Artificial neural network as an alternative to multiple regression analysis in optimizing formulation parameters of cytarabine liposomes

The objective of the study was to optimize the formulation parameters of cytarabine liposomes by using artificial neural networks (ANN) and multiple regression analysis using 3(3) factorial design (FD). As model formulations, 27 formulations were prepared. The formulation variables, drug (cytarabine)/lipid (phosphatidyl choline [PC] and cholesterol [Chol]) molar ratio (X1), PC/Chol in percentage ratio of total lipids (X2), and the volume of hydration medium (X3) were selected as the independent variables; and the percentage drug entrapment (PDE) was selected as the dependent variable. A set of causal factors was used as tutorial data for ANN and fed into a computer. The optimization was performed by minimizing the generalized distance between the predicted values of each response and the optimized one that was obtained individually. In case of 3(3) factorial design, a second-order full-model polynomial equation and a reduced model were established by subjecting the transformed values of independent variables to multiple regression analysis, and contour plots were drawn using the equation. The optimization methods developed by both ANN and FD were validated by preparing another 5 liposomal formulations. The predetermined PDE and the experimental data were compared with predicted data by paired t test, no statistically significant difference was observed. ANN showed less error compared with multiple regression analysis. These findings demonstrate that ANN provides more accurate prediction and is quite useful in the optimization of pharmaceutical formulations when compared with the multiple regression analysis method.     

Literature mining and database annotation of protein phosphorylation using a rule-based system

MOTIVATION: A large volume of experimental data on protein phosphorylation is buried in the fast-growing PubMed literature. While of great value, such information is limited in databases owing to the laborious process of literature-based curation. Computational literature mining holds promise to facilitate database curation. RESULTS: A rule-based system, RLIMS-P (Rule-based LIterature Mining System for Protein Phosphorylation), was used to extract protein phosphorylation information from MEDLINE abstracts. An annotation-tagged literature corpus developed at PIR was used to evaluate the system for finding phosphorylation papers and extracting phosphorylation objects (kinases, substrates and sites) from abstracts. RLIMS-P achieved a precision and recall of 91.4 and 96.4% for paper retrieval, and of 97.9 and 88.0% for extraction of substrates and sites. Coupling the high recall for paper retrieval and high precision for information extraction, RLIMS-P facilitates literature mining and database annotation of protein phosphorylation.     

Filling-in Void and Sparse Regions in Protein Sequence Space by Protein-Like Artificial Sequences Enables Remarkable Enhancement in Remote Homology Detection Capability

Protein functional annotation relies on the identification of accurate relationships, sequence divergence being a key factor. This is especially evident when distant protein relationships are demonstrated only with three-dimensional structures. To address this challenge, we describe a computational approach to purposefully bridge gaps between related protein families through directed design of protein-like “linker” sequences. For this, we represented SCOP domain families, integrated with sequence homologues, as multiple profiles and performed HMM-HMM alignments between related domain families. Where convincing alignments were achieved, we applied a roulette wheel-based method to design 3,611,010 protein-like sequences corresponding to 374 SCOP folds. To analyze their ability to link proteins in homology searches, we used 3024 queries to search two databases, one containing only natural sequences and another one additionally containing designed sequences. Our results showed that augmented database searches showed up to 30% improvement in fold coverage for over 74% of the folds, with 52 folds achieving all theoretically possible connections. Although sequences could not be designed between some families, the availability of designed sequences between other families within the fold established the sequence continuum to demonstrate 373 difficult relationships. Ultimately, as a practical and realistic extension, we demonstrate that such protein-like sequences can be “plugged-into” routine and generic sequence database searches to empower not only remote homology detection but also fold recognition. Our richly statistically supported findings show that complementary searches in both databases will increase the effectiveness of sequence-based searches in recognizing all homologues sharing a common fold.     

Superoxide radical production and performance index of Photosystem II in leaves from magnetoprimed soybean seeds

Priming of soybean seeds with static magnetic field exposure of 200 mT (1 h) and 150 mT (1 h) resulted in plants with enhanced performance index (PI). The three components of PI i.e the density of reaction centers in the chlorophyll bed (RC/ABS), exciton trapped per photon absorbed (φpo) and efficiency with which a trapped exciton can move in electron transport chain (Ψo) were found to be 17%, 27%  and 16% higher, respectively in leaves from 200 mT (1h) treated compared to untreated seeds.  EPR spectrum of  O₂^{. –} - PBN  adduct revealed that the O₂^{. –} radical level was lower by 16% in the leaves of plants that emerged from magnetic field treatment. Our study revealed that magnetoprimed seeds have a long lasting stimulatory effect on plants as reduced superoxide production and higher performance index contributed to higher efficiency of light harvesting that consequently increased biomass in plants from treated plants.     

Metabolism of cholesterol by callus culture of Holarrhena antidysenterica

A chemically defined medium was established for the growth of tissue cultures of Holarrhena antidysenterica. Administration of cholesterol-[4-¹⁴C] to 10-day-old callus yielded radioactive 24-methylenecholesterol, 28-isofucosterol, sitosterol, stigmasterol, and conessine, thereby indicating that the conversion of cholesterol into sitosterol is mediated through 24-methylenecholesterol and 28-isofucosterol in this system.     

The relationship between phytoplankton distribution and water column characteristics in North West European shelf sea waters

Phytoplankton underpin the marine food web in shelf seas, with some species having properties that are harmful to human health and coastal aquaculture. Pressures such as climate change and anthropogenic nutrient input are hypothesized to influence phytoplankton community composition and distribution. Yet the primary environmental drivers in shelf seas are poorly understood. To begin to address this in North Western European waters, the phytoplankton community composition was assessed in light of measured physical and chemical drivers during the "Ellett Line" cruise of autumn 2001 across the Scottish Continental shelf and into adjacent open Atlantic waters. Spatial variability existed in both phytoplankton and environmental conditions, with clear differences not only between on and off shelf stations but also between different on shelf locations. Temperature/salinity plots demonstrated different water masses existed in the region. In turn, principal component analysis (PCA), of the measured environmental conditions (temperature, salinity, water density and inorganic nutrient concentrations) clearly discriminated between shelf and oceanic stations on the basis of DIN:DSi ratio that was correlated with both salinity and temperature. Discrimination between shelf stations was also related to this ratio, but also the concentration of DIN and DSi. The phytoplankton community was diatom dominated, with multidimensional scaling (MDS) demonstrating spatial variability in its composition. Redundancy analysis (RDA) was used to investigate the link between environment and the phytoplankton community. This demonstrated a significant relationship between community composition and water mass as indexed by salinity (whole community), and both salinity and DIN:DSi (diatoms alone). Diatoms of the Pseudo-nitzschia seriata group occurred at densities potentially harmful to shellfish aquaculture, with the potential for toxicity being elevated by the likelihood of DSi limitation of growth at most stations and depths.     

Cloning and characterization of differentially expressed genes of internal breakdown in mango fruit (Mangifera indica)

Internal breakdown in mango fruits has become a major concern in recent years. This disorder renders the fruits unfit for human consumption. The overall loss due to this disorder is about 35-55%. Environmental and physiological factors like high temperature, humidity, respiration and low transpiration rates have been attributed to cause spongy tissue due to reduced loss of heat from fruits. Biochemical studies have shown that there is a reduction in pH, total soluble solids, ascorbic acid, total sugars and carotenoids, low reducing and non-reducing sugar contents, lower amylase and invertase activities and high acid and starch content in spongy tissue affected pulp. There are no reports on molecular studies to determine changes in gene expression in these tissues. The present study was conducted using PCR based subtractive hybridization and RNA gel blot analysis of a few selected genes. The latter showed a higher expression of catalase, ubiquitin, alcohol dehydrogenase, coproporphyrinogen oxidase and keratin associated protein. A lower expression of ribosomal gene, fructose bisphosphate aldolase and cysthathionine gamma synthase was also noticed in spongy tissue. Biochemical studies indicated a lower amylase activity and a lower content of the total and reducing sugars in spongy tissue as compared to healthy tissue. Analyses of results indicate that oxidative stress may be one of the causes for formation of spongy tissue, which affects the expression of many genes. The role of these genes in spongy tissue formation is discussed.     

An assay combining high-performance liquid chromatography and mass spectrometry to measure DNA interstrand cross-linking efficiency in oligonucleotides of varying sequences

The main method of evaluating the DNA interstrand cross-linking ability of cancer chemotherapeutic agents in naked DNA currently involves the electrophoresis of relatively long radiolabeled duplex DNA fragments (typically approximately 2000 bp) on neutral gels after incubation with the agent of interest. Denaturation by heating is carried out prior to loading, and a neutral gel allows reannealing of cross-linked DNA. To avoid the use of radioactivity we have developed a new method based on ion pair reversed phase liquid chromatography (RPLC) and mass spectrometry (MS) that allows characterization and quantitation of drug-DNA interstrand cross-links formed within short oligonucleotide duplexes (i.e., 12 bp). Advantages of this assay include rapid throughput, as compared to electrophoretic methods, and the use of readily available short nonradiolabeled oligonucleotides of any sequence, thereby facilitating investigation of sequence selectivity. A further advantage is that all species separated by the chromatographic process can be positively identified by MS. Using this new method, we have investigated the rate of DNA cross-linking and sequence selectivity of the interstrand cross-linking agent SJG-136, a pyrrolobenzodiazepine (PBD) dimer currently in phase I clinical trials. The assay was found to be sufficiently sensitive and selective to allow separation of the unbound and drug-bound oligonucleotide species by high-performance liquid chromatography (HPLC) and to allow positive identification of these individual species by MS. A further benefit, as compared with electrophoretic methods, is that kinetic information can be obtained and compared for different binding sequences.