Effect of cooling water chlorination on primary productivity of entrained phytoplankton at Kalpakkam,east coast of India

We studied the effect of various concentrations of chlorine, used in the cooling system of Madras Atomic Power Station (MAPS), on the primary productivity of entrained phytoplankton from October 1988 to December 1990. The rates of primary productivity (gross) were lower at the discharge point than in the intake area. The reduction in productivity ranged from 30 to 70% at low-dose chlorination (residual chlorine 0.05–0.20 mg l⁻¹ at the discharge point). During shock-dose chlorination, when residual chlorine at the discharge point ranged from 1.10 to 1.50 mg 1⁻¹ the reduction in productivity was 80–83%. A reduction of 16–17% was recorded without chlorination.     

Growth, natural relationships, cellular fatty acids and metabolic adaptation of sulfate-reducing bacteria that utilize long-chain alkanes under anoxic conditions

Natural relationships, improvement of anaerobic growth on hydrocarbons, and properties that may provide clues to an understanding of oxygen-independent alkane metabolism were studied with two mesophilic sulfate-reducing bacteria, strains Hxd3 and Pnd3. Strain Hxd3 had been formerly isolated from an oil tank; strain Pnd3 was isolated from marine sediment. Strains Hxd3 and Pnd3 grew under strictly anoxic conditions on n-alkanes in the range of C₁₂–C₂₀ and C₁₄–C₁₇, respectively, reducing sulfate to sulfide. Both strains shared 90% 16 S rRNA sequence similarity and clustered with classified species of completely oxidizing, sulfate-reducing bacteria within the δ-subclass of Proteobacteria. Anaerobic growth on alkanes was stimulated by α-cyclodextrin, which served as a non-degradable carrier for the hydrophobic substrate. Cells of strain Hxd3 grown on hydrocarbons and α-cyclodextrin were used to study the composition of cellular fatty acids and in vivo activities. When strain Hxd3 was grown on hexadecane (C₁₆H₃₄), cellular fatty acids with C-odd chains were dominant. Vice versa, cultures grown on heptadecane (C₁₇H₃₆) contained mainly fatty acids with C-even chains. In contrast, during growth on 1-alkenes or fatty acids, a C-even substrate yielded C-even fatty acids, and a C-odd substrate yielded C-odd fatty acids. These results suggest that anaerobic degradation of alkanes by strain Hxd3 does not occur via a desaturation to the corresponding 1-alkenes, a hypothetical reaction formerly discussed in the literature. Rather an alteration of the carbon chain by a C-odd carbon unit is likely to occur during activation; one hypothetical reaction is a terminal addition of a C₁ unit. In contrast, fatty acid analyses of strain Pnd3 after growth on alkanes did not indicate an alteration of the carbon chain by a C-odd carbon unit, suggesting that the initial reaction differed from that in strain Hxd3. When hexadecane-grown cells of strain Hxd3 were resuspended in medium with 1-hexadecene, an adaptation period of 2 days was observed. Also this result is not in favor of an anaerobic alkane degradation via the corresponding 1-alkene. Received: 25 June 1998 / Accepted: 29 July 1998     

Pseudomonas sp. Strain 273, an Aerobic α,ω-DichloroalkaneDegrading Bacterium

A gram-negative, aerobic bacterium was isolated from soil; this bacterium grew in 50% (vol/vol) suspensions of 1,10-dichlorodecane (1,10-DCD) as the sole source of carbon and energy. Phenotypic and small-subunit ribosomal RNA characterizations identified the organism, designated strain 273, as a member of the genus Pseudomonas. After induction with 1,10-DCD, Pseudomonas sp. strain 273 released stoichiometric amounts of chloride from C₅ to C₁₂ α,ω-dichloroalkanes in the presence of oxygen. No dehalogenation occurred under anaerobic conditions. The best substrates for dehalogenation and growth were C₉ to C₁₂ chloroalkanes. The isolate also grew with nonhalogenated aliphatic compounds, and decane-grown cells dechlorinated 1,10-DCD without a lag phase. In addition, cells grown on decane dechlorinated 1,10-DCD in the presence of chloramphenicol, indicating that the 1,10-DCD-dechlorinating enzyme system was also induced by decane. Other known alkane-degrading Pseudomonas species did not grow with 1,10-DCD as a carbon source. Dechlorination of 1,10-DCD was demonstrated in cell extracts of Pseudomonas sp. strain 273. Cell-free activity was strictly oxygen dependent, and NADH stimulated dechlorination, whereas EDTA had an inhibitory effect.     

Polyphasic approach for assessing changes in an autochthonous marine bacterial community in the presence of Prestige fuel oil and its biodegradation potential

A laboratory experiment was conducted to identify key hydrocarbon degraders from a marine oil spill sample (Prestige fuel oil), to ascertain their role in the degradation of different hydrocarbons, and to assess their biodegradation potential for this complex heavy oil. After a 17-month enrichment in weathered fuel, the bacterial community, initially consisting mainly of Methylophaga species, underwent a major selective pressure in favor of obligate hydrocarbonoclastic microorganisms, such as Alcanivorax and Marinobacter spp. and other hydrocarbon-degrading taxa (Thalassospira and Alcaligenes), and showed strong biodegradation potential. This ranged from >99% for all low- and medium-molecular-weight alkanes (C₁₅–C₂₇) and polycyclic aromatic hydrocarbons (C₀- to C₂- naphthalene, anthracene, phenanthrene, dibenzothiophene, and carbazole), to 75–98% for higher molecular-weight alkanes (C₂₈–C₄₀) and to 55–80% for the C₃ derivatives of tricyclic and tetracyclic polycyclic aromatic hydrocarbons (PAHs) (e.g., C₃-chrysenes), in 60 days. The numbers of total heterotrophs and of n-alkane-, aliphatic-, and PAH degraders, as well as the structures of these populations, were monitored throughout the biodegradation process. The salinity of the counting medium affects the counts of PAH degraders, while the carbon source (n-hexadecane vs. a mixture of aliphatic hydrocarbons) is a key factor when counting aliphatic degraders. These limitations notwithstanding, some bacterial genera associated with hydrocarbon degradation (mainly belonging to α- and γ-Proteobacteria, including the hydrocarbonoclastic Alcanivorax and Marinobacter) were identified. We conclude that Thalassospira and Roseobacter contribute to the degradation of aliphatic hydrocarbons, whereas Mesorhizobium and Muricauda participate in the degradation of PAHs.     

Impact of Anaerobic and Aerobic Processes on PolyChloroBiphenyl Removal in Contaminated Sewage Sludge

Aerobic and anaerobic biodegradation of six priority PCBs was investigated in continuous stirred tank reactors fed with naturally contaminated sewage sludge. Anaerobic and aerobic abiotic losses were higher for the lightly chlorinated PCBs but remained for all PCBs below 20%. Under strict methanogenic conditions, PCB removals were about 40% whatever PCB molecular weight or their degree of chlorination. However, considering abiotic losses, the heaviest PCBs were more efficiently anaerobically biodegraded probably because of higher dechlorination rates. The aerating sludge process enhanced removal of the lightest chlorinated PCBs from 40% up to 100%, while removal rates of the heaviest PCBs remained around 40%. Although the mesophilic aerobic process exhibits better removal efficiencies because of operating conditions, the results suggest that PCB biodegradation was strongly limited by their bioavailability in naturally contaminated sludge, under both redox conditions. Indeed, since PCB removal was closely linked to the solid reduction rates, PCB bioavailability was likely the limiting factor for biodegradation. As a consequence, the raw PCB concentrations (in mg kg^–1_{dry weight}) which are concerned by legislative procedures did not decrease sufficiently in both processes to reach a limit value fulfilling the current French/European regulation about PCB contents in sewage sludge before spreading on agricultural land.     

Increased Immunoreactivities of Cleaved αII-Spectrin and Cleaved Caspase-3 in the Aged Dog Spinal Cord

The activation of caspase-3 is considered to be a reliable marker for apoptotic cell death, and a 120-kDa fragment of αII-spectrin is generated by caspase-3 mediated cleavage of this structural protein. In the present study, we compared cleaved αII-spectrin (120-kDa) and cleaved caspase-3-immunoreactive cells and their protein levels in the cervical (C₅–C₆) and lumbar (L₃–L₄) levels of the spinal cord in adult (1–2 year-old) and aged (10–12 year-old) dogs (German shepherds). Weak cleaved αII-spectrin and cleaved caspase-3 immunoreactivity was found in neurons of the adult group; however, their immunoreactivity was distinctively increased in the neuronal cytoplasm in the aged group compared to those in the adult group, although the distribution pattern of their neurons was similar between the adult and age group. In addition, cleaved αII-spectrin and cleaved caspase-3 levels in the aged spinal cord were markedly increased compared to those in the adult group. These findings suggest that the increases of cleaved αII-spectrin and cleaved caspase-3 immunoreactivity may be related to aging of the spinal cord in dogs.     

De novo formation of organochlorines in a sewage treatment plant

The de novo formation of organochlorines was observed in a municipalsewage treatment plant. Due to this formation, the amount of organically boundhalogens (AOX) increased 15-fold inside the sewage treatment plant. Per day,more than 6 kg of organically-bound chlorine were produced. Thisformation is not based on a metabolism of present organochlorines, it is a denovo formation out of inorganic chloride and organic substrates. The AOXtriggerconcentration in sewage sludge in Germany is 500 mgkg^–1 and was sometimes exceeded by a factor of 10. Noknown anthropogenic organohalogens were found which could explain the elevatedAOX concentrations. Instead many chlorinated compounds could be identifiedwhichwere not known to be of anthropogenic origin. The compound with the highestconcentration was the 3,4-dichlorophenylacetic acid (3,4-CPAc). In one case,more than 1 g kg^–1 of this compound was detected.A slaughterhouse that emits phenylacetic acid is probably the origin of thatformation. In model experiments phenylacetic acid was chlorinated with HOCl butchlorinated phenylacetic acids other than 3,4-CPAc were found. Therefore it canbe excluded that the chlorination in the sewage treatment plant takes place byan abiotic reaction with hypochlorite that might have been introduced there. Weassume that the occurring microorganisms are responsible for the de novoformation in the sewage treatment plant. The obtained knowledge could also beuseful to understand natural chlorination processes.     

The Abiotic Formation of Hydrocarbons from Dissolved CO2 Under Hydrothermal Conditions with Cobalt-Bearing Magnetite

Conversion of CO₂ to organic compounds in hydrothermal systems is important in understanding prebiotic chemical evolution leading to the origin of life. However, organic compounds with carbon number of more than 3 have never been produced from dissolved CO₂ in simulated hydrothermal experiments. In this paper, we report that not only CH₄, C₂H₆ and C₃H₈, but also n-C₄H₁₀ and n-C₅H₁₂ could be produced from dissolved CO₂ and H₂ in the presence of cobalt-bearing magnetite at 300°C and 30 MPa. It is shown that unbranched alkanes in Anderson–Schulz–Flory distribution were the dominant hydrocarbon products produced from dissolved CO₂ catalyzed by cobalt-bearing magnetite under certain hydrothermal conditions. It is proposed that magnetite with other transition metals may act potentially as effective mineral catalysts for abiotic formation of organic compounds from dissolved CO₂ in hydrothermal systems.     

A sensitive bioassay for spider mite (Tetranychus urticae) repellency: a double bond makes a difference

Choice bioassays were used to determine repellency of homologous n-alkanes (C₈H₁₈–C₂₁H₄₄) to spider mites. When tested at 400 μg/cm², the C₁₅–C₁₉ alkanes were highly repellent; the C₁₆ n-alkane, n-hexadecane, was most repellent. Subsequently the EC₅₀ values, the concentration at which 50% of the mites were repelled, were determined for the C₁₅–C₁₉ n-alkanes and their analogous 1-n-alkenes (C₁₅H₃₀–C₁₉H₃₈). The EC₅₀ value for 1-heptadecene, the C₁₇ 1-n-alkene, was the lowest observed. Except for the 17-carbon hydrocarbons, the EC₅₀ values for the n-alkanes were less than those for their analogous 1-n-alkenes. Depending on the compounds evaluated, there was as much as a six-fold difference of repellency between an n-alkane its analogous 1-n-alkene. Thus, the bioassay has sufficient sensitivity to detect behavioral differences associated with the presence or absence of a single double bond. The EC₅₀ values for the most repellent hydrocarbons were similar to that reported for 2,3-dihydrofarnesoic acid, a naturally occurring repellent isolated from trichome secretions of a wild tomato, Solanum habrochaites, and also were similar to concentrations used to evaluate arthropod repellents. Consequently, this bioassay may be useful for providing a better understanding of the relationships between structures and activities of natural products that are repellent to spider mites.     

Voltage-dependent calcium channels in young and old human red cells

Presence of subtypes of voltage-dependent Ca channels was investigated in young and old human red cells, employing immunological and flux-kinetics methods. Western blots showed specific reaction toward polyclonal rabbit antibodies raised against a highly conserved residue of α_1C, subunit of high-voltage activated Ca channels (pan α₁) and against conserved residues of α_1C and α_1E subunits. No specific reaction was detected with antibodies against conserved residues of α_1A, α_1B, or α_1D subunits. Only a single band (approx 260 kDa) was revealed on anti-pan α_1A or anti-α_1E blots, whereas two bands (200 and 230 kDa) were detected by α_1C exposure, Blots from old cells always showed diminished band intensity. Channel activity was assessed by studying the effect of voltage-dependent Ca channels blockers' under conditions likely to alter the red cell membrane potential, through incubation in media of different composition. In a 150 mM NaCl+5 mM KCl medium, blockers of L-, R-, and Q-type caused a 15–50% reductions of ⁴⁵Ca influx into cells, which had the Ca pump inactivated by either exhaustive adenosine triphosphate depletion or presence of vanadate plus substrates. Additionally, some P/Q-and N-type blockers also reduced Ca influx to various extents (25–60%). Old cells were generally insensitive to L-type but not to non-L-type, blockers. Raising external K to about 70–80 mM reduced by 50–100% inhibition by L-type blockers. Incubation in a gluconate medium containing 150 mM Na+5 mM K practically abolished the action of L-type blockers, but only slightly reducing that by non-L-type. The results, clearly demonstrate presence of L- and R-type Ca channels, apparently occurring in different functional states in young and old cells. Other non-L-type channels were also demonstrated only by pharmacological means. A possible physiological role for these channels is discussed.     

Gratuitous dechlorination of chloroethanes by methanogenic granular sludge

The dechlorinating activity of a methanogenic granular sludge from a methanol-fed upflow anaerobic sludge blanket reactor was investigated with chlorinated ethanes. This unadapted methanogenic consortium degraded all chloroethanes tested. The product formation rates decreased with the number of chlorine substituents. The more highly chlorinated ethanes were also converted, although at a lower rate, in the presence of autoclaved (dead) sludge, indicating the involvement of reduced heat-stable cofactors like vitamin B₁₂ and F₄₃₀. Direct chemical dechlorination of hexa-, penta- and tetrachloroethanes was also observed in medium without sludge, although at a much lower rate. The results show the importance of cometabolic and abiotic (chemical) conversions for the transformation of chlorinated ethanes by the methanogenic consortium. The types of reaction and the products formed were correlated with the Gibbs free-energy change (ΔG ^0′). Reductive hydrogenolysis and dichloroelimination were important dechlorinating mechanisms. Generally, these reactions have a higher ΔG ^0′ value than dehydrochlorination reactions, which occurred less frequently during the transformation of chloroethanes by the methanogenic granular sludge. Received: 8 June 1998 / Received revision: 7 September 1998 / Accepted: 13 September 1998     

Production of flavour compounds by yogurt starter cultures

The present work studied the production of carbonyl compounds and saturated volatile free fatty acids by pure cultures of Streptococcus thermophilus and Lactobacillus bulgaricus, and by starter cultures for Bulgarian yogurt during cultivation and cooling. The mixed cultures formed volatile aromatic compounds more actively than the pure cultures. A guiding factor in the preparation of the starter cultures was the biochemical activity of Lactobacillus bulgaricus in synthesizing the major carbonyl compounds, acetaldehyde, diacetyl and the volatile fatty acids C₂–C₁₀. The activity of the yogurt cultures in synthesizing carbonyl compounds was at its highest during milk coagulation and cooling, up to 7 h. However, maximum concentration was reached by 22–31 h. In the cooled 22–h starter cultures, acetaldehyde predominated (1415.0–1734.2 μg per 100 g) followed by diacetyl (165.0–202.0 μg per 100 g), acetoin (170.0–221.0 μg per 100 g), acetone (66.0–75.5 μg per 100 g), ethanol (58.0 μg per 100 g), and butanone-2 (3.6–3.8 μg per 100 g). The thermophilic streptococcus and lactobacillus cultures, and the starter cultures contained predominantly acetic, butyric and caproic acids. Received 19 June 1997/ Accepted in revised form 10 January 1998     

Influence of growth regulators and elicitors on cell growth and α-tocopherol and pigment productions in cell cultures of <Emphasis Type="Italic">Carthamus tinctorius</Emphasis> L.

The present study examined the effects of plant growth hormones, incubation period, biotic (Trametes versicolor, Mucor sp., Penicillium notatum, Rhizopus stolonifer, and Fusarium oxysporum) and abiotic (NaCl, MgSO₄, FeSO₄, ZnSO₄, and FeCl₃) elicitors on cell growth and α-tocopherol and pigment (red and yellow) productions in Carthamus tinctorius cell cultures. The cell growth and α-tocopherol and pigment contents improved significantly on Murashige and Skoog (MS) liquid medium containing 50.0 μM α-naphthalene acetic acid (NAA) and 2.5 μM 6-Benzyladenine (BA) at 28 days of incubation period. Incorporation of T. versicolor (50 mg l⁻¹) significantly enhanced the production of α-tocopherol (12.7-fold) and red pigment (4.24-fold). Similarly, supplementation of 30 mg l⁻¹ T. versicolor (7.54-fold) and 70 mg l⁻¹ Mucor sp. (7.40-fold) significantly increased the production of yellow pigment. Among abiotic elicitors, NaCl (50–70 mg l⁻¹) and MgSO₄ (10–30 mg l⁻¹) significantly improved production of α-tocopherol (1.24-fold) and red pigment (20-fold), whereas yellow pigment content increased considerably by all the abiotic elicitor treatments. Taken together, the present study reports improved productions of α-tocopherol and the pigment as a stress response of safflower cell cultures exposed to these elicitors.     

Structural and immunochemical characterization of β-1,2-linked mannobiosyl phosphate residue in the cell wall mannan of Candida glabrata

A mannan of Candida glabrata IFO 0622 digested by Arthrobacter exo-α-mannosidase and a β-1,2-linked mannobiose obtained from the parent mannan by acid treatment was analyzed using ¹³C nuclear magnetic resonance spectroscopy. The results show that the β-1,2-linked mannobiosyl residue is esterified to a phosphate group through position C-1 in the α-configuration, Manβ1– 2Manα1–HPO₃–. The results of immunochemical assays of these mannans using the commercial antigenic factor sera of the genus Candida (Candida Check, Iatron) indicate that the main recognition site of serum no. 6 in this kit is the mannotetraosyl side-chain Manβ1–2Manα1– 2Manα1–2Man in C. glabrata mannan and also suggest that the phosphate-containing unit (such as Manβ1– 2Manα1–HPO₃– in this mannan) behaves as one of the antigenic determinants of serum no. 6, but not of serum no. 5. Therefore, the present and previous findings indicate that serum no. 5 recognizes relatively longer β-1,2-linked oligomannosyl side-chains, Manβ1–[2Manβ1–]_n 2Man (n = 1–6), attached to the phosphate groups previously observed in the cell wall mannans of Candida albicans, Candida stellatoidea, and Candida tropicalis. Received: 18 March 1997 / Accepted: 16 September 1997     

Effects of Carbon Source, Carbon Concentration, and Chlorination on Growth Related Parameters of Heterotrophic Biofilm Bacteria

Abstract To investigate growth of heterotrophic biofilm bacteria, a model biofilm reactor was developed to simulate a drinking water distribution system. Controlled addition of three different carbon sources (amino acids, carbohydrates, and humics) at three different concentrations (500, 1,000, and 2,000 ppb carbon) in the presence and absence of chlorine were used in separate experiments. An additional experiment was run with a 1:1:2 mixture of the above carbon sources. Biofilm and effluent total and culturable cells in addition to total and dissolved organic carbon were measured in order to estimate specific growth rates (SGRs), observed yields, population densities, and bacterial carbon production rates. Bacterial carbon production rates (μg C/L day) were extremely high in the control biofilm communities (range = 295–1,738). Both growth rate and yield decreased with increasing carbon concentrations. Therefore, biofilm growth rates were zero-order with respect to the carbon concentrations used in these experiments. There was no correlation between growth rate and carbon concentration, but there was a significant negative correlation between growth rate and biofilm cell density (r=−0.637, p= 0.001 control and r=−0.57, p= 0.021 chlorinated biofilms). Growth efficiency was highest at the lowest carbon concentration (range = 12–4.5%, amino acids and humics respectively). Doubling times ranged from 2.3–15.4 days in the control biofilms and 1–12.3 days in the chlorinated biofilms. Growth rates were significantly higher in the presence of chlorine for the carbohydrates, humics, and mixed carbon sources (p= 0.004, < 0.0005, 0.013, respectively). The concept of r/K selection theory was used to explain the results with respect to specific growth rates and yields. Humic removal by the biofilm bacteria (78% and 56% for the control and chlorinated biofilms, respectively) was higher than previously reported literature values for planktonic bacteria. A number of control experiments indicated that filtration of drinking water was as effective as chlorination in controlling bacterial biofilm growth. Received: 26 March 1999; Accepted: 3 August 1999; Online Publication: 15 February 2000     

Mutagenicity on chlorination of products formed by ozonation of naphthoresorcinol in water

The mutagenicity of products formed by chlorination after ozonation of naphthoresorcinol in aqueous solution was assayed with Salmonella typhimurium strains TA98 and TA100 in the presence and absence of S9 mix from phenobarbital- and 5,6-benzoflavone-induced rat liver. Ozonated and subsequently chlorinated naphthoresorcinol was directly mutagenic, as was ozonated naphthoresorcinol, in both strains tested. The mutagenic activity at chlorination with 8 equivalents of chlorine per mole of naphthoresorcinol after ozonation was markedly higher than that at only ozonation. Of the identified ozonation products of naphthoresorcinol, muconic acid, after chlorination with 2 or 4 equivalents of chlorine per mole of the compound, induced direct mutagenicity against TA98 and TA100. The chlorination of glyoxal with 0.5 and 1 chlorine equivalents per mole of the compound was shown to produce direct mutagenicity toward TA98. The identification of the chlorination products of these compounds is also discussed.     

Anaerobic reductive dehalogenation of polychlorinated dioxins

Polychlorinated dibenzo-p-dioxins and -furans (PCDD/Fs) are among the most harmful environmental contaminants. Their widespread distribution due to unintentional or unknown release coincides with environmental persistence, acute and chronic toxicity to living organisms, and long-term effects due to the compounds’ tendency for bioaccumulation and biomagnification. While microbial aerobic degradation of PCDD/Fs is mainly reported for the turnover of low chlorinated congeners, this review focuses on anaerobic reductive dehalogenation, which may constitute a potential remediation strategy for polychlorinated compounds in soils and sediments. Microorganisms in sediments and in microcosms or enrichment cultures have been shown to be involved in the reductive dechlorination of dioxins. Bacteria related to the genus Dehalococcoides are capable of the reductive transformation of dioxins leading to lower chlorinated dioxins including di- and monochlorinated congeners. Thus, reductive dehalogenation might be one of the very few mechanisms able to mediate the turnover of polychlorinated dioxins by reducing their toxicity and paving the way for a subsequent breakdown of the carbon skeleton.     

Effect of triterpenoid glycosides on α- and β-amylase activity and total protein content in wheat seedlings

Influence of the aleanolic acid glycosides from Silphium perfoliatum L. (silphioside B, C, E and G) and their progenins on the amylase activity and total protein content in wheat seedlings was studied. Treatment of the Triticum aestivum L. seeds with 1–10 μM water solutions of mono- and diglycosides (mono- and bisdesmosines) elevated the α-amylase and total amylase activities in seedlings. Silphioside E containing three glucose moieties in its molecule did not change α-amylase activity, but it did if bis-triglycoside acetylated carbohydrate (as in silphioside C). Effects of 5–10 μM solutions of the active glycosides was comparable with that of exogenous gibberellin A₃ and 6-benzylaminopurine.     

Formulation Development of Morphine Sulfate Sustained-Release Tablets and Its Bioequivalence Study in Healthy Thai Volunteers

The objectives of this study were to develop morphine sulfate sustained-release tablet formulations and to evaluate the bioequivalence compared with a commercial brand. The physicochemical properties of the formulated and commercial tablets were determined and compared. The bioequivalence investigation was carried out in 15 healthy male volunteers who received a single dose in a randomized two-way crossover design. After dosing, serial blood samples were collected for a period of 24 h. Morphine concentration was assayed by high-performance liquid chromatography with electrochemical detector. The log-transformed C _max and AUC_s were statistically compared by analysis of variance, and the 90% confidence intervals (CIs) of the ratio of the log-transformed C _max and AUC_s between the most promising developed formulation and the commercial product were determined. It was found that the dissolution rate profile of a developed formulation was similar to the commercial brand. Their similarity and difference factors were well within limits. In the bioequivalence study, the AUC_last and AUC_inf between the test and the reference products were not statistically different (p = 0.227 and p = 0.468, respectively), with the 90% CIs of 83.4–102.6% and 87.7–139.4%, respectively. However, the C _max of the two formulations was significantly different (p = 0.019). The 90% CI of the developed formulation was 72.0–93.0% compared to the commercial product. In vitro dissolution of locally prepared morphine sulfate sustained-release tablets was comparable to commercial brand. However, the results justified the conclusion of lack of bioequivalence of the developed product to the commercial one.     

Tetrachloroethene reductive dehalogenase of Dehalospirillum multivorans: substrate specificity of the native enzyme and its corrinoid cofactor

The substrate specificity of the tetrachloroethene reductive dehalogenase of Dehalospirillum multivoransand its corrinoid cofactor were studied. Besides reduced methyl viologen, titanium(III) citrate could serve as electron donor for reductive dehalogenation of tetrachloroethene (PCE) and trichloroethene to cis-1,2-dichloroethene. In addition to chlorinated ethenes, chlorinated propenes were reductively dechlorinated solely by the native enzyme. trans-1,3-Dichloropropene, 1,1,3-trichloropropene and 2,3-dichloropropene were reduced to a mixture of mono-chloropropenes, 1,1-dichloropropene, and 2-chloropropene, respectively. Other halogenated compounds that were rapidly reduced by the enzyme were also dehalogenated abiotically by the heat-inactivated enzyme and by commercially available cyanocobalamin. The rate of this abiotic reaction was dependent on the number and type of halogen substituents and on the type of catalyst. The corrinoid cofactor purified from the tetrachloroethene dehalogenase of D. multivorans exhibited an activity about 50-fold higher than that of cyanocobalamin (vitamin B(12)) with trichloroacetate as electron acceptor, indicating that the corrinoid cofactor of the PCE dehalogenase is not cyanocobalamin. Corrinoids catalyzed the rapid dehalogenation of trichloroacetic acid. The rate was proportional to the amount of, e.g. cyanocobalamin; therefore, the reductive dehalogenation assay can be used for the sensitive and rapid quantification of this cofactor.