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PubMed2005

Source identification of Indian opium based on chromatographic fingerprinting of amino acids.

Total and free pool of amino acids was determined in Indian opium samples using liquid chromatography (LC) with post-column opthalaldehyde derivatization followed by its fluorimetric detection. The limit of detection (LOD) was found to be in the range of 2-10 pmol with a signal to noise ratio of 3:1 and limit of quantitation (LOQ) was found to be in the range of 7-31 pmol with a signal to noise ratio of 10:1. The recovery of amino acids was found to be in the range of 86-103%. A total of 124 Indian opium samples were collected from the states of Madhya Pradesh (MP), Uttar Pradesh (UP) and Rajasthan (Raj), covering 14 licit opium growing divisions of India were chromatographically fingerprinted for the presence of various amino acids. The amino acids identified in sample hydrosylate included D, T, S, S, G, A, V, I, L, Y, F, H, K and R, while the analysis of free pool of amino acids (80% aqueous ethanol extract) indicated the presence of D, T, S, E, A, V, I, L, Y, H, K respectively. Multiple discriminant analysis was applied to the quantitative total amino acid data to determine an optimal classifier in order to evaluate the source of Indian opium. The foremost amino acid variables that accounted for the true discrimination were identified as D, E, G, A, F and K in evaluating the geographical origin of Indian opium and the predictive value based on the discriminant analysis was found to be 90% in relation to the source of opium samples. Chemometrics performed with amino acid analytical data was used successfully in discriminating the licit opium growing divisions of India into three major groups, viz. groups I, II and III. The methodology developed may find wide application in forensic analysis.

Amino Acids↗
PubMed2003

[Identification of Chinese medicinal material radix Angelicae sinensis by X-ray diffraction Fourier fingerprint pattern method].

OBJECTIVE: To develop a new identification and analysis method of Chinese medicinal material Radix Angelicae Sinenesis. METHODS: Powder X-ray Diffraction Fourier Fingerpring pattern. RESULTS: Experiments and analysis were carried out on five samples. The standard X-ray diffraction Fourier pattern and characteristic diffraction peaks of Radix Angelicae Sinensis were obtained. CONCLUSION: This method can be used for identification of Chinese medicinal material Radix Angelicae Sinensis.

Angelica↗
PubMed2003

Fingerprinting of prokaryotic 16S rRNA genes using oligodeoxyribonucleotide microarrays and virtual hybridization.

An oligonucleotide microarray hybridization system to differentiate microbial species was designed and tested. Seven microbial species were studied, including one Bacillus and six Pseudomonas strains. DNA sequences near the 5' end of 16S rRNA genes were aligned and two contiguous regions of high variability, flanked by highly conserved sequences, were found. The conserved sequences were used to design PCR primers which efficiently amplified these polymorphic regions from all seven species. The amplicon sequences were used to design 88 9mer hybridization probes which were arrayed onto glass slides. Single-stranded, fluorescence-tagged PCR products were hybridized to the microarrays at 15 degrees C. The experimental results were compared with the DeltaG(0) values for all matched and mismatched duplexes possible between the synthetic probes and the 16S target sequences of the seven test species, calculated using a 'virtual hybridization' software program. Although the observed hybridization patterns differed significantly from patterns predicted solely on the basis of perfect sequence matches, a unique hybridization fingerprint was obtained for each of the species, including closely related Pseudomonas species, and there was a reasonable correlation between the intensity of observed hybridization signals and the calculated DeltaG(0) values. The results suggest that both perfect and mismatched pairings can contribute to microbial identification by hybridization fingerprinting.

Bacillus↗
PubMed1998

Identification of phototrophic sulfur bacteria through the analysis of lmwRNA band patterns

Several phototrophic sulfur bacteria were identified preliminarily through the analysis of the low-molecular-weight RNA fraction (lmwRNA) of bacterial cells. This fraction includes the ribosomal 5S RNA and several transfer RNAs. These molecules were separated by high-resolution electrophoresis in polyacrylamide gels, and the resulting band patterns were used as fingerprints for the identification of the organisms. We examined a large number of well-characterized reference strains together with a broad range of purple sulfur bacterial isolates from freshwater and marine environments. A cluster analysis was run using the similarity matrix calculated from the band patterns. Despite the shortcomings of the method, close relatives were clustered together yielding a number of groups consistent with the phylogenetic arrangement established through the analyses of a few available 16S rRNA gene sequences. Thus, the classification obtained gives further support to rearrangement of the group as the analyses of 16S rRNA gene sequences had previously suggested. We conclude that the analysis of lmwRNA band patterns is a rapid and simple tool for grouping and preliminarily identifying new isolates of phototrophic sulfur bacteria.

Journal Article↗
PubMed2002

Small molecule affinity fingerprinting. A tool for enzyme family subclassification, target identification, and inhibitor design.

Classifying proteins into functionally distinct families based only on primary sequence information remains a difficult task. We describe here a method to generate a large data set of small molecule affinity fingerprints for a group of closely related enzymes, the papain family of cysteine proteases. Binding data was generated for a library of inhibitors based on the ability of each compound to block active-site labeling of the target proteases by a covalent activity based probe (ABP). Clustering algorithms were used to automatically classify a reference group of proteases into subfamilies based on their small molecule affinity fingerprints. This approach was also used to identify cysteine protease targets modified by the ABP in complex proteomes by direct comparison of target affinity fingerprints with those of the reference library of proteases. Finally, experimental data were used to guide the development of a computational method that predicts small molecule inhibitors based on reported crystal structures. This method could ultimately be used with large enzyme families to aid in the design of selective inhibitors of targets based on limited structural/function information.

Algorithms↗
PubMed2003

Senescence-specific gene expression fingerprints reveal cell-type-dependent physical clustering of up-regulated chromosomal loci.

Replicative senescence is the state of irreversible proliferative arrest that occurs as a concomitant of progressive telomere shortening. By using cDNA microarrays and the gabriel system of computer programs to apply domain-specific and procedural knowledge for data analysis, we investigated global changes in gene transcription occurring during replicative senescence in human fibroblasts and mammary epithelial cells (HMECs). Here we report the identification of transcriptional "fingerprints" unique to senescence, the finding that gene expression perturbations during senescence differ greatly in fibroblasts and HMECs, and the discovery that despite the disparate nature of the chromosomal loci affected by senescence in fibroblasts and HMECs, the up-regulated loci in both types of cells show physical clustering. This clustering, which contrasts with the random distribution of genes down-regulated during senescence or up-regulated during reversible proliferative arrest (i.e., quiescence), supports the view that replicative senescence is associated with alteration of chromatin structure.

Breast↗
PubMed2003

Identification of impurities in phenylacetylene by species-selected mass-analyzed threshold ionization spectroscopy.

Two-color resonant two-photon mass-analyzed threshold ionization (MATI) spectroscopy was used to identify styrene impurities in a commercial phenylacetylene sample. With the mass-gating setup, the recorded MATI spectra are free of contamination from molecules of different mass. Analysis of these spectra gives information on the precise adiabatic ionization and the active cation vibrations, which can be used as fingerprints for the identification of molecular ions.

Acetylene↗
PubMedDate not supplied

Feasibility of using GC/FT-IR for drug analysis in the forensic toxicology laboratory.

New developments in cryogenic sample deposition for gas chromatography/Fourier transform infrared (GC/FT-IR) spectroscopy have increased the sensitivity of the technique 100-1000-fold, to match or surpass that of gas chromatography/mass spectrometry (GC/MS). The current methods of GC/MS have led to some false positive identifications in drug testing labs. New methods employing GC/FT-IR will provide absolute identification through infrared fingerprinting with routine detection in the ppb range. GC/FT-IR methods are being developed which indicate low picogram amounts of material are detectable. Some preliminary data have shown that reference-quality spectra can be obtained from samples containing 200 ng/mL of amphetamines, and spectra from samples below the 25 ng/mL level for amphetamines can be obtained for identification purposes. These and other applications will be addressed along with limit of detection (LOD), limit of quantitation (LOQ), and linearity in comparison to commonly used techniques.

Chromatography, Gas↗
PubMed2005

[Study on the horizontal transmission of oral Streptococcus mutans in children].

OBJECTIVE: To find out horizontal transmission of oral Streptococcus mutas (S. mutans) in nursery children through analyzing the similarity of S. mutans genotypes. METHODS: The study group included 24 nursery children between 3 and 4 years of age. Dental plaque samples were collected with sterile toothpick and cultured on MSB plates for 48 h. Individual Streptococcus mutans group colonies representative of the colonial morphologies were subcultured on TPY plates. These strains were identified to species level biochemically. AP-PCR fingerprinting was preformed after identification. S. mutans isolates from different children with very similar fingerprinting profiles were examined by chromosomal DNA fingerprinting analysis. RESULTS: Streptococcus mutans group were isolated in oral cavities of 66.7% children, 58.3% in caries-free and 75.0% in caries children. A total of 4' S. mutans isolates from 24 subjects were analyzed by AP-PCR, and 29 different amplitypes were identifyied, 45.8% carried tw. genotypes. There were 2 genotypes of S. mutans isolated repeatedly among 12 nursery children. CONCLUSION: The presence of matching genotypes of S. mutans among nursery children suggests horizontal transmission.

Child, Preschool↗
PubMed2006

[Studies on fingerprints of Bupleurum chinense from Hebei Province with HPLC-UV].

OBJECTIVE: To establish a HPLC fingerprint analysis method for identification of Bupleurum chinense of Hebei Province and compare the fingerprints of Radix Bupleuri collected from different habitats so as to establish a sensitive and specific method for controlling the quality of Radix Bupleuri. METHODS: The HPLC-UV fingerprints of Radix Bupleuri from different habitats were obtained from Waters 1525 instruments. The HPLC separation was performed on a C18 analytical column gradient eluted with a mixture consisting of acetonitrile and water at a flow rate of 1.0 ml/min with UV detector at 203 nm. The temperature of column was 30 degrees C. RESULTS: The mutual mode of HPLC-UV fingerprints was set up, and the similar degrees to the crude drugs of different habitats were compared. CONCLUSION: It is simple and quick to differ Bupleurum chinense from different habitats with the method that can be used as a quality control item for Radix Bupleuri.

Bupleurum↗
PubMed2005

After the Indian Ocean tsunami: Singapore's contribution to the international disaster victim identification effort in Thailand.

This paper describes the international disaster victim identification (DVI) response mounted in Thailand, with particular reference to Singapore's contribution to this process, in the wake of the Asian tsunami of 26 December, 2004, which devastated parts of more than 10 countries in and around the Indian Ocean and claimed more than 200,000 lives. Although Singapore was unscathed by this natural calamity, over 30 Singaporean visitors were counted amongst the thousands of deceased victims, mostly in Thailand. The systematic application of forensic pathology, forensic dentistry, DNA profiling, and fingerprinting to human identification, especially of the bodies of various nationalities that were in advanced states of putrefaction, was crucial to the entire DVI process. The authors perceive that the resource implications arising from such a disaster, which is unprecedented in both its scale and reach in the international history of DVI, are immense. Forward planning, adequate funding and international cooperation are essential to mounting an effective response to any major mass disaster of the future.

Bayes Theorem↗
PubMed2003

A PCR-based method for identification of bifidobacteria from the human alimentary tract at the species level.

A polymerase chain reaction (PCR)-based method was developed for the identification of isolates of Bifidobacterium at the species level. Using two Bifidobacterium-specific primers directed against the 16S ribosomal gene (Bif164 and Bif662), a PCR product was obtained from the type strains of 12 different bifidobacterial species that have been found in the human alimentary tract. After purification of the PCR products, the DNA was restricted with five different restriction enzymes. The size of the different restriction fragments was used as a fingerprint for the identification of individual bifidobacterial species. The amplified ribosomal DNA restriction analysis method was subsequently used to speciate bifidobacterial isolates from child's feces and from an in vitro model of the large intestine.

Bifidobacterium↗
PubMed1992

Restriction endonuclease-fragment polymorphisms of oral viridans streptococci, compared by conventional and field-inversion gel electrophoresis.

Oral streptococci formerly classified as Streptococcus sanguis or Streptococcus mitis have recently been divided into four species. Two additional species have also been proposed for this group. Each species is genetically distinct, but they have many traits in common, which makes it difficult for clinical isolates to be identified by phenotypic tests. Genotypic comparison may provide an alternative approach. This study used DNA fingerprint analysis for comparison of genotypes of 21 reference strains--classified as Streptococcus gordonii, Streptococcus sanguis, Streptococcus oralis, "Streptococcus parasanguis", or "Streptococcus crista" in previous DNA hybridization studies--and 17 clinical and laboratory strains placed in those groups on the basis of phenotypic tests. HinDIII and PvuII digests were run in conventional horizontal agarose gels. SfiI digests of reference strains and two laboratory strains were run in field-inversion gels. Fingerprint patterns were compared by visual examination, cluster analysis of densitometric traces, and lane-matching software. Only two "S. crista" strains and two parent mutant lineages showed fingerprint patterns that were identical by visual examination. Fingerprint patterns of all other strains were unique. Cluster analysis results could not be considered valid, since replicate lanes in different gels were not grouped together. HinDIII and PvuII digests contained too many bands for correct matching by lane-matching software. SfiI digests were correctly matched by computer, with the same results as visual examination. Results indicate that the diversity of strains within these streptococcal species was too great to permit species identification by DNA fingerprint patterns. This genotypic diversity suggests that isolates from unrelated hosts may have been separate for long periods of time.(ABSTRACT TRUNCATED AT 250 WORDS)

Bacterial Proteins↗
PubMed2006

Secretory protein enrichment and analysis: an optimized approach applied on cancer cell lines using 2D LC-MS/MS.

Reliable methods for profiling secretory proteins are highly desirable for the identification of biomarkers of disease progression. Secreted proteins are often masked by high amounts of protein supplements in the culture medium. We have developed an efficient method for the enrichment and analysis of the secretome of different cancer cell lines, free of essential contaminants. The method is based on the optimization of cell incubation conditions in protein-free medium. Secreted proteins are concentrated and fractionated using a reversed-phase tC2 Sorbent, followed by peptide mass fingerprinting for protein identification. An average of 88 proteins were identified in each cancer cell line, of which more than 76% are known to be secreted, possess a signal peptide or a transmembrane domain. Given the importance of secreted proteins as a source for early detection and diagnosis of disease, this approach may help to discover novel candidate biomarkers with potential clinical significance.

Cell Culture Techniques↗