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Biomedical subjects

R G Berger

Publications and source records attributed to R G Berger.

At least 19 recordsLinked to original sources
PubMed2004

Degradation of alpha-pinene oxide and [2H7]-2,5,6-trimethyl-hept-(2E)-enoic acid by Pseudomonas fluorescens NCIMB 11761.

When submerged cultured Pseudomonas fluorescens NCIMB 11761 was fed-batch supplemented with alpha-pinene oxide, a rapid formation of 2,6-dimethyl-5-methylene-hept-(2Z)-enal (I) (isonovalal) was observed. Biotransformation and isomerisation of (I) to the (2E)-isomer (II) (novalal) were enhanced by Lewatit OC 1064, a macroporous polystyrene adsorbent. Accelerated isomerisation in the presence of an amino donor (glycine) at pH 7.3 pointed to a merely chemical mechanism. A maximum yield of 48 g of aldehydesl(-1) was achieved, but quantitative analysis of the volatile fraction showed that the molar conversion of the pinene oxide substrate reached no more than 67%. To fill this gap of the mass balance, the acidic fraction was isolated. It contained several compounds which suggested a beta-oxidation-like catabolism starting from 2,6-dimethyl-5-methylene-hept-(2E)-enoic acid (III) (novalic acid). Using [2H7]-2,5,6-dimethyl-hept-(2E)-enoic acid as a conversion substrate and gas chromatography coupled to atomic emission detection and mass spectrometry a degradation pathway via labelled 3,4-dimethylpentenoic and methylpropanoic acids was evidenced. This pathway may play a predominant role in isoprenoid degradation by soil bacteria.

Aldehydes↗
PubMed2003

Cleavage of beta,beta-carotene to flavor compounds by fungi.

More than 50 filamentous fungi and yeasts, known for de novo synthesis or biotransformation of mono-, sesqui-, tri-, or tetraterpenes, were screened for their ability to cleave beta,beta-carotene to flavor compounds. Ten strains discolored a beta,beta-carotene-containing growth agar, indicating efficient degradation of beta,beta-carotene. Dihydroactinidiolide was formed as the sole conversion product of beta,beta-carotene in submerged cultures of Ganoderma applanatum, Hypomyces odoratus, Kuehneromyces mutabilis, and Trametes suaveolens. When mycelium-free culture supernatants from five species were applied for the conversions, nearly complete degradation of beta,beta-carotene was observed after 12 h. Carotenoid-derived volatile products were detected in the media of Ischnoderma benzoinum, Marasmius scorodonius, and Trametes versicolor. beta-Ionone proved to be the main metabolite in each case, whereas beta-cyclocitral, dihydroactinidiolide, and 2-hydroxy-2,6,6-trimethylcyclohexanone were formed in minor quantities. Using a photometric bleaching test, the beta,beta-carotene cleaving enzyme activities of M. scorodonius were partially characterized.

Agar↗
PubMed2003

A labeling study to elucidate the biosynthesis of 4-(4-hydroxyphenyl)-butan-2-one (raspberry ketone) by Nidula niveo-tomentosa.

Submerged cells of the basidiomycete Nidula niveo-tomentosa, a microbial producer of 4-(4-hydroxyphenyl)-butan-2-one, were supplemented with (13)C-labeled L-phenylalanines and with [1-(13)C]glucose. Labeled transformation products were detected by a novel method of analyzing stable isotope-labeled metabolites, gas chromatography (GC) coupled to an atomic emission detector, and by GC-mass spectrometry. A benzoate moiety was side chain elongated according to the poly-beta-keto scheme. The presence of an acetyl coenzyme A-carboxylase inhibitor shifted the spectrum of products to benzyl compounds. Hence, the fungal pathway differs from the one established for plant tissues.

Basidiomycota↗
PubMed2001

Phenyl propenoic side chain degradation of ferulic acid by Pycnoporus cinnabarinus - elucidation of metabolic pathways using [5-2H]-ferulic acid.

The white-rot fungus Pycnoporous cinnabarinus (DMS-1184) was submerged cultured for 22 days under controlled conditions in a bioreactor. After 6, 9, and 15 days of culture the growth medium was supplemented with [5-2H]-labelled ferulic acid (I). The major phenolic compounds identified labelled were four lignans, the methyl esters of ferulic (I) and vanillic acid (VIII), (E)-coniferyl aldehyde (II), (E)-coniferyl alcohol (III), vanillic acid (VIII), vanillin (IX) and vanillyl alcohol (X). The detection of considerable amounts of labelled 4-hydroxy-3-methoxyacetophenone (VII) in the late growth phase suggested the increasing formation and decarboxylation of free 4-hydroxy-3-methoxybenzoylacetic acid (VI) and, thus, a beta-oxidation-like degradation of ferulic acid (I) or its methyl ester to vanillic acid (VIII). 4-Hydroxy-3-methoxybenzoylacetic acid methyl ester (VI) and 3-hydroxy-(4-hydroxy-3-methoxyphenyl)-propanoic acid methyl ester (V) were synthesised and then identified as metabolites in the culture medium. The fungal degradation of the phenyl propenoic side chain of ferulic acid (I), a principal key step of lignin decomposition, appeared to proceed analogous to fatty acids.

Basidiomycota↗
PubMed2001

Intelligent use of NSAIDs--where do we stand?

Non-steroidal anti-inflammatory drugs (NSAIDs) are drugs commonly prescribed for a variety of medical conditions. They are potent pharmacological agents efficacious for inflammatory conditions, but have significant gastrointestinal (GI), renal and haematological toxicity that must not be taken lightly. The newer, more cyclooxygenase-(COX)-2-selective NSAIDs, have no effects on platelet function and little GI toxicity, but do have renal physiological effects. The superiority of one NSAID over another has not been clinically demonstrated in musculoskeletal conditions, nor has the efficacy of NSAIDs in non-inflammatory rheumatic conditions been shown to be better than that of simple analgesics. NSAIDs are indicated for primary therapy of inflammatory rheumatic diseases and the more selective COX-2 agents should be employed as first choice when economically feasible. NSAIDs should not be used indiscriminately for non-inflammatory osteoarthritis or musculoskeletal injuries, particularly in the elderly patient, in whom alternative, less toxic therapy should be sought.

Aged↗
PubMed2000

A comparative study on the disintegration of filamentous fungi.

Different methods for cell disintegration were tested for their efficacy on filamentous fungi, including percussion grinding, homogenization using an Ultra-Turrax, chemical treatment and lyophylization. The release of protein from Ganoderma applanatum and Pycnoporus cinnabarinus and the activity of cytoplasmatic glucose-6-phosphate dehydrogenase in the crude extracts were monitored to determine the efficiency of each disintegration technique used. Fungal cells proved to be particularly resistant towards some disintegration methods commonly used for yeasts and bacteria. Best results were obtained using a percussion grinder, if necessary, in combination with an Ultra-Turrax pretreatment.

Basidiomycota↗
PubMed2000

Generation of odorous acyloins by yeast pyruvate decarboxylases and their occurrence in sherry and soy sauce.

Volatile acyloins (alpha-hydroxy ketones) were obtained by condensing either aldehydes with pyruvate or 2-keto acids with acetaldehyde in a reaction catalyzed by yeast pyruvate decarboxylases (EC 4.1.1.1). Odor qualities and threshold values of 34 acyloins were evaluated, and 23 of them possessed distinct flavor properties. Sherry and soy sauce flavors were analyzed: 2-hydroxy-3-pentanone and 3-hydroxy-2-pentanone were identified in soy sauce for the first time; these and 2-hydroxy-5-methyl-3-hexanone and 3-hydroxy-1-phenyl-2-butanone were isolated from sherry for the first time. The biocatalytic efficiencies of crude pyruvate decarboxylase preparations from Zygosaccharomyces bisporus, Saccharomyces cerevisiae, Kluyveromyces lactis, and Kluyveromyces marxianus were compared. Product yields comparable to those of conversions with purified pyruvate decarboxylase demonstrated the suitability of crude enzyme extracts as cost-effective biocatalysts in acyloin formation. Conversion rates of >50% showed that the potential of this type of enzyme to catalyze the formation of aliphatic acyloins has been underestimated before.

Alcoholic Beverages↗
PubMed2000

Purification, characterisation and cDNA sequencing of pyruvate decarboxylase from Zygosaccharomyces bisporus.

Cells of the wild-type yeast strain Zygosaccharomyces bisporus CBS 702 form alpha-hydroxy ketones from aromatic amino acid precursors during fermentation. Pyruvate decarboxylase (PDC, E.C. 4.1.1.1), the key enzyme of this biotransformation catalysing the non-oxidative decarboxylation of pyruvate and other 2-oxo-acids, was purified and characterised. The active enzyme is homotetrameric (alpha4) with a molecular mass of about 244 kDa. Activation of PDC by its substrate pyruvate results in a sigmoidal dependence of the reaction rate from substrate concentration (apparent Km value 1.73 mM; Hill coefficient 2.10). A cDNA library was screened using a PCR-based procedure, and a 1856 bp cDNA of PDC was identified and sequenced. The cDNA encodes a polypeptide of 563 amino acid residues (monomeric unit). Sequence alignments demonstrate high homologies (> 80%) to PDC genes from Saccharomyces cerevisiae, Kluyveromyces lactis and Kluyveromyces marxianus.

DNA, Complementary↗
PubMed1999

Effects of R-(+)-limonene on submerged cultures of the terpene transforming basidiomycete Pleurotus sapidus.

The biotransformation of limonene by the basidiomycete Pleurotus sapidus yielded cis/trans-carveol and carvone as the main products. The transformation period was extended from 4 days after direct addition to 12 days by gas phase addition of the substrate. After 2 days of transformation, 97% of the substrate had accumulated in the mycelium, while only 3% were present in the culture medium. Substrate toxicity led to a decrease of dry matter. Adaptation of the precultures with small amounts of substrate doubled the concentration of carveol and increased the concentration of carvone by a factor of 3-4. Total product concentrations of > 100 mg l-1 were reached.

Adaptation, Physiological↗
PubMed1999

Biotransformation of citronellol by the basidiomycete Cystoderma carcharias in an aerated-membrane bioreactor.

The basidiomycete Cystoderma carcharias transformed citronellol into 3,7-dimethyl-1,6,7-octanetriol as the main product. 3,7-Dimethyl-6,7-epoxy-1-octanol was identified as important intermediary product of the biotransformation, and the allylic diols 2,6-dimethyl-2-octene-1,8-diol, 3,7-dimethyl-5-octene-1,7-diol and 3,7-dimethyl-7-octene-1,6-diol were found to be minor products. Microbial formation of rose oxide, a flavour-impact component, was observed for the first time. The formation of the main products was inhibited by 70% after addition of 0.1 mmol l-1 cytochrome monooxygenase inhibitors. Formation of 3,7-dimethyl-1,6,7-octanetriol was effective in a bioreactor with aeration over a coil of a hydrophobic microporous polypropene capillary membrane. Production rates of up to 150 mg l-1 day-1 were reached and led to a product concentration of 866 mg l-1 (conversion rate: 52%). The total loss of the added volatile substrate via the exhaust air was 4.5% when this aeration method was used.

Acyclic Monoterpenes↗
PubMed1998

Biotechnological production of flavours and fragrances.

The biotechnological generation of natural aroma compounds is rapidly expanding. Aroma chemicals, such as vanillin, benzaldehyde (bitter almond, cherry) and 4-(R)-decanolide (fruity-fatty) are marketed on a scale of several thousand tons per year. Their possible production by single-step biotransformations, bioconversions and de novo synthesis using microorganisms, plant cells or isolated enzymes is shown. The perspectives of bioprocesses for the oxifunctionalisation of lower terpenes by genetically modified organisms and economic aspects are discussed.

Bacteria↗
PubMed1997

Analysis of oxylipins by high-performance liquid chromatography with evaporative light-scattering detection and particle beam-mass spectrometry.

The metabolism of 13S-hydroperoxy-9Z,11E,15Z-octadecatrienoic acid was investigated in a crude enzyme extract from mung bean seedlings (Phaseolus radiatus L.). Hydroperoxide-metabolizing activity was mainly due to a hydroperoxide lyase and, to a lesser extent, to an allene oxide synthase and a peroxygenase. Oxylipins originating from hydrolysis and cyclization of the allene oxide synthase product 12,13-epoxy-9Z,11,15Z-octadecatrienoic acid and from peroxygenase catalysis were identified by high-performance liquid chromatography (HPLC) particle beam-mass spectrometry (PB-MS) and quantified by normal-phase HPLC with an evaporative light-scattering detector (ELSD). An advantage of this methodology was the possibility to avoid extensive derivatization procedures commonly used for the gas chromatographic analysis of oxylipins. Owing to a comparable sample inlet system, the ELSD served an important analytical pilot function for the PB-MS: Qualitatively identical chromatographic patterns were obtained with both detection systems. The HPLC system enabled the separation of methyl 12-oxo-phytodienoate, methyl 11-hydroxy-12-oxo-9Z,15Z-octadecadienoate, methyl 12-oxo-13-hydroxy-9Z,15Z-octadecadienoate, methyl 9-hydroxy-12-oxo-10E,15Z-octadecadienoate, methyl 13-hydroxy-9Z,11E,15Z-octadecatrienoate, methyl 15,16-epoxy-13-hydroxy-9Z,11E-octadecadienoate, and methyl 13-hydroperoxy-9Z,11E,15Z-octadecatrienoate on a Lichrospher DIOL column within 33 min. Compared with a diode array detector, the ELSD proved to be more sensitive, in the case of methyl 12-oxo-13-hydroxy-9Z, 15Z-octadecadienoate by a factor of about 15. In addition, volatile metabolites were analyzed by capillary gas chromatography. The yield of the hydroperoxide lyase product 2E-hexenal was 49%, whereas the sum of oxylipins reached about 15%.

Aldehyde-Lyases↗
PubMed1996

Geosmin and Related Volatiles in Bioreactor-Cultured Streptomyces citreus CBS 109.60.

Streptomyces citreus CBS 109.60 produced geosmin and a complex pattern of other volatile compounds during cultivation in a 2.5-liter laboratory bioreactor. Volatiles were isolated from disrupted cells, from the culture medium, and from the waste air of the bioreactor by adsorption on Lewatit OC 1064MD. Quantitative and qualitative analyses were carried out using capillary gas chromatography and coupled gas chromatography-mass spectroscopy. S. citreus produced 56 volatile compounds, which were mainly terpenoids but also included aliphatic ketones, alcohols, esters, pyrazines, furan(on)es, and aromatic types during the growth phase. The major components were geosmin and a germacradienol. A biosynthetic pathway for geosmin including eudesmanolides is proposed.

Journal Article↗
PubMed1995

Separation and characterisation of bovine histone H1 subtypes by combined ion-exchange and reversed-phase chromatography and mass spectrometry.

In order to separate and identify histone H1 subtypes from calf thymus we used both electrospray mass spectrometry (ES-MS) and matrix-assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI-TOF-MS) after a three-step chromatographic procedure consisting of reversed-phase high-performance liquid chromatography (RP-HPLC), size-exclusion chromatography (SEC) and ion-exchange chromatography (IEC). Under the RP-HPLC conditions described, we obtained two baseline-separated H1-fractions which were characterised by MALDI-TOF-MS. The determined masses ranged from 22,850 to 22,590 for the first fraction and from 22,070 to 21,250 for the second fraction. Further, it was shown that the first fraction contained at least four and the second one at least five subtypes of the histone class H1. Four homogeneous pure H1 subtypes were obtained by a combination of IEC followed by SEC and RP-HPLC. The molecular masses of these four subtypes determined by ES-MS were 22,606, 22,761, 21,347 and 21,263. We obtained six additional molecular masses of histone H1 subtypes from three heterogeneous fractions, namely 22,066, 21,802, 20,586 and 19,817 by ES-MS and 22,800 and 22,675 by MALDI-TOF-MS. The retention times of these fractions and the molecular masses were in agreement with the data obtained from RP-HPLC fractions by MALDI-TOF-MS.

Animals↗
PubMed1995

Chemical phosphorylation of the peptides GGXA (X = S, T, Y): an evaluation of different chemical approaches.

An evaluation was made of the two methods most commonly used for phosphorylation of hydroxyamino acids in peptides, i.e. the tetrazole-catalysed phosphitylation by di-tert-butyl-N,N-diethylphosphoramidite followed by oxidation and the phosphorylation by dibenzylphosphochloridate. As model system the sequence GGXA (X = S, T, Y) was used which represents a random-coil sequence avoiding the influence on the reaction kinetics of secondary structure formation. In the case of serine- and threonine-containing peptides, both synthetic methods gave comparable yields of the desired phosphopeptides. The phosphorylation of tyrosine was achieved more favorably via the phosphoramidite method. However, phosphotyrosine peptides are most easily obtained by peptide synthesis using Fmoc-Tyr(PO3Me2)OH as building block. The dibenzylphosphochloridate method yields the expected phosphopeptides as the only peptide derivative and in addition, a great number of unidentified by-products which can be removed by ion-exchange chromatography. The phosphoramidite method consistently resulted in three peptide derivatives, i.e. the desired phosphopeptide, the phosphitylated peptide and a bridged derivative with two GGXA fragments linked through a phosphodiester bridge. The derivatives were characterised by RP and ion-exchange chromatography, 31P- and 1H-NMR spectroscopy, and ion-spray and electrospray mass spectrometry. Interestingly, even these mild ionisation techniques resulted in partial fragmentation. The observed fragmentation pathways seem to be a diagnostic tool for the identification of phosphorylation sites in peptides. Both the phosphorylated serine and threonine peptide lost phosphoric acid (98 mass units), the tyrosine peptide lost phenyl phosphate (174 mass units).(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1995

Multimedia medical case authorship and simulator program.

For the last several years, third and fourth year medical students rotating on the rheumatology/immunology service at the University of North Carolina School of Medicine have been using a laptop computer as a teaching adjunct to their formal training in rheumatology. The laptop contains diagnostic programs, reference management and clinical note generation facilities, remote medline access, and most recently, multimedia case simulations. These simulations have been created by the use of a case authoring and simulation system which is presented in this demonstration. The program is divided into simulator and designer modules and uses graphics and sound to portray such data as physical examination findings, blood smears, radiographs, heart sounds, etc. The simulator module includes diagnostic sections with feedback to the student as well as robust patient management trees with an occasional circuitous route for patient outcome. The student receives a numerical score based on deviations from the correct path and optimal cost as designated by the case designer. The system simulates complete management of a patient from the first encounter until treatment is complete. During each encounter, a student obtains the patient's history, physical examination findings, orders tests and reviews their results, makes a differential diagnosis, and treats the patient. The patient's progress and further treatment options at any time are dependent on the treatment option selected by the student at an earlier stage. Students are given the costs of ancillary tests and hospitalization before they order them. Words or phrases can be marked as hypertext and the student can get more information about the marked words by a mouse click. The designer interface of the program creates the clinical case by prompts and requests for information from the designer who needs no programming skills. The designer is almost always an expert faculty member who bases the simulated case on a real patient. Default and normal values for clinical findings and tests facilitate the case design. Graphics and sound files are easily incorporated into any historical, physical examination or test window. Management trees are automatically generated during the case design and the author is able to rapidly change to a new management scene for editing with a mouse click. Optimal critical path to correct cost effective diagnoses and treatment is designated and used as the standard to which the students' choices are compared. The software runs on PC's with 386 or better processors and is based in either Windows or OS/2 operating environments. The display can be VGA or SVGA, color or monochrome. Sound is played from 8 or 16 bit sound files. It can be readily extrapolated to any clinical medical specialty in the medical school or postgraduate curricula. The capabilities of this system to create unlimited, branched management and multiple clinical scenarios combined with the ease of authorship make this program unique and an excellent teaching tool.

Computer Simulation↗
PubMed1994

Report on the clinical workstation and clinical data repository utilization at UNC Hospitals.

On December 1, 1993, we implemented version 2.1 of the Clinical Workstation-Clinical Data Repository application in the Ambulatory Care Center. This version of the workstation allowed access of laboratory data from the clinical data repository that had been populated by a real-time HL7 interface between the Clinical Data Repository and the Laboratory Information System. This implementation completed a major part of the Clinical Workstation project. Also in December, we implemented a security system that records the date and time, user logon code, clinical workstation functions used, and the patient medical record number on whom data were displayed. In addition to the security function, this system has proven to be a valuable tool in evaluating the utilization of the clinical workstation and is the source of the data presented in this paper.

Clinical Laboratory Information Systems↗