Comparative evaluation of extraction methods for total proteins from Crocus sativus L. (Saffron)

Broadly speaking proteomic studies are one of the various techniques of utmost importance for understanding complex biological processes that occur under inductive conditions and revealing the multidimensional aspects of Crocus sativus in biological systems. In order to get an insight into the molecular changes and to characterize the variations in protein expression of C. sativus, a detailed proteomic analysis on one-dimensional gel electrophoresis is one of the basic steps to accomplish. We have compared total protein profiles of C. sativus extracted by three different recipes and analyzed on 10% sodium dodecyl sulfate polyacrylamide gels. Gels were subjected to densitometric analysis for further characterization. Among three different protocols NP-40 extraction buffer recipe resulted in the extraction of proteins most efficiently with minimum background and streaking. There was maximum solubilization of proteins with high efficiency. Such a profile can be used for high precision analysis of differential protein expression. This work is an attempt to assist researchers in effective extraction of proteins from C. sativus. As a researcher faces a perplexing array of choices as where to start we describe a method based on our collective analysis of the different protein protocols. This paper presents a method that could be applied at the outset of any proteomic study.     

In vitro label‐free screening of chemotherapeutic drugs using Raman microspectroscopy: Towards a new paradigm of spectralomics

This overview groups some of the recent studies highlighting the potential application of Raman microspectroscopy as an analytical technique in preclinical development to predict drug mechanism of action and in clinical application as a companion diagnostic and in personalised therapy due to its capacity to predict cellular resistance and therefore to optimise chemotherapeutic treatment efficacy. Notably, the anthracyclines, doxorubicin and actinomycin D, elicit similar spectroscopic signatures of subcellular interaction characteristic of the mode of action of intercalation. Although cisplatin and vincristine show markedly different signatures, at low exposure doses, their signatures at higher doses show marked similarities to those elicited by the intercalating anthracyclines, confirming that anticancer agents can have different modes of action with different spectroscopic signatures, depending on the dose. The study demonstrates that Raman microspectroscopy can elucidate subcellular transport and accumulation pathways of chemotherapeutic agents, characterise and fingerprint their mode of action, and potentially identify cell‐resistant strains. The consistency of the spectroscopic signatures for drugs of similar modes of action, in different cell lines, suggests that this fingerprint can be considered a “spectralome” of the drug‐cell interaction suggesting a new paradigm of representing spectroscopic responses.      

Vegetation mapping above tree line in Nandiar valley,western Himalayas. A multivariate approach

This paper communicates an analytical exploration of the vegetation above tree line in Nandiar valley western Himalayas with respect to climatic, edaphic and topographic factors. The alpine pasture stretches above the tree limits between elevations of 2850–3800 m. Thirteen stands were selected on the basis of physiognomy of vegetation in 2014–15. The leading life form and leaf spectra were therophytes and microphyll respectively. Four plant communities were identified through two ways indicator species analysis at cut level 2. Bray-Curtis ordination showed that maximum stands have common species. Detrended correspondence analysis grouped species having similar habitats in ordination space. Canonical correspondence analysis constrained species along different environmental variables. The significant result were contribution by phosphorous (P = 0.01) and wind speed (P = 0.08). Few plant species like Betula utilis and Podophyllum emodi are threatened due to its medicinal collection. The alpine pasture of Nandiar valley is under severe stress due to anthropogenic and grazing factors. Overall, this research article call for greater attention to the types of conservation actives occurring and the ways they are working together to protect and recover the global alpine biodiversity.     

Watershed ‘chemical cocktails’: forming novel elemental combinations in Anthropocene fresh waters

In the Anthropocene, watershed chemical transport is increasingly dominated by novel combinations of elements, which are hydrologically linked together as ‘chemical cocktails.’ Chemical cocktails are novel because human activities greatly enhance elemental concentrations and their probability for biogeochemical interactions and shared transport along hydrologic flowpaths. A new chemical cocktail approach advances our ability to: trace contaminant mixtures in watersheds, develop chemical proxies with high-resolution sensor data, and manage multiple water quality problems. We explore the following questions: (1) Can we classify elemental transport in watersheds as chemical cocktails using a new approach? (2) What is the role of climate and land use in enhancing the formation and transport of chemical cocktails in watersheds? To address these questions, we first analyze trends in concentrations of carbon, nutrients, metals, and salts in fresh waters over 100 years. Next, we explore how climate and land use enhance the probability of formation of chemical cocktails of carbon, nutrients, metals, and salts. Ultimately, we classify transport of chemical cocktails based on solubility, mobility, reactivity, and dominant phases: (1) sieved chemical cocktails (e.g., particulate forms of nutrients, metals and organic matter); (2) filtered chemical cocktails (e.g., dissolved organic matter and associated metal complexes); (3) chromatographic chemical cocktails (e.g., ions eluted from soil exchange sites); and (4) reactive chemical cocktails (e.g., limiting nutrients and redox sensitive elements). Typically, contaminants are regulated and managed one element at a time, even though combinations of elements interact to influence many water quality problems such as toxicity to life, eutrophication, infrastructure corrosion, and water treatment. A chemical cocktail approach significantly expands evaluations of water quality signatures and impacts beyond single elements to mixtures. High-frequency sensor data (pH, specific conductance, turbidity, etc.) can serve as proxies for chemical cocktails and improve real-time analyses of water quality violations, identify regulatory needs, and track water quality recovery following storms and extreme climate events. Ultimately, a watershed chemical cocktail approach is necessary for effectively co-managing groups of contaminants and provides a more holistic approach for studying, monitoring, and managing water quality in the Anthropocene.     

Burkholderia pseudomallei: Its Detection in Soil and Seroprevalence in Bangladesh

Background

Melioidosis, caused by Burkholderia pseudomallei, is an endemic disease in Bangladesh. No systematic study has yet been done to detect the environmental source of the organism and its true extent in Bangladesh. The present study attempted to isolate B. pseudomallei in soil samples and to determine its seroprevalence in several districts in Bangladesh.

Methodology and Results

Soil samples were collected from rural areas of four districts of Bangladesh from where culture confirmed melioidosis cases were detected earlier. Multiple soil samples, collected from 5–7 sampling points of 3–5 sites of each district, were cultured in Ashdown selective media. Suspected colonies of B. pseudomallei were identified by biochemical and serological test, and by polymerase chain reaction (PCR) using 16s rRNA specific primers. Blood samples were collected from 940 healthy individuals of four districts to determine anti- B. pseudomallei IgG antibody levels by indirect enzyme linked immunosorbent assay (ELISA) using sonicated crude antigen. Out of 179 soil samples, B. pseudomallei was isolated from two samples of Gazipur district which is located 58 km north of capital Dhaka city. Both the isolates were phenotypically identical, arabinose negative and showed specific 550bp band in PCR. Out of 940 blood samples, anti- B. pseudomallei IgG antibody, higher than the cut-off value (>0.8), was detected in 21.5% individuals. Seropositivity rate was 22.6%-30.8% in three districts from where melioidosis cases were detected earlier, compared to 9.8% in a district where no melioidosis case was either detected or reported (p<0.01). Seropositivity increased with the advancement of age from 5.3% to 30.4% among individuals aged 1–10 years and > 50 years respectively. The seropositivity rates were 26.0% and 20.6% in male and female respectively, while it was 20–27% among different occupational groups. No significant association was observed with gender (χ2 = 3.441, p = 0.064) or any occupational group (χ² = 3.835, p = 0.280).

Conclusion

This is the first study demonstrating the presence of B. pseudomallei in the environmental (soil) samples of Bangladesh. It also suggested that a large proportion of people, residing in these districts, were exposed to the organism.     

Enkephalins-modulation of Plasmodium cynomolgi antigens-induced colony-stimulating factors elaboration by macrophages.

Methionine-enkephalin (M-Enk) and its analogue compound 82/205 (10(-5) and 10(-6) M) inhibited elaboration of Plasmodium cynomolgi total antigens soluble in culture medium (P.c.SA)-induced colony-stimulating factors (CSFs) by monkey blood monocyte-derived macrophages, in vitro. Paradoxically, lower concentrations (10(-7)-10(-9) M) of both the peptides greatly augmented CSFs elaboration; 82/205 appeared to be nearly 2.3-fold more potent. Naloxone (10(-5) M) pretreatment of macrophages inhibited only the M-Enk- and 82/205-induced enhanced CSFs elaboration, suggesting an opiate receptors-mediated mechanism of action. None of the peptides or naloxone (10(-5)-10(-9) M) had any direct effect on the CSF elaboration by unstimulated macrophages.     

The subcellular distribution and partial characterization of cholinesterase activities of canine platelets

The multiple cholinesterase activities in canine platelets have been investigated. Platelets were homogenized by rapid decompression under nitrogen, glass tube/Teflon pestle, and glycerol lysis techniques. Rapid decompression under nitrogen technique was found to be the most efficient and gentle method for cell disruption. Homogenates were subfractionated using sodium diatrizoate density gradients. Marker enzyme assays and pulse labeling experiments with 5-hydroxy[¹⁴C]tryptamine and [¹²⁵I]thrombin on prepared subcellular fractions confirmed that the soluble, plasma membrane and the granule-1 fractions were all in reasonably pure form. Furthermore, labeling of the plasma membrane with [¹²⁵I]thrombin is cited as the first successful attempt at attaining a significantly bound marker for this structure. Cholinesterase activity distributions measured in these fractions indicated that about 30% of the activity was present in the plasma membrane, 50% in granule-1 and 5% in soluble fractions. Kinetic data of cholinesterase activities obtained from intact platelets, plasma membrane preparations and platelet release supernatants indicated that they are strikingly similar.