Authentic Specimens Research Articles

Analysis of bile acid glutathione thioesters by liquid chromatography/electrospray ionization–tandem mass spectrometry

The formation of thioester-linked glutathione (GSH) conjugates of bile acids (BAs) is presumed to occur via trans-acylation reactions between GSH and reactive acyl-linked metabolites of BAs. The present study examines the chemical reactivity of cholyl-adenylate and cholyl-CoA thioester, acyl-linked metabolites of cholic acid (CA), with GSH to form CA-GSH conjugate in vitro. The authentic specimen of CA-GSH was synthesized along with GSH conjugates of four common BAs found in the human body. Their structures were confirmed by proton-nuclear magnetic resonance spectroscopy and electrospray ionization (ESI)–tandem mass spectrometry in positive- and negative-ion modes. Incubation of cholyl-adenylate or cholyl-CoA thioester with GSH was carried out at pH 7.5 and 37 °C for 30 min, with analysis of the reaction mixture by liquid chromatography/ESI–tandem mass spectrometry, where CA-GSH was detected on the product ion mass chromatograms monitored with stable and abundant dehydrated positive-ion [ M + H H 2O] + at m/ z 680.3 and fragmented negative-ion [GSH H] − at m/ z 306.0, and was definitely identified by CID spectra by comparison with those of the authentic sample. The results show that both cholyl-adenylate and cholyl-CoA thioester are able to acylate GSH in vitro.

Determination of therapeutics with growth‐hormone secretagogue activity in human urine for doping control purposes

The administration of growth-promoting agents such as human growth hormone as well as compounds with respective secretagogue activity is prohibited in sports according to the regulations of the World Anti-Doping Agency. Acetylcholine esterase inhibitors have been demonstrated to stimulate growth-hormone secretion in elderly humans, and new orally active drugs have been developed to provide alternatives to therapeutic injections of growth-hormone preparations. Preventive anti-doping strategies include method development for emerging drugs and potentially misused compounds. Hence, the mass spectrometric dissociation behavior of three acetylcholine esterase inhibitors (donepezil, galantamine and rivastigmine) and a structural analogue to the growth-hormone secretagogue SM-130686 were studied using high-resolution/high-accuracy orbitrap mass spectrometry. These data provided substantial information for screening procedures, complementing common methods of sports drug testing. Using liquid-liquid extraction and subsequent liquid chromatography/tandem mass spectrometry (LC/MS/MS) analysis, the four target analytes were determined at urinary concentrations of 15-20 ng/mL, recoveries ranged from 55-97%, and assay precisions were calculated at 5.2-15.8% (intraday) and 10.2-21.6% (interday) for all compounds. The applicability of the developed assay to authentic urine specimens was tested using two administration study urine samples after application of Reminyl (galantamine) and Aricept (donepezil). In both cases, the administered drug and the respective desmethylated metabolites were detected.

Determination of Propoxyphene in Oral Fluid

The determination of propoxyphene in oral fluid using solid-phase extraction and gas chromatography-mass spectrometry is described for the first time. The method employs collection of oral fluid with the Quantisal device, immunoassay screening of the specimen, confirmation of the positive screened samples after extraction using cation exchange/hydrophobic solid-phase extraction columns, optimized derivative formation, and gas chromatography-mass spectrometry in electron impact mode. Validated parameters including selectivity, linearity, accuracy, intra- and interday precision, extraction efficiency, and limit of quantitation were all within acceptable limits. The method was applied to authentic specimens taken from an individual prescribed propoxyphene following surgery.

LC/ESI-tandem mass spectrometric determination of bile acid 3-sulfates in human urine: 3β-Sulfooxy-12α-hydroxy-5β-cholanoic acid is an abundant nonamidated sulfate

We developed a highly sensitive and quantitative method to detect bile acid 3-sulfates in human urine employing liquid chromatography/electrospray ionization-tandem mass spectrometry. This method allows simultaneous analysis of bile acid 3-sulfates, including nonamidated, glycine-, and taurine-conjugated bile acids, cholic acid (CA), chenodeoxycholic acid (CDCA), deoxycholic acid (DCA), ursodeoxycholic acid (UDCA), and lithocholic acid (LCA), using selected reaction monitoring (SRM) analysis. The method was applied to analyze bile acid 3-sulfates in human urine from healthy volunteers. The results indicated an unknown compound with the nonamidated common bile acid 3-sulfates on the chromatogram obtained by the selected reaction monitoring analysis. By comparison of the retention behavior and MS/MS spectrum of the unknown peak with the authentic specimen, the unknown compound was identified as 3β,12α-dihydroxy-5β-cholanoic acid 3-sulfate.

2,7-Dimethyl-3,8-dinitrodipyrazolo[1,5-a:1′,5′-d]pyrazine-4,9-dione: A new labelling reagent for liquid chromatographic analysis of amino acids

The use of 2,7-dimethyl-3,8-dinitrodipyrazolo[1,5-a:1′,5′-d]pyrazine-4,9-dione as pre-column reagent for LC analysis of amino acids is proposed. The compound reacts (30 min at 68 °C in presence of 0.04 M triethylamine in a dimethylsulfoxide–water mixture) with primary and secondary amino function and the stable resulting adducts can be chromatographed under reversed-phase conditions and detected at λ = 280 nm. The derivatization conditions were optimized by a series of experiments. The effect of temperature, triethylamine concentration and reagent on the reaction was investigated. The yield of the glycine derivative was found to be quantitative at a reagent amino acid molar ratio of about 6 by comparison with an authentic specimen of synthesized glycine adduct. Application of the method to quality control of commercially available oral polyaminoacid formulations is described.

Stable Isotope Analyses of Heroin Seized from the Merchant Vessel Pong Su

A new type of heroin HCl seized in Australia was examined by stable isotope analysis. The final origin/process classification of these samples by chromatographic signature profiles of the impurity/manufacturing by-products was previously determined to be "unknown" by two independent national laboratories. Various drug enforcement authorities speculated that the heroin might be from a new region or new illicit process due to the unusual chromatographic impurity profiles that were present. Samples from 20 different kilogram packages were examined for isotopic content to determine if the samples fit isotopic patterns of known origins or if they were unique to any known origins. Authentic specimens from Southeast Asian (N=59), Southwest Asia (N=37), South America (N=104), and Mexico (N=21) we concomitantly examined for comparison purposes. Both continuous flow elemental analysis-isotope ratio mass spectrometry and gas chromatography-isotope ratio mass spectrometry techniques were utilized. Heroin samples were also converted to morphine, without apparent isotopic fractionation, utilizing methanolic HCl for gas chromatography-isotope ratio mass spectrometry. The Pong Su samples were found to be isotopically and isotopically/alkaloidally distinct from the known origin/process classifications of Southwest Asian, Southeast Asian, South American, and Mexican.

Evaluation of Buprenorphine CEDIA Assay versus GC-MS and ELISA using Urine Samples from Patients in Substitution Treatment

As buprenorphine becomes more clinically used in heroin substitution treatment, there is an increasing need for methods suitable for high-volume screening. In this study, a new immunochemical test based on CEDIA technology was evaluated for the use in clinical urine drug testing. The method was compared with an existing ELISA method and a gas chromatography-mass spectrometry (GC-MS) method on urine specimens from patients in heroin substitution treatment. The precision of the CEDIA assay was < 9% both within- and between-day at levels at and above the cutoff limit of 5 microg/L. The concordance in qualitative results with an existing ELISA method was 96.8%. The CEDIA measuring range was extended by diluting urine samples 100-fold with saline, and the results agreed well (slope of regression line was 1.09, r(2) = 0.968) with GC-MS. The sensitivity of CEDIA in detecting authentic specimen containing buprenorphine at levels >or= 5 microg/L was 99.5%. Cross-reactivity causing false-positive response was discovered in patients receiving prescribed dihydrocodeine. The urine concentration of total buprenorphine in urine from patients prescribed daily doses between 0.2 and 24 mg ranged from 0.5 to 2900 microg/L. The concentration of the metabolite norbuprenorphine was usually higher, and the median ratio of buprenorphine to norbuprenorphine was 0.23 (95% were below 1). We conclude that the CEDIA assay is suitable for application in high-volume screening of buprenorphine for urine drug testing.

Type studies of the polypores described by E.J.H. Corner from Asia and West Pacific Areas. VII. Species described in Trametes (1)

Identifications of 30 species of polypores described by Corner were made by type examinations. The following new combinations are proposed: Antrodiella depauperata, Daedalea dochmia, D. fulvirubida, D. lusor, D. pseudodochmia, Fomitella fumosipora, F. rhodophaea, Microporus internuntius, and Trichaptum brastagii. The following names are accepted in the original genus: Trametes arcana, T. barbulata, T. daedaleoides, and T. jejuna. The following species are taxonomic synonyms of alreadydescribed species: Trametes atriceps, a synonym of Trametes menziesii; T. badiuscula, a synonym of Abundisporus roseoalbus; T. biogilvoides, a synonym of Coriolopsis glabro-rigens; T. elevata, a synonym of Daedalea dochmia; T. flammula, a synonym of C. glabro-rigens; T. fulvidochmia, a synonym of D. dochmia; T. linguiformis, a synonym of Trichaptum suberosum; and T. luridochracea, a synonym of Lenzites acutus. Trametes benetosta is a synonym of Fomitella rhodophaea or its allied species. The following species are dubious because of the poor or sterile conditions of their type specimens: Trametes allantospora, T. cristobalensis, T. farinolens, T. febris, T. fuligineicana, and T. granulifera. No authentic specimens were traced for T. argenteiceps, T. benevestita, T. castaneifumosa, T. decorticans, and T. flavidinigra. Descriptions and line drawings are given for poorly known species.

Rapid Simultaneous Determination of Amphetamine, Methamphetamine, 3,4-Methylenedioxyamphetamine, 3,4-Methylenedioxymethamphetamine, and 3,4-Methylenedioxyethylamphetamine in Urine by Fast Gas chromatography-mass Spectrometry

The use of fast gas chromatography-mass spectrometry (FGC-MS) was investigated to improve the efficiency of analysis of urine specimens that previously screened presumptively positive for amphetamine (AMP), methamphetamine (MAMP), 3,4-methylenedioxyamphetamine (MDA), 3,4-methylenedioxymethamphetamine (MDMA), and/or 3,4 methylenedioxyethylamphetamine (MDEA) by immunoassay testing. Specimens were pretreated with basic sodium periodate, extracted using a positive-pressure manifold/cation-exchange solid-phase cartridge methodology, and derivatized using 4-carbethoxyhexafluorobutyryl chloride (4-CB). The analytical method was compared to traditional GC-MS analysis and evaluated with respect to assay chromatography, linearity, sensitivity, precision, accuracy, and reproducibility. The limits of detection were 62.5 ng/mL for MDA and 31.25 ng/mL for AMP, MAMP, MDMA, and MDEA. All of the target analytes were linear to 12,000 ng/mL with the exception of MAMP which was linear to 10,000 ng/mL. The intra-assay precision of a 500 ng/mL multiconstituent control (n=15) ranged from 522.6 to 575.9 ng/mL with a coefficient of variation of less than 3.8%. Authentic human urine specimens (n=187) previously determined to contain the target analytes were re-extracted and analyzed by both FGC-MS and the currently utilized GC-MS method. No significant differences in specimen concentration were observed between these analytical methods. No interferences were seen when the performance of the FGC-MS method was challenged with ephedrine, pseudoephedrine, phenylpropanolamine, and phentermine. When compared to traditional GC-MS analysis, FGC-MS analysis provided a dramatic reduction in retention time for amphetamine (1.8 min vs. 4.12 min). For example, the FGC-MS method reduced overall run time for a batch of 56 specimens from 12.0 h to 7.25 h. This reduction in analysis time makes FGC-MS an attractive alternative to traditional GC-MS by allowing a laboratory greater flexibility in the purchase and use of capital equipment and in the assignment of laboratory personnel, all resulting in greater overall efficiency by decreasing reporting times for AMP, MAMP, and designer amphetamine positive specimens.

The genus Pyrenomyxa and its affinities to other cleistocarpous Hypoxyloideae as inferred from morphological and chemical traits

Types and authentic specimens of Hypoxylon piceum, Pulveria porrecta, and Pyrenomyxa invocans were studied for morphological traits and extrolite (= secondary metabolite) profiles generated by analytical HPLC with UV-visual and mass spectrometric detection. The orange stromatal pigments of P. invocans are rubiginosin A and mitorubrinol. It lacks three different types of extrolites (BNT, macrocarpone and hypomiltin) that are known from Hypoxylon taxa and occur in H. piceum and P. porrecta. In agreement with morphological traits, the latter two names are regarded as synonymous and transferred to Pyrenomyxa. Another species from Eastern Russia, Pyrenomyxa morganii sp. nov., is recognized. It contains yet unidentified azaphilones besides BNT and orsellinic acid, and its culture produces 5-methylmellein and a virgariella-like anamorph. These findings suggest a close relationship of Pyrenomyxa to Hypoxylon and emphasize the utility of chemotaxonomic traits for fungal taxonomy in general. Pyrenomyxa is accepted ad interim until the phylogenetic relationships among Hypoxylon have been further evaluated by means of chemotaxonomic, morphological and molecular methods.

The genus Pyrenomyxa and its affinities to other cleistocarpous Hypoxyloideae as inferred from morphological and chemical traits

Types and authentic specimens of Hypoxylon piceum, Pulveria porrecta, and Pyrenomyxa invocans were studied for morphological traits and extrolite (= secondary metabolite) profiles generated by analytical HPLC with UV-visual and mass spectrometric detection. The orange stromatal pigments of P. invocans are rubiginosin A and mitorubrinol. It lacks three different types of extrolites (BNT, macrocarpone and hypomiltin) that are known from Hypoxylon taxa and occur in H. piceum and P. porrecta. In agreement with morphological traits, the latter two names are regarded as synonymous and transferred to Pyrenomyxa. Another species from Eastern Russia, Pyrenomyxa morganii sp. nov., is recognized. It contains yet unidentified azaphilones besides BNT and orsellinic acid, and its culture produces 5-methylmellein and a virgariella-like anamorph. These findings suggest a close relationship of Pyrenomyxa to Hypoxylon and emphasize the utility of chemotaxonomic traits for fungal taxonomy in general. Pyrenomyxa is accepted ad interim until the phylogenetic relationships among Hypoxylon have been further evaluated by means of chemotaxonomic, morphological and molecular methods.

Ethyl Sulfate: A Metabolite of Ethanol in Humans and a Potential Biomarker of Acute Alcohol Intake

This study identified ethyl sulfate (EtS) in human urine and compared the excretion characteristics of EtS with that of ethanol and ethyl glucuronide (EtG). Urine samples were collected from healthy subjects after a single ethanol dose, and also selected from routine clinical samples. Simultaneous analysis of EtS and EtG was performed by direct electrospray liquid chromatography-mass spectrometry in the negative ion mode, with selected-ion monitoring of the pseudomolecular ions at m/z 125 for EtS (M(w) 126 g/mol) and m/z 221 for EtG (M(w) 222 g/mol). The identity of EtS in authentic urine specimens was established by co-chromatography with reference substance, the presence of product ions (m/z 97 and 80 from m/z 125) with correct relative intensity, and a correct sulfur isotope ratio for (34)S (m/z 127). After healthy subjects drank ethanol, EtS showed a much longer, dose-dependent elimination half-life than the parent compound. No EtS was detected in urines collected after abstention from ethanol for several days prior to sampling. Among 354 consecutive clinical samples, 86 were positive for both EtS and EtG with a mean EtG/EtS molar ratio of 2.3 (median 1.7). Another three urine samples were only positive for EtS and four only for EtG. The present results confirm that sulfate conjugation is a normal but minor metabolic pathway for ethanol in humans, and EtS a common constituent in the urine after alcohol intake. It is also indicated that the concurrent determination of EtS and EtG will improve sensitivity, when being used as biomarkers of recent drinking.

An Automated Alcohol Dehydrogenase Method for Ethanol Quantification in Urine and Whole Blood

In a previous work, an automated alcohol dehydrogenase method for the quantification of ethanol in whole blood (blood) specimens was presented. In the present work the application of the method to urine specimens has been investigated. Also, method robustness to routine analysis of urine and blood specimens during a period of eight months is shown. The limits of detection and quantification for urine were 0.0012 g/dL and 0.0042 g/dL, respectively. Relative standard deviations for the repeatability and within-laboratory reproducibility were in the ranges 1.4-4.1% and 1.8-4.6%, respectively. The method was compared with two headspace gas chromatography-flame ionization detection methods using authentic forensic urine specimens (n = 305) and blood specimens (n = 3186). Passing-Bablok regression for the concentration range 0.01-0.48 g/dL (urine) and 0.002-0.40 g/dL (blood) showed a statistically significant difference, for urine y = 0.9313 (0.9250 - 0.9377)x + 0.0038 (0.0029-0.0044) and for blood y = 0.9493 (0.9491 - 0.9495)x + 0.0032 (0.00318-0.00323), at 95% confidence level. The results of the external quality control specimens were in accordance with the reported theoretical concentrations.

PANDURALIMULUS BABCOCKI N. GEN. AND SP., A NEW LIMULACEAN HORSESHOE CRAB FROM THE PERMIAN OF TEXAS

Fossil horseshoe crabs are rare, and new discoveries have the potential for providing important taxonomic and biostratigraphic information. The superfamily Limulacea Zittel, 1885, embraces the lineal ancestors of the modern genera, including Limulus Muller, 1785, within the Limulidae Zittel, 1885. The geologically oldest of the limulaceans are arrayed within the late Paleozoic Paleolimulidae Raymond, 1944. The first discovery, as well as all authentic specimens, of fossil paleolimulids have been from the Late Paleozoic rocks of Kansas and Illinois. This paper not only describes a new genus and species of horseshoe crab but also records the first occurrence of the family in north-central Texas, and clarifies placement of taxa within the Paleolimulidae. Specimens were collected in Baylor County aproximately 15 km northeast of Seymour, Texas, from a layer of clastic sediment within the Lower Permian (Leonardian) Maybelle Limestone, the member comprising the lower part of the Lueders Formation (National Museum of Natural History site USNM 41884) (Fig. 1). The Lueders Formation ranges from 50 to 275 ft (15–82 m) thick and is comprised of alternating shale and limestone (Stafford, 1960). The limestones of the Maybelle Member are thick, massive, fossiliferous, and resistant (Beede and Waite, 1918). The paleoenvironment in which these rocks were deposited was most likely deltaic; gymnosperms, pteridosperms, horseshoe crabs, bivalves, and other fossil fragments occur together in a fine-grained, blue-gray matrix suggesting a marginal marine setting. Babcock and Merriam (2000) suggested that xiphosurans might be preserved in tidally influenced coastal settings like this because high depositional rates would favor preservation of the delicate cuticle. Figure 1 —Generalized stratigraphic section showing the position from which the xiphosurans were collected ### Subclass XiphosuraLatreille, 1802 Order Xiphosurida Latreille, 1802 Suborder Limulina Richter and Richter, 1929 Superfamily Limulacea Zittel, 1885 Family Paleolimulidae Raymond, 1944 #### Diagnosis Ophthalmic ridges meeting anterior to cardiac lobe; …

Identification of a novel conjugate in human urine: bile acid acyl galactosides

We report a novel conjugate, bile acid acyl galactosides, which exist in the urine of healthy volunteers. To identify the two unknown peaks obtained in urine specimens from healthy subjects, the specimens were subjected to solid phase extraction and then to liquid chromatographic separation. The eluate corresponding to the unknown peaks on the chromatogram was collected. Following alkaline hydrolysis and liquid chromatography (LC)/electrospray ionization (ESI)-mass spectrometric (MS) analysis, cholic acid (CA) and deoxycholic acid (DCA) were identified as liberated bile acids. When a portion of the alkaline hydrolyzate was subjected to a derivatization reaction with 1-phenyl-3-methyl-5-pyrazolone, a derivative of galactose was detected by LC/ESI-MS. Finally, the liquid chromatographic and mass spectrometric properties of these unknown compounds in urine specimens were compared to those of authentic specimens and the structures were confirmed as CA 24-galactoside and DCA 24-galactoside. These results strongly imply that bile acid 24-galactosides, a novel conjugate, were synthesized in the human body.

An analysis of flavonoid compounds in leaves of Japonolirion (Petrosaviaceae)

Japonolirion, comprising Japonolirion osense Nakai, which occurs on serpentinite at two widely separated localities in Japan, has been considered as an isolated taxon, but more recently has been proved by molecular evidence to be a sister group to an achlorophyllous, mycoheterotrophic genus, Petrosavia. In an effort to research possible characters linking these groups, we analyzed the flavonoid compounds obtained from leaves of Japonolirion using UV spectra, mass spectrometry and 1H and 13C nuclear magnetic resonance, and acid hydrolysis of the original glycosides as well as direct thin layer chromatography and high performance liquid chromatography comparisons with authentic specimens. As a result, we identified seven flavonoids, of which two were major components identified as 6-C-glucosylquercetin 3-O-glucoside and isoorientin. The remaining five were minor components identified as 6-C-glucosylkaempferol 3-O-glucoside, quercetin 3-O-glucoside, quercetin 3-O-arabinoside, vicenin-2 and orientin. Both 6-C-glucosylquercetin 3-O-glucoside and 6-C-glucosylkaempferol 3-O-glucoside were recorded for the first time in nature. Because of their restricted occurrence in angiosperms, both C-glycosylflavonols and 3-O-glycosides of C-glycosylflavonols may be significant chemical markers for assessing relationships of J. osense.

Fast Quantification of Ethanol in Whole Blood Specimens by the Enzymatic Alcohol Dehydrogenase Method. Optimization by Experimental Design

A sensitive, fast, simple, and high-throughput enzymatic method for the quantification of ethanol in whole blood (blood) on Hitachi 917 is presented. Alcohol dehydrogenase (ADH) oxidizes ethanol to acetaldehyde using the coenzyme nicotinamide adenine dinucleotide (NAD), which is concurrently reduced to form NADH. Method development was performed with the aid of factorial design, varying pH, and concentrations of NAD+ and ADH. The linear range increased and reaction end point decreased with increasing NAD+ concentration and pH. The method was linear in the concentration range 0.0024-0.4220 g/dL. The limits of detection and quantification were 0.0007 g/dL and 0.0024 g/dL, respectively. Relative standard deviations for the repeatability and within-laboratory reproducibility were in the ranges 0.7-5.7% and 1.6-8.9%, respectively. The correlation coefficient when compared with headspace gas chromatography-flame ionization detection methods was 0.9903. Analysis of authentic positive blood specimens gave results that were slightly lower than those of the reference method.

Chemical study of Eriobotrya japonica

In continuation of the search for sources of biologically active compounds, we studied flowers of E. japonica L. (Japanese mimosa, Rosaceae), which is cultivated in Azerbaidzhan as a decorative. In contrast with many plants, it flowers in November—December [1, 2]. Dried flowers (0.5 kg) were collected in December 2002 and extracted successively with hexane, chloroform, and ethanol. Compound 1 was obtained from the hexane extract; 2, from chloroform. The ethanol extract was evaporated to dryness and dissolved in water. The aqueous solution was treated with ethylacetate. Solvent was removed. Fraction crystallization from ethanol produced 3 and 4. Compound 1 was small needle-like white crystals that were soluble in hexane, chloroform, and ethanol and insoluble in water, mp 138-139°C (ethanol), [α]D -37° (c 0.25, CHCl3); acetate, mp 126-128°C (ethanol). It appeared on TLC [developer phosphotungstic acid solution (25%) at 100-105°C] as a pink spot and gave a positive reaction with Burchard—Liebermann and Salkovskii reagents. Compound 2 was needle-like white crystals that were soluble in chloroform and ethanol and insoluble in hexane and water. It gave a positive reaction with Burchard—Liebermann and Salkovskii reagents. It appeared on chromatograms as a pink spot, mp 280-282(C (ethanol), [α]D -64° (c 0.94, CHCl3). Compound 3 was white crystals, mp 232-234°C (ethanol), [α]D -59° (c 0.6, CH3OH), Rf 0.73 (n-C4H9OH:CH3CO2H:H2O, 4:1:5), soluble in water and ethanol. Hydrolysis by H2SO4 (4%, 3h) cleaved it into quercetin and L-arabinose. Yield of quercetin, 63.5%. Compound 4 was white crystals, mp 219-221°C (ethanol), [α]D -38° (c 0.15, CH3OH), Rf 0.69 (n-C4H9OH:CH3CO2H:H2O, 4:1:5), soluble in water and ethanol. Acid hydrolysis by H2SO4 (5%) quickly cleaved it into quercetin and D-glucose. Yield of aglycon, 64.3%. Based on the physicochemical properties and chromatographic behavior (comparison with authentic specimens), 1 was identified as β-sitosterol; 2, ursolic acid; 3, quercetin-3-O-α-L-arabinoside; 4, quercetin-3-O-β-D-glucoside [3, 4]. These compounds are widely distributed in various plants. E. japonica L. from Azerbaidzhan flora was studied for the first time [5].

DISCOVERY OF THE TYPE SPECIMEN OF PINNULARIA GASTRUM EHRENBERG, THE TYPE SPECIES OF THE GENUS PLACONEIS MERESCHKOWSKY

A specimen was discovered in the Ehrenberg Collection at BHUPM that can serve as lectotype of Pinnularia gastrum, the type species of Placoneis Mereschkowsky. Previously, Cox was unable to find an authentic specimen in the Ehrenberg Collection and therefore proposed to conserve the binomial with a Donkin specimen at the BM. The usefulness of conserving names to settle uncertainties of typification is discussed. In addition, Pinnularia placentula, a species morphologically similar to P. gastrum, is lectotypified with Ehrenberg's original drawing. Since the type specimen is lost, Cox's proposed neotype can serve as epitype.

Anaerobic/aerobic sequential treatment of a cotton textile mill wastewater

AbstractThe treatment of a wastewater taken from a cotton textile mill was investigated using an anaerobic/aerobic sequential system during an operational period of 87 days. The process units consisted of an upflow anaerobic sludge blanket (UASB) reactor and a continuous stirred tank reactor (CSTR). Wastewater characterization was performed before feeding the reactor system. Glucose‐COD, and azo dyes were added to the textile wastewater for comparative purposes in the final period of operation. The pH values in the effluent of the UASB reactor were suitable for optimal anaerobic treatment in all runs. The biodegradable part of the COD in wastewater was removed effectively, with the anaerobic stage improving the biodegradability of wastewater entering the aerobic stage. The UASB reactor permitted COD and color removals of 9–51% and 46–55%, respectively, at a hydraulic retention time (HRT) of 30 h. COD removal efficiencies were between 40 and 85% and color removal efficiencies were 39–81% in normal and artificially‐colored wastewaters at a total HRT of 5.75 days in the UASB/CSTR reactor system. Benzidine produced from the cleavage of azo bond in the anaerobic stage was effectively removed in the aerobic stage, and was identified by comparison of its HPLC spectrum with that of an authentic specimen. Copyright © 2004 Society of Chemical Industry