Proximate Composition, Antioxidant Properties, and Hepatoprotective Activity of Three Species of Shellfish of the Pacific Coast of Russia.

The objective of the present study was to investigate the proximate composition, antiradical properties and hepatoprotective activity of three species of shellfish, Corbicula japonica, Spisula sachalinensis, and Anadara broughtonii, from the coastal areas of Far East Russia. Biologically active peptides such as taurine (3.74 g/100 g protein) and ornithine (2.12 g/100 g protein) have been found in the tissues of A. broughtonii. C. japonica contains a high amount of ornithine (5.57 g/100 g protein) and taurine (0.85 g/100 g protein). The maximum DPPH and ABTS radical scavenging activity (36.0 µg ascorbic acid/g protein and 0.68 µmol/Trolox equiv/g protein, respectively) was determined for the tissue of C. japonica. The protein and peptide molecular weight distribution of the shellfish tissue water extracts was investigated using HPLC. It was found that the amount of low molecular weight proteins and peptides were significantly and positively correlated with radical scavenging activity (Pearson's correlation coefficient = 0.96), while the amount of high molecular weight proteins negatively correlated with radical scavenging activity (Pearson's correlation coefficient = -0.86). Hepatoprotective activity, measured by the survival rate of HepG2 hepatocytes after cotreatment with t-BHP, was detected for C. japonica. The highest protection (95.3 ± 2.4%) was achieved by the cold water extract of C. japonica at the concentration of 200 mg/mL. Moreover, oral administration of hot water extract of C. japonica to rats before the treatment with CCl4 exhibited a markedly protective effect by lowering serum levels of ALT and AST, inhibiting the changes in biochemical parameters of functional state of rat liver, including MDA, SOD, GSH and GST.

Extracting bio-zinc and taurine from Pinctada martensii meat.

Recently, intensive processing of marine resources has attracted considerable attention. In order to further utilize the byproducts of aquatic shellfish (Pinctada martensii meat) with high value, this study proposes a method of extracting zinc and taurine from P. martensii meat. Zinc was first extracted from P. martensii meat with an ultrasonic crusher, and then taurine was further extracted by ultrasonic-assisted water extraction from the remaining shellfish. After optimization, the biological zinc extraction rate reached 8.63%, and the taurine extraction rate reached 0.71%. Meanwhile, the parameters for cation exchange separation and taurine purification were optimized, in which the injection volume, pH value, and elution rate were set to 8.0 mL, 4.5, and 3.0 mL/min, respectively. Ultimately, the purity the extracted taurine reached 98.16%. This study provides a novel method for the extraction of biological zinc and taurine by deep processing of shellfish meat.

Six new polyhydroxylated steroids conjugated with taurine, microdiscusols A-F, from the Arctic starfish Asterias microdiscus.

Six new polyhydroxylated steroids conjugated with taurine, microdiscusols A-F, were isolated from the alcoholic extract of the Arctic starfish Asterias microdiscus. Three of them have been found to have additional sulfate groups in tetracyclic cores. The structures of new compounds were established by 1D and 2D NMR and HRESIMS techniques. The found steroid taurine conjugates resemble, by their structures, salts of bile acids and alcohols of lower vertebrates.

Improved Extraction Repeatability and Spectral Reproducibility for Liquid Extraction Surface Analysis-Mass Spectrometry Using Superhydrophobic-Superhydrophilic Patterning.

A major problem limiting reproducible use of liquid extraction surface analysis (LESA) array sampling of dried surface-deposited liquid samples is the unwanted spread of extraction solvent beyond the dried sample limits, resulting in unreliable data. Here, we explore the use of the Droplet Microarray (DMA), which consists of an array of superhydrophilic spots bordered by a superhydrophobic material giving the potential to confine both the sample spot and the LESA extraction solvent in a defined area. We investigated the DMA method in comparison with a standard glass substrate using LESA analysis of a mixture of biologically relevant compounds with a wide mass range and different physicochemical properties. The optimized DMA method was subsequently applied to urine samples from a human intervention study. Relative standard deviations for the signal intensities were all reduced at least 3-fold when performing LESA-MS on the DMA surface compared with a standard glass surface. Principal component analysis revealed more tight clusters indicating improved spectral reproducibility for a human urine sample extracted from the DMA compared to glass. Lastly, in urine samples from an intervention study, more significant ions (145) were identified when using LESA-MS spectra of control and test urine extracted from the DMA. We demonstrate that DMA provides a surface-assisted LESA-MS method delivering significant improvement of the surface extraction repeatability leading to the acquisition of more robust and higher quality data. The DMA shows potential to be used for LESA-MS for controlled and reproducible surface extraction and for acquisition of high quality, qualitative data in a high-throughput manner.

Two Major Bile Acids in the Hornbills, (24R,25S)-3α,7α,24-Trihydroxy-5ß-cholestan-27-oyl Taurine and Its 12α-Hydroxy Derivative.

Two major bile acids were isolated from the gallbladder bile of two hornbill species from the Bucerotidae family of the avian order Bucerotiformes Buceros bicornis (great hornbill) and Penelopides panini (Visayan tarictic hornbill). Their structures were determined to be 3α,7α,24-dihydroxy-5ß-cholestan-27-oic acid and its 12α-hydroxy derivative, 3α,7α,12α,24-tetrahydroxy-5ß-cholestan-27-oic acid (varanic acid, VA), both present in bile as their corresponding taurine amidates. The four diastereomers of varanic acid were synthesized and their assigned structures were confirmed by X-ray crystallographic analysis. VA and its 12-deoxy derivative were found to have a (24R,25S)-configuration. 13 additional hornbill species were also analyzed by HPLC and showed similar bile acid patterns to B. bicornis and P. panini. The previous stereochemical assignment for (24R,25S)-VA isolated from the bile of varanid lizards and the Gila monster should now be revised to the (24S,25S)-configuration.

Qualitative and Quantitative Analysis of the Major Constituents in Shexiang Tongxin Dropping Pill by HPLC-Q-TOF-MS/MS and UPLC-QqQ-MS/MS.

Shexiang Tongxin dropping pill (STP) is a traditional Chinese medicine formula that consists of total saponins of ginseng, synthetic Calculus bovis, bear gall, Venenum bufonis, borneol and Salvia miltiorrhiza. STP has been widely used in China and Southeast Asia for the treatment of cardiovascular diseases. In this study, a qualitative analytical method using high performance liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry was developed for identification of the major constituents in STP. Based on the retention time and MS spectra, 41 components were identified by comparison with reference compounds and literature data. Moreover, using ultra-performance liquid chromatography coupled with triple-quadrupole tandem mass spectrometry in multiple-reaction monitoring mode, we quantified 13 of the identified constituents (ginsenoside Rg1, ginsenoside Rk3, cinobufagin, arenobufagin, bufalin, resibufogenin, tanshinone IIA, taurine, tauroursodeoxycholic acid, taurocholic acid, cholic acid, deoxycholic acid, and chenodeoxycholic acid). These results suggest that this new approach is applicable for the routine analysis and quality control of STP products and provides fundamental data for further in vivo pharmacokinetical studies.

Isolation and Total Synthesis of Stolonines A-C, Unique Taurine Amides from the Australian Marine Tunicate Cnemidocarpa stolonifera.

Cnemidocarpa stolonifera is an underexplored marine tunicate that only occurs on the tropical to subtropical East Coast of Australia, with only two pyridoacridine compounds reported previously. Qualitative analysis of the lead-like enhanced fractions of C. stolonifera by LC-MS dual electrospray ionization coupled with PDA and ELSD detectors led to the identification of three new natural products, stolonines A-C (1-3), belonging to the taurine amide structure class. Structures of the new compounds were determined by NMR and MS analyses and later verified by total synthesis. This is the first time that the conjugates of taurine with 3-indoleglyoxylic acid, quinoline-2-carboxylic acid and ß-carboline-3-carboxylic acid present in stolonines A-C (1-3), respectively, have been reported. An immunofluorescence assay on PC3 cells indicated that compounds 1 and 3 increased cell size, induced mitochondrial texture elongation, and caused apoptosis in PC3 cells.

Optimal excitation and emission wavelengths to analyze amino acids and optimize neurotransmitters quantification using precolumn OPA-derivatization by HPLC.

We describe an analytical methodology to obtain high sensitivity and better resolution through the study of fluorometric excitation (λex) and emission (λem) spectrum wavelengths of OPA-amino acids. The spectrum emission study revealed a maximum signal peak at 450 nm for aspartate and glutamine. For glycine, taurine, and GABA, the maximum signal peak was at 448 and for glutamate at 452 nm. The remaining amino acids analyzed showed a maximum emission around 450 nm. The best signal obtained within the spectrum excitation experiments was using 229- to 450-nm λex-λem. The drawbacks observed at these wavelengths were a baseline drift and negative peaks occurrence. Thus, the excitation wavelength of 240 nm was chosen (240- to 450-nm λex-λem) as a compromise between a very good signal response and a baseline stability to resolve the 18 amino acids studied. Furthermore, this protocol was properly validated. On the other hand, the elution gradient program used for neuroactive amino acids (aspartate, glutamate, glycine, taurine and GABA) showed separation to the baseline, in a 15-min run in all of them. Other amino acids, up to 18, also exhibited a very good separation in a 25-min run. In conclusion, we propose the use of 240- to 450-nm λex-λem wavelengths, in OPA-amino acids analysis, as the most suitable protocol to obtain the best signal response, maintaining an optimum chromatographic resolution.

Reduction of bilirubin ditaurate by the intestinal bacterium Clostridium perfringens.

Bilirubin is degraded in the human gut by microflora into urobilinoids. In our study we investigated whether the bilirubin-reducing strain of Clostridium perfringens can reduce bilirubin ditaurate (BDT), a bile pigment of some lower vertebrates, without hydrolysis of the taurine moiety. C. perfringes was incubated under anaerobic conditions with BDT; reduction products were quantified by spectrophotometry and separated by TLC. Based on Rf values of BDT reduction products and synthetic urobilinogen ditaurate, three novel taurine-conjugated urobilinoids were identified. It is likely that bilirubin-reducing enzyme(s) serve for the effective disposal of electrons produced by fermentolytic processes in these anaerobic bacteria.

New N-acyl taurine from the sea urchin Glyptocidaris crenularis.

A new N-acyl taurine (1), together with a new natural product, l-(beta-D-ribofuranosyl)-1,2,4-triazole (4), and two known compounds (2 and 3), were isolated from the sea urchin, Glyptocidaris crenularis. The new N-acyl taurine was elucidated as 2-(5R,15S-dihydroxyeicosanoylamino) ethanesulfonic acid on the basis of spectroscopic (NMR, MS) analyses and the modified Mosher ester method. Compound 2 showed significant toxicity against brine shrimp larvae.

Enhancement of menadione stress tolerance in yeast by accumulation of hypotaurine and taurine: co-expression of cDNA clones, from Cyprinus carpio, for cysteine dioxygenase and cysteine sulfinate decarboxylase in Saccharomyces cerevisiae.

Taurine is known to function as a protectant against various stresses in animal cells. In order to utilize taurine as a compatible solute for stress tolerance of yeast, isolation of cDNA clones for genes encoding enzymes involved in biosynthesis of taurine was attempted. Two types of cDNA clones corresponding to genes encoding cysteine dioxygenase (CDO1 and CDO2) and a cDNA clone for cysteine sulfinate decarboxylase (CSD) were isolated from Cyprinus carpio. Deduced amino acid sequences of the two CDOs and that of CSD showed high similarity to those of CDOs and those of CSDs from other organisms, respectively. The coding regions of CDO1, CDO2, and CSD were subcloned into an expression vector, pESC-TRP, for Saccharomyces cerevisiae. Furthermore, to enhance the efficiency of synthesis of taurine in S. cerevisiae, a CDO-CSD fusion was designed and expressed. Expression of CDO and CSD proteins, or the CDO-CSD fusion protein was confirmed by Western blot analysis. HPLC analysis showed that the expression of the proteins led to enhancement of the accumulation level of hypotaurine, a precursor of taurine, rather than taurine. The yeast cells expressing corresponding genes showed tolerance to oxidative stress induced by menadione, but not to freezing-thawing stress.

Short communication: rapid preparation of preventive and therapeutic whole-killed retroviral vaccines using the microbicide taurine chloramine.

A current urgent priority is to develop microbicides and vaccines to combat retroviruses like human immunodeficiency virus (HIV). We show that the cysteine-selective natural compound, taurine chloramine (T-NCl), can be effective in this task. A number of proteins in all retroviruses contain highly conserved cysteine-rich regions that are essential for infection and replication. Our data show that by targeting these essential cysteine residues, T-NCl (2 or 5 mM) acts as a highly effective and safe microbicide that fully blocks the infectivity of high HIV-1 titers (10(6) TCID(50) units/ml) but is not injurious to eukaryotic cells. We also demonstrate that T-NCl can be used to prepare a highly effective whole-killed vaccine against murine AIDS (MAIDS) that shows both preventive and therapeutic efficacy. The vaccine consists of a T-NCl-inactivated retrovirus suspension in host cell lysate. The novelty of our approach lies in the ease and speed of vaccine preparation and its avoidance of harsh inactivation or purification steps that can alter native viral conformation. Our approach is therefore likely to overcome a number of intractable obstacles to the preparation of an effective whole-killed HIV vaccine, such as surviving infective viral particles, rapid viral mutation rates, numerous viral strains, and harsh purification steps. Our approach may also permit the rapid preparation of autologous, or custom-made, vaccines for individual patients.

Pharmacokinetics of taurolidine following repeated intravenous infusions measured by HPLC-ESI-MS/MS of the derivatives taurultame and taurinamide in glioblastoma patients.

BACKGROUND AND OBJECTIVE: Taurolidine is known to have antimicrobial activity. Furthermore, at lower concentrations, it has been found to exert a selective antineoplastic effect in vitro and in vivo. The aim of this study was to investigate the pharmacokinetics of taurolidine in vivo following repeated intravenous infusion in a schedule used for the treatment of glioblastoma. As a prerequisite, the pharmacokinetics of taurolidine in human blood plasma and whole blood in vitro was investigated. PATIENTS AND METHODS: The pharmacokinetics of taurolidine and its derivatives taurultame and taurinamide were investigated in human blood plasma and in whole blood in vitro using blood from a healthy male volunteer. During repeated intravenous infusion therapy with taurolidine, plasma samples were taken every hour for a period of 13 hours per day in seven patients (three male, four female; mean age 48.4 +/- 12.8 years, range 27-66 years) with a glioblastoma. Following dansyl derivatisation, the concentrations of taurultame and taurinamide were determined using a new method based on high-performance liquid chromatography (HPLC) online coupled to electrospray ionisation tandem mass spectrometry (ESI-MS/MS) in the multiple reaction monitoring mode. Under the experimental conditions used, taurolidine could not be determined directly and was back-calculated from the taurultame and taurinamide values. RESULTS: The new HPLC-ESI-MS/MS method demonstrated high accuracy and reproducibility. In vitro plasma concentrations of taurultame and taurinamide remained constant over the incubation period. In whole blood in vitro, a time-dependent formation of taurinamide was observed. At the start of the incubation, the taurultame-taurinamide ratio (TTR) was 0.95 at an initial taurolidine concentration of 50 microg/mL, and 1.69 at 100 microg/mL. The concentration of taurultame decreased at the same rate as the taurinamide concentration increased, showing logarithmic kinetics. The calculated taurolidine concentration remained largely constant over the 6-hour incubation period. During repeated infusions in patients, calculated plasma concentrations of taurolidine showed a strong increase after the start of each infusion and continued to increase until the end of infusion, followed by a rapid decline. The TTR was found to fluctuate between 0.1 and 0.3, depending on the relation to the previous or next infusion period. The volume of distribution was markedly higher for taurolidine, taurultame and taurinamide than the plasma volume. CONCLUSIONS: Taurolidine displayed a stable pattern of derivatives in plasma in vitro, whereas in whole blood, a time- and concentration-dependent conversion was apparent. In patients, the calculated average taurolidine plasma concentration, achieved with the repeated infusion regimen, was in the antineoplastic-effective concentration range. The tissue concentrations of taurolidine and taurultame are expected to be higher than the plasma concentrations, taking into account the calculated volumes of distribution. Repeated infusion of taurolidine is the therapeutically adequate mode of administration for the indication of glioblastoma.

Site-specific sampling of taurine from rat brain followed by on-line sample pre-concentration, throughout in-capillary derivatization and capillary electrophoresis.

A method of pinpoint-sampling followed by on-line pre-concentration of the sample, throughout in-capillary derivatization and capillary electrophoretic separation was evaluated by demonstrating the detection of taurine, 2-aminoethanesulfonic acid at a specific location of a rat brain. The direct sampling of taurine from the rat brain was accomplished by using voltage injection associated with two kinds of driving forces, electrophoretic flow and electroosmotic flow (EOF). The capillary tube (75 microm of inner diameter x 375 microm of outer diameter) of the capillary electrophoresis (CE) apparatus was already filled with a CE run buffer, viz., 40 mM phosphate-borate buffer (pH 10) containing 2mM o-phthalaldehyde (OPA)/N-acetylcysteine (NAC) as the derivatization reagent. One end of a platinum wire (0.5mm o.d.), used as the anode, and the inlet end of capillary tube (from which a 1.0 cm long polyimide coating was removed), were pricked down onto the surface of either the cerebrum or cerebellum of a rat brain at a location of very small dimension. When a low voltage (5 kV, 30s) was applied, taurine began to move from the rat brain into the capillary tube, and, simultaneously, electric focusing of taurine occurred by the action of "the pH-junction effect" at the inlet end of the capillary tube. After completing the injection, both the platinum wire and capillary tube were detached from the brain and dipped into the run buffer in an anode reservoir filed with the same solution as that in the capillary tube for the CE apparatus. Then, by applying a high voltage (20 kV) between the ends of the capillary tube, taurine was automatically derivatized to yield the fluorescent derivative, separated and detected with fluorescence (E(x)=340 nm, E(m)=455 nm) during migration throughout the capillary tube. The migration profiles obtained from cerebrum and cerebellum appeared to be different, but the peak corresponding to taurine was identified on both electropherograms. The efficacy of the present method including sample on-line pre-concentration prior to throughout in-capillary derivatization CE was first verified with several preliminary experiments by using samples of taurine in water, saline and a piece of 1.5% agar-gel block, as an alternate standard for the rat brain used in this study.

Current studies on anti-endotoxic chemical components of traditional Chinese medicine in China.

Endotoxin (ET) was found to have wide bioactivities and ET antagonists have become the pop research topic in life science. The chemical components of traditional Chinese medicine (TCM) were the substance basis of its pharmacology. This review demonstrated the study state of about 18 chemical components from TCM, eg, organic acids of Radix Isatidis, anisodamine, matrine, tetramethypyrazine, colchicine, and glycine, etc, which showed anti-endotoxin effects through different routes. But now the most of them were limited to the laboratory. In the future, the trends of development should not only enlarge the range of research, but also strengthen the clinical study.

[Improvement of a method for separation and determination of amino acids].

A method for separation and determination of amino acids using pre-column derivatization with 6-aminoquinoline-N-hydroxysuccinimido-carbamate (AccQ. Tag) was improved. The technique developed for rapid separation and determination of amino acids by AccQ. Tag method is described. The 18 amino acids and taurine are rapidly separated and determined by AccQ. Tag special chromatographic column(3.9 mm i.d. x 150 mm) within 17.5 min(the analysis period is 22.5 min) by changing the flow rate from 1.0 mL/min to 2.0 mL/min. In place of the AccQ. Tag special chromatographic column, the Nova-Pak C18 column(3.9 mm i.d. x 150 mm), Nova-Pak C18 column(4.6 mm i.d. x 150 mm), Symmetry C18 column(3.9 mm i.d. x 150 mm) or Waters Xterra RP-18 column, can be used to separate and determine the amino acids rapidly in the same manner. But among them the Symmetry C18 column is the best substitute for AccQ. Tag column. The separation time is 14.5 min and the analysis period is 20.0 min only. The price is much cheaper than the AccQ. Tag column.

Improvement of impaired memory in mice by taurine.

Taurine was extracted from Pegasus laternarius Cuvier to study its effects on learning and memory in mice. Mice were treated with different doses of taurine (10 mg/kg, 20 mg/kg, 40 mg/kg). The mice were treated with various chemical agents (pentobarbital, cycloheximide, sodium nitrite, alcohol) to disrupt the normal memory process. We measured the effect of taurine on step-down latency (SDL) and escape latency (EL) in a passive avoidance task after 10 or 30 days. Treatment with taurine alone did not change either SDL or EL. Taurine protected mice from the memory disruption induced by alcohol, pentobarbital, sodium nitrite, and cycloheximide but had no obvious effect on motor coordination, exploratory activity, or locomotor activity as measured using the rota-rod test and the hole board test. We conclude that taurine can be effective in attenuating the amnesia produced by alcohol, pentobarbital, cycloheximide, and sodium nitrite without compromising the behavioral aspects of the animals tested.

Exclusion and retention of compensatory kosmotropes by HPLC columns.

With water as the elution solvent, zwitterionic solutes and polyols were retained on HPLC columns, more than was water, by totally hydrophobic packing materials. Relative retentions were systematically affected by oxygen functional groups in the packing material, explicable as specific retention of water. Reproducible elution sequences of 20 solutes at a variety of hydrophobic surfaces (aromatic and both long- and short-alkyl aliphatic surfaces) showed there is a general process, consistent with interactions with hydration water at the surface having solvent properties distinct from bulk water. Early eluting solutes included glycine, sarcosine and taurine. Glycine betaine followed both these and N,N-dimethylglycine. The natural betaines propionobetaine and dimethylsulfoniopropionate also preceded glycine betaine. Dimethylsulfoxide was strongly retained, as (to a lesser extent) was proline betaine. Polyols eluted in the sequence sorbitol, trehalose, glycerol. Changes in the chemical nature of the surface or base material affected relative retentions of water and solutes. The presence of hydrogen-bonding functions increased retention of polyols, as well as water, relative to zwitterionic solutes. Specific effects retention, constraining models based on the formation of low-density water.