the past 4 years, during which both clinical and environmental samples were collected at biweekly intervals. The clinical epidemiology portion of the research has been published (Sack et al., J. Infect. Dis. 187:96-101, 2003). The results of environmental sampling and analysis of the environmental and clinical data have revealed significant correlations of water temperature, water depth, rainfall, conductivity, and copepod counts with the occurrence of cholera toxin-producing bacteria (presumably V. cholerae). The lag periods between increases or decreases in units of factors, such as temperature and salinity, and occurrence of cholera correlate with biological parameters, e.g., plankton population blooms. The new information on the ecology of V. cholerae is proving useful in developing environmental models for the prediction of cholera epidemics.     

Influence of Heparin Mimetics on Assembly of the FGF·FGFR4 Signaling Complex

Fibroblast growth factor (FGF) signaling regulates mammalian development and metabolism, and its dysregulation is implicated in many inherited and acquired diseases, including cancer. Heparan sulfate glycosaminoglycans (HSGAGs) are essential for FGF signaling as they promote FGF·FGF receptor (FGFR) binding and dimerization. Using novel organic synthesis protocols to prepare homogeneously sulfated heparin mimetics (HM), including hexasaccharide (HM₆), octasaccharide (HM₈), and decasaccharide (HM₁₀), we tested the ability of these HM to support FGF1 and FGF2 signaling through FGFR4. Biological assays show that both HM₈ and HM₁₀ are significantly more potent than HM₆ in promoting FGF2-mediated FGFR4 signaling. In contrast, all three HM have comparable activity in promoting FGF1·FGFR4 signaling. To understand the molecular basis for these differential activities in FGF1/2·FGFR4 signaling, we used NMR spectroscopy, isothermal titration calorimetry, and size-exclusion chromatography to characterize binding interactions of FGF1/2 with the isolated Ig-domain 2 (D2) of FGFR4 in the presence of HM, and binary interactions of FGFs and D2 with HM. Our data confirm the existence of both a secondary FGF1·FGFR4 interaction site and a direct FGFR4·FGFR4 interaction site thus supporting the formation of the symmetric mode of FGF·FGFR dimerization in solution. Moreover, our results show that the observed higher activity of HM₈ relative to HM₆ in stimulating FGF2·FGFR4 signaling correlates with the higher affinity of HM₈ to bind and dimerize FGF2. Notably FGF2·HM₈ exhibits pronounced positive binding cooperativity. Based on our findings we propose a refined symmetric FGF·FGFR dimerization model, which incorporates the differential ability of HM to dimerize FGFs.     

CD8+ CTLs are essential for protective immunity against Encephalitozoon cuniculi infection

Encephalitozoon cuniculi is a protozoan parasite that has been implicated recently as a cause of opportunistic infection in immunocompromised individuals. Protective immunity in the normal host is T cell-dependent. In the present study, the role of individual T cell subtypes in immunity against this parasite has been studied using gene knockout mice. Whereas CD4-/- animals resolved the infection, mice lacking CD8+ T cells or perforin gene succumbed to parasite challenge. The data obtained in these studies suggest that E. cuniculi infection induces a strong and early CD8+ T response that is important for host protection. The CD8+ T cell-mediated protection depends upon the CTL activity of this cell subset, as the host is rendered susceptible to infection in the absence of this function. This is the first report in which a strong dependence upon the cytolytic activity of host CD8+ T cells has been shown to be important in a parasite infection.     

QTL mapping of fire blight resistance in apple

Fire blight caused by the bacterium Erwinia amylovora is a severe threat to apple and pear orchards worldwide. Apple varieties exhibit a wide range of relative susceptibility/tolerance to fire blight. Although, no monogenic resistance against fire blight has been identified yet, recent evidence indicates the existence of quantitative resistance. Potential sources of fire blight resistance include several wild Malus species and some apple cultivars. F1 progenies of ‘Fiesta’בDiscovery’ were inoculated with the Swiss strain Ea 610 and studied under controlled conditions to identify quantitative trait loci (QTLs) for fire blight resistance. Disease was evaluated at four time points after inoculation. Shoot lesion length and the area under disease progress curve (AUDPC) values were used for QTL analysis. One significant (LOD score of 7.5–8.1, p<0.001) QTL was identified on the linkage group 7 of ‘Fiesta’ (F7). The F7 QTL explained about 37.5–38.6% of the phenotypic variation.     

Quantification of biochemical compounds in Bauhinia Variegata Linn flower extract and its hepatoprotective effect

Liver disorders may occur as a result of exposure to chemical compounds capable of inducing the oxidative stress and hepatic injuries. The aim of present study was to investigate the effects of flower extracts of B. Variegata for the treatment of liver injury induced by the CCl₄. About 1 ml/kg body weight (b.w) of CCl4 was induced to experimental mice by intraperitoneal way for 14 days. The methanol and chloroform extracts (100, 200 and 300 mg/kg b.w) were administered to experimental animals for 14 days along with standard drug Silymarine (100 mg/kg b.w). The extracts alone showed no evidence of hepatic toxicity but animals exposed to CCl₄ without the treatment with B. Variegata presented variations in levels of liver enzymes, antioxidant enzymes, proteins and blood cells as well as injuries in liver cells were also observed during histopathological study. However, after the treatments especially with 300 mg/kg b.w of methanol flower extracts levels of liver markers (ALT, AST and ALP), antioxidant enzymes and blood cells decreases and turned towards normal levels. Whereas level of total proteins and bilirubin was improved and damaged liver cells were repaired. The curative activity of flower extracts can be correlated to the higher potential of antioxidants and occurrence of Quercetin and some other organic compounds those were investigated from flower extracts of B. Variegata during HPLC and GC-MS analysis. The finding of this study supports the use of B. Variegata flower formulation in folk medicines.     

Structural and functional analysis of NS1 and NS2 proteins of H1N1 subtype

Influenza A virus (H1N1), a genetic reassortment of endemic strains of human, avian and swine flu, has crossed species barrier to human and apparently acquired the capability of human to human transmission. Some strains of H5N1 subtype are highly virulent because NS1 protein inhibits antiviral interferon α/β production. Another protein NS2 mediates export of viral ribonucleoprotein from nucleus to the cytoplasm through export signal. In this paper, we have studied structure-function relationships of these proteins of H1N1 subtype and have determined the cause of their pathogenicity. Our results showed that non-conservative mutations slightly stabilized or destabi- lized structural domains of NS1 or NS1-dsRNA complex, hence slightly increased or decreased the function of NS1 protein and consequently enhanced or reduced the pathogenicity of the H1N1 virus. NS2 protein of different strains carried non-conservative mutations in different domains, resulting in slight loss of function. These mutations slightly decreased the pathogenicity of the virus. Thus, the results confirm the structure-function relationships of these viral proteins.