Enhanced chaotrope tolerance and (S)-2-hydroxypropiophenone production by recombinant Pseudomonas putida engineered with Pprl from Deinococcus radiodurans.

Pseudomonas putida is a soil bacterium with multiple uses in fermentation and biotransformation processes. P. putida ATCC 12633 can biotransform benzaldehyde and other aldehydes into valuable α-hydroxyketones, such as (S)-2-hydroxypropiophenone. However, poor tolerance of this strain toward chaotropic aldehydes hampers efficient biotransformation processes. To circumvent this problem, we expressed the gene encoding the global regulator PprI from Deinococcus radiodurans, an inducer of pleiotropic proteins promoting DNA repair, in P. putida. Fine-tuned gene expression was achieved using an expression plasmid under the control of the LacIQ /Ptrc system, and the cross-protective role of PprI was assessed against multiple stress treatments. Moreover, the stress-tolerant P. putida strain was tested for 2-hydroxypropiophenone production using whole resting cells in the presence of relevant aldehyde substrates. P. putida cells harbouring the global transcriptional regulator exhibited high tolerance toward benzaldehyde, acetaldehyde, ethanol, butanol, NaCl, H2 O2 and thermal stress, thereby reflecting the multistress protection profile conferred by PprI. Additionally, the engineered cells converted aldehydes to 2-hydroxypropiophenone more efficiently than the parental P. putida strain. 2-Hydroxypropiophenone concentration reached 1.6 g L-1 upon a 3-h incubation under optimized conditions, at a cell concentration of 0.033 g wet cell weight mL-1 in the presence of 20 mM benzaldehyde and 600 mM acetaldehyde. Product yield and productivity were 0.74 g 2-HPP g-1 benzaldehyde and 0.089 g 2-HPP g cell dry weight-1 h-1 , respectively, 35% higher than the control experiments. Taken together, these results demonstrate that introducing PprI from D. radiodurans enhances chaotrope tolerance and 2-HPP production in P. putida ATCC 12633.

Optimized enantioselective (S)-2-hydroxypropiophenone synthesis by free- and encapsulated-resting cells of Pseudomonas putida.

BACKGROUND: Aromatic α-hydroxy ketones, such as S-2-hydroxypropiophenone (2-HPP), are highly valuable chiral building blocks useful for the synthesis of various pharmaceuticals and natural products. In the present study, enantioselective synthesis of 2-HPP was investigated by free and immobilized whole cells of Pseudomonas putida ATCC 12633 starting from readily-available aldehyde substrates. Whole resting cells of P. putida, previously grown in a culture medium containing ammonium mandelate, are a source of native benzoylformate decarboxylase (BFD) activity. BFD produced by induced P. putida resting cells is a highly active biocatalyst without any further treatment in comparison with partially purified enzyme preparations. These cells can convert benzaldehyde and acetaldehyde into the acyloin compound 2-HPP by BFD-catalyzed enantioselective cross-coupling reaction. RESULTS: The reaction was carried out in the presence of exogenous benzaldehyde (20 mM) and acetaldehyde (600 mM) as substrates in 6 mL of 200 mM phosphate buffer (pH 7) for 3 h. The optimal biomass concentration was assessed to be 0.006 g dry cell weight (DCW) mL- 1. 2-HPP titer, yield and productivity using the free cells were 1.2 g L- 1, 0.56 g 2-HPP/g benzaldehyde (0.4 mol 2-HPP/mol benzaldehyde), 0.067 g 2-HPP g- 1 DCW h- 1, respectively, under optimized biotransformation conditions (30 °C, 200 rpm). Calcium alginate (CA)-polyvinyl alcohol (PVA)-boric acid (BA)-beads were used for cell entrapment. Encapsulated whole-cells were successfully employed in four consecutive cycles for 2-HPP production under aerobic conditions without any noticeable beads degradation. Moreover, there was no production of benzyl alcohol as an unwanted by-product. CONCLUSIONS: Bioconversion by whole P. putida resting cells is an efficient strategy for the production of 2-HPP and other α-hydroxyketones.

Two new cytotoxic glycosides isolated from the green walnut husks of Juglans mandshurica Maxim.

Two new glycosides including an alcohol glycoside and a phenolic glycoside: hexyl-1-O-α-d-arabinofuranosyl-(1 â†’ 6)-ß-d-glucopyranoside (1), 4-hydroxypropiophenone-4-O-ß-d-glucopyranosyl(1 â†’ 6)-ß-d-glucopyranoside(2), along with six known naphthalenyl glucosides (3-8) were isolated from green walnut husks of Juglans mandshurica, and their structures were elucidated on the basis of spectroscopic studies. All compounds were evaluated for their inhibitory effects on tumour cells (BGC-823, HepG-2, MCF-7). The results showed that new compounds 1 and 2 had superior inhibitory activity in comparison with other naphthalenyl glucosides.

Regio- and Stereoselective Aliphatic-Aromatic Cross-Benzoin Reaction: Enzymatic Divergent Catalysis.

The catalytic asymmetric synthesis of chiral 2-hydroxy ketones by using different thiamine diphosphate dependent enzymes, namely benzaldehyde lyase from Pseudomonas fluorescens (PfBAL), a variant of benzoylformate decarboxylase from Pseudomonas putida (PpBFD-L461A), branched-chain 2-keto acid decarboxylase from Lactococcus lactis (LlKdcA) and a variant of pyruvate decarboxylase from Acetobacter pasteurianus (ApPDC-E469G), was studied. Starting with the same set of substrates, substituted benzaldehydes in combination with different aliphatic aldehydes, PfBAL and PpBFD-L461A selectively deliver the (R)- and (S)-2-hydroxy-propiophenone derivatives, respectively. The (R)- and (S)-phenylacetylcarbinol (1-hydroxy-1-phenylacetone) derivatives are accessible in a similar way using LlKdcA and ApPDC-E469G, respectively. In many cases excellent stereochemical purities (>98 % enantiomeric excess) could be achieved. Hence, the regio- and stereochemistry of the product in the asymmetric aliphatic-aromatic cross-benzoin reaction can be controlled solely by choice of the appropriate enzyme or enzyme variant.

Immobilization and characterization of benzoylformate decarboxylase from Pseudomonas putida on spherical silica carrier.

If an adequate biocatalyst is identified for a specific reaction, immobilization is one possibility to further improve its properties. The immobilization allows easy recycling, improves the enzyme performance, and it often enhances the stability of the enzyme. In this work, the immobilization of the benzoylformate decarboxylase (BFD) variant, BFD A460I-F464I, from Pseudomonas putida was accomplished on spherical silica. Silicagel is characterized by its high mechanical stability, which allows its application in different reactor types without restrictions. The covalently bound enzyme was characterized in terms of its activity, stability, and kinetics for the formation of chiral 2-hydroxypropiophenone (2-HPP) from benzaldehyde and acetaldehyde. Moreover, temperature as well as pressure dependency of immobilized BFD A460I-F464I activity and enantioselectivity were analyzed. The used wide-pore silicagel shows a good accessibility of the immobilized enzyme. The activity of the immobilized BFD A460I-F464I variant was determined to be 70% related to the activity of the free enzyme. Thereby, the enantioselectivity of the enzyme was not influenced by the immobilization. In addition, a pressure-induced change in stereoselectivity was found both for the free and for the immobilized enzyme. With increasing pressure, the enantiomeric excess (ee) of (R)-2-HPP can be increased from 44% (0.1 MPa) to 76% (200 MPa) for the free enzyme and from 43% (0.1 MPa) to 66% (200 MPa) for the immobilized enzyme.

Influence of reaction conditions on the enantioselectivity of biocatalyzed C-C bond formations under high pressure conditions.

Benzoylformate decarboxylase (BFD, EC 4.1.1.7) is a homotetrameric thiamine diphosphate (ThDP)-dependent enzyme which catalyzes the synthesis of chiral 2-hydroxyketones accepting a broad range of aldehydes as substrates. In this study the synthesis of 2-hydroxypropiophenone (2-HPP) from benzaldehyde and acetaldehyde was catalyzed by three BFD variants namely BFD F464I, BFD A460I and BFD A460I-F464I. This paper reports the effect of hydrostatic pressure up to 290 MPa when the reactions were carried out at different benzaldehyde concentrations (5-40 mM) as well as at different pH values (7.0-8.5). Acetaldehyde concentration was fixed at 400 mM in all biotransformations. Reactions performed at high benzaldehyde concentrations and at high hydrostatic pressures showed an increase in (R)-2-HPP formation catalyzed by all BFD variants. For BFD A460I-F464I we observed an increase in the ee of (R)-2-HPP up to 80%, whereas at atmospheric conditions this variant synthesizes (R)-2-HPP with an ee of only 50%. Alkaline conditions (up to pH 8.5) and high hydrostatic pressures resulted in an increase of (R)-2-HPP synthesis, especially in the case of BFD A460I and BFD F464I.

Influence of the hydrostatic pressure and pH on the asymmetric 2-hydroxyketone formation catalyzed by Pseudomonas putida benzoylformate decarboxylase and variants thereof.

Benzoylformate decarboxylase (BFD) from Pseudomonas putida is a thiamine diphosphate-dependent (ThDP) enzyme that catalyzes the asymmetric C--C bond formation to (S)-2-hydroxypropiophenone [(S)-HPP] starting from benzaldehyde and acetaldehyde. The enantioselectivity of BFD was shown to be a function of temperature and substrate concentration. It can additionally be changed by site-directed mutagenesis on hot spot positions in the active site. In this article, we present the effect of hydrostatic pressure up to 250 MPa on the enantioselectivity for the recombinant wtBFD as well as for the variants BFD F464I, BFD A460I, and BFD A460I-F464I. A general tendency toward lower amounts of (S)-HPP could be observed at increasing pressures. For two of these variants an increase in pressure even caused an inversion in the enantioselectivity and thus increasing enantiomeric excesses, respectively. A pressure-induced increase in enantioselectivity could therefore be observed for the first time in biocatalysis to the best of our knowledge. Furthermore, the pH is shown to be a parameter that also significantly influences the enantioselectivity of the reaction mentioned above.

Métodos convencionais e moleculares para o diagnóstico da tuberculose pulmonar: um estudo comparativo / Conventional and molecular techniques in the diagnosis of pulmonary tuberculosis: a comparative study

OBJETIVO: Comparar quatro métodos laboratoriais no diagnóstico de tuberculose pulmonar. MÉTODOS: Foram realizadas pesquisa direta pelas colorações de Ziehl-Neelsen e auramina, cultura para micobactérias em meio Lõwenstein-Jensen (LJ) e polymerase chain reaction (PCR, reação em cadeia da polimerase) para Mycobacterium tuberculosis em 160 amostras de secreção respiratória de pacientes com suspeita de tuberculose pulmonar. As cepas isoladas foram identificadas por método radiométrico utilizando-se p-nitro-alfa-acetilamino-beta-hidroxipropiofenona (NAP) e métodos clássicos. A sensibilidade dos métodos foi comparada com o padrão ouro para o diagnóstico da tuberculose pulmonar, definido por critérios clínicos, radiológicos e microbiológicos. RESULTADOS: Dos 160 pacientes, 142 foram diagnosticados com tuberculose pulmonar de acordo com o padrão ouro. As técnicas de Ziehl-Neelsen e auramina, cultura em meio LJ e PCR apresentaram sensibilidade de 54,2 por cento, 58,4 por cento, 67,6 por cento e 77,5 por cento, respectivamente, quando comparados ao critério diagnóstico adotado. A especificidade dos quatro métodos foi de 100 por cento. A concordância na identificação da micobactéria entre PCR e o método radiométrico utilizando NAP foi alta (96,8 por cento). A sensibilidade da PCR foi de 50,8 por cento nas amostras com baciloscopia negativa e de 98,8 por cento naquelas com baciloscopia positiva. Nas amostras com resultados negativos na baciloscopia e cultura, a sensibilidade da PCR foi menor que nas com resultados positivos (25,6 por cento e 99,0 por cento, respectivamente). CONCLUSÕES: A PCR é método promissor no diagnóstico da tuberculose pulmonar, mesmo em amostras paucibacilares. Além disso, apresenta a vantagem da identificação simultânea e rapidez do resultado.

OBJECTIVE: To compare four laboratory methods in the diagnosis of pulmonary tuberculosis. METHODS: Respiratory secretion specimens were collected from 160 patients suspected of having pulmonary tuberculosis. Direct testing for Mycobacterium tuberculosis was carried out using Ziehl-Neelsen and auramine staining. In addition, culture in Lõwenstein-Jensen (LJ) medium and polymerase chain reaction (PCR) were used. The strains isolated were identified by means of a radiometric method using p-nitro-alpha-acetylamino-beta-hydroxypropiophenone (NAP) and classical methods. The sensitivity of the methods was compared to the gold standard for the diagnosis of pulmonary tuberculosis, based on clinical, radiological and microbiological criteria. RESULTS: Of the 160 patients, 142 were diagnosed with pulmonary tuberculosis according to the gold standard. The sensitivity of Ziehl-Neelsen staining, auramine staining, culture in LJ medium and PCR was 54.2 percent, 58.4 percent, 67.6 percent and 77.5 percent, respectively, when compared with the diagnostic criterion adopted. All four methods presented 100 percent specificity. In the identification of mycobacteria, there was high (96.8 percent) concordance between PCR and the radiometric method using NAP. The sensitivity of PCR was 50.8 percent in samples with negative sputum smear microscopy results and 98.8 percent in those with positive results. The sensitivity of PCR was lower in specimens with negative results in sputum smear microscopy and culture than in those with positive results (25.6 percent and 99.0 percent, respectively). CONCLUSIONS: We found PCR to be a promising method for the diagnosis of pulmonary tuberculosis, even in paucibacillary specimens. Simultaneous identification and faster results are additional advantages of this method.

Use of in-house PCR for identification of Mycobacterium tuberculosis in BACTEC broth cultures of respiratory specimens.

We evaluated the ability of a PCR assay to identify Mycobacterium tuberculosis complex (MTBC) from positive BACTEC 12B broth cultures. A total of 107 sputum samples were processed and inoculated into Ogawa slants and BACTEC 12B vials. At a growth index (GI) > or=30, 1.0 ml of the 12B broth was removed, stored, and assayed with PCR. Molecular results were compared to those obtained by phenotypic identification methods, including the BACTEC NAP method. The average times required to perform PCR and NAP were compared. Of the 107 broth cultures evaluated, 90 were NAP positive, while 91 were PCR positive for MTBC. Of particular interest were three contaminated BACTEC 12B broth cultures yielding microorganisms other than acid-fast bacilli growth with a MTBC that were successfully identified by PCR, resulting in a mean time of 14 days to identify MTBC before NAP identification. These results suggest that PCR could be used as an alternative to the NAP test for the rapid identification of MTBC in BACTEC 12B cultures, particularly in those that contained both MTBC and nontuberculous mycobacteria.

Use of in-house PCR for identification of Mycobacterium tuberculosis in BACTEC broth cultures of respiratory specimens

We evaluated the ability of a PCR assay to identify Mycobacterium tuberculosis complex (MTBC) from positive BACTEC® 12B broth cultures. A total of 107 sputum samples were processed and inoculated into Ogawa slants and BACTEC® 12B vials. At a growth index (GI) > 30, 1.0 ml of the 12B broth was removed, stored, and assayed with PCR. Molecular results were compared to those obtained by phenotypic identification methods, including the BACTEC® NAP method. The average times required to perform PCR and NAP were compared. Of the 107 broth cultures evaluated, 90 were NAP positive, while 91 were PCR positive for MTBC. Of particular interest were three contaminated BACTEC® 12B broth cultures yielding microorganisms other than acid-fast bacilli growth with a MTBC that were successfully identified by PCR, resulting in a mean time of 14 days to identify MTBC before NAP identification. These results suggest that PCR could be used as an alternative to the NAP test for the rapid identification of MTBC in BACTEC® 12B cultures, particularly in those that contained both MTBC and nontuberculous mycobacteria.

Conventional and molecular techniques in the diagnosis of pulmonary tuberculosis: a comparative study.

OBJECTIVE: To compare four laboratory methods in the diagnosis of pulmonary tuberculosis. METHODS: Respiratory secretion specimens were collected from 160 patients suspected of having pulmonary tuberculosis. Direct testing for Mycobacterium tuberculosis was carried out using Ziehl-Neelsen and auramine staining. In addition, culture in Löwenstein-Jensen (LJ) medium and polymerase chain reaction (PCR) were used. The strains isolated were identified by means of a radiometric method using p-nitro-alpha-acetylamino-beta-hydroxypropiophenone (NAP) and classical methods. The sensitivity of the methods was compared to the gold standard for the diagnosis of pulmonary tuberculosis, based on clinical, radiological and microbiological criteria. RESULTS: Of the 160 patients, 142 were diagnosed with pulmonary tuberculosis according to the gold standard. The sensitivity of Ziehl-Neelsen staining, auramine staining, culture in LJ medium and PCR was 54.2%, 58.4%, 67.6% and 77.5%, respectively, when compared with the diagnostic criterion adopted. All four methods presented 100% specificity. In the identification of mycobacteria, there was high (96.8%) concordance between PCR and the radiometric method using NAP. The sensitivity of PCR was 50.8% in samples with negative sputum smear microscopy results and 98.8% in those with positive results. The sensitivity of PCR was lower in specimens with negative results in sputum smear microscopy and culture than in those with positive results (25.6% and 99.0%, respectively). CONCLUSIONS: We found PCR to be a promising method for the diagnosis of pulmonary tuberculosis, even in paucibacillary specimens. Simultaneous identification and faster results are additional advantages of this method.

Lipase-catalyzed chemo- and enantioselective acetylation of 2-alkyl/aryl-3-hydroxypropiophenones.

The chemo- and enantioselective capabilities of porcine pancreatic lipase (PPL) in tetrahydrofuran, and Candida rugosa lipase (CRL) in diisopropyl ether have been investigated for the acetylation of racemic 2-alkyl/aryl-3-hydroxypropiophenones, which are important precursors in the synthesis of biologically active chromanones and isoflavanones. A highly chemoselective acetylation of primary hydroxy group in preference to phenolic hydroxy group leading to the formation of enantiomerically enriched monoacetates has been observed.

Conversion of 4-hydroxyacetophenone into 4-phenyl acetate by a flavin adenine dinucleotide-containing Baeyer-Villiger-type monooxygenase.

An arylketone monooxygenase was purified from Pseudomonas putida JD1 by ion exchange and affinity chromatography. It had the characteristics of a Baeyer-Villiger-type monooxygenase and converted its substrate, 4-hydroxyacetophenone, into 4-hydroxyphenyl acetate with the consumption of one molecule of oxygen and oxidation of one molecule of NADPH per molecule of substrate. The enzyme was a monomer with an M(r) of about 70,000 and contained one molecule of flavin adenine dinucleotide (FAD). The enzyme was specific for NADPH as the electron donor, and spectral studies showed rapid reduction of the FAD by NADPH but not by NADH. Other arylketones were substrates, including acetophenone and 4-hydroxypropiophenone, which were converted into phenyl acetate and 4-hydroxyphenyl propionate, respectively. The enzyme displayed Michaelis-Menten kinetics with apparent K(m) values of 47 microM for 4-hydroxyacetophenone, 384 microM for acetophenone, and 23 microM for 4-hydroxypropiophenone. The apparent K(m) value for NADPH with 4-hydroxyacetophenone as substrate was 17.5 microM. The N-terminal sequence did not show any similarity to other proteins, but an internal sequence was very similar to part of the proposed NADPH binding site in the Baeyer-Villiger monooxygenase cyclohexanone monooxygenase from an Acinetobacter sp.

Methods for the rapid identification of mycobacterial species.

The use of rapid tests for the identification of mycobacteria has been advocated primarily for Mycobacterium tuberculosis; however, they have been accepted less widely than expected. Chromatographic methods are best suited for larger laboratories, whereas nucleic acid probes may be used by laboratories that can justify the cost based on volume and pricing. Nucleic acid sequencing offers the possibility of providing the most exact identification of species of mycobacteria, but its use is limited to reference laboratories that have the applicable resources. Because of their clinical importance, all mycobacteria should be identified using the most rapid methods available.

Alternative use of polymerase chain reaction instead of rho-nitro-alpha-acetylamino-beta-hydroxypropiophenone test for the early detection of Mycobacterium tuberculosis in BACTEC 12B cultures.

Compared with conventional culture media, the TB BACTEC system has demonstrated improved isolation rates as well as an earlier detection time for mycobacterial species. However, the identification of M. tuberculosis by the rho-nitro-alpha-acetylamino-beta-hydroxypropiophenone (NAP) test in the TB BACTEC 460 system may require 6 days for interpretable results. We evaluated the usefulness of a polymerase chain reaction (PCR) assay for earlier identification of M. tuberculosis in positive BACTEC 12B cultures. A total of 262 TB BACTEC culture specimens with GIs > or = 10 were assayed by PCR, and the results were compared with those of the NAP test. The aliquot from BACTEC 12B vials was boiled for 10 min, and 2 microliters of the boiled suspension was used for the PCR assay. One set of primers based on the IS 6110 sequence of M. tuberculosis was used to amplify a 457 bp fragment of DNA. Of the 173 TB BACTEC culture specimens which were identified as M. tuberculosis by the NAP test. 171 were PCR positive. Of the 21 TB BACTEC cultures identified as MOTT by the NAP test. 19 were PCR negative, but 2 were PCR positive: these two cultures were shown to grow both M. tuberculosis and MOTT in BACTEC 12B vials. Of the remaining 68 cultures which were contaminated with AFB-negative bacteria, the PCR identified M. tuberculosis in 13, in agreement with the NAP results in the reprocessed specimens. Overall, the PCR results in the 262 BACTEC culture specimens with GIs > or = 10 were sensitive in 99.5% (186/187) and specific in 100% (68/68). The mean time for the identification of M. tuberculosis in TB BACTEC cultures with GIs > or = 10 was 7 h by the PCR compared to 5.9 days by the NAP test. These results suggest that the PCR could be used as an alternative to the NAP test for the rapid identification of M. tuberculosis in BACTEC 12B cultures, particularly in those which contained both M. tuberculosis and MOTT or M. tuberculosis and AFB-negative bacteria.

[Disseminated Mycobacterium genavense infection in patients with HIV infection. Description of 5 cases and review of the literature]. / Infección diseminada por Mycobacterium genavense en pacientes con infección por HIV. Descripción de 5 casos y revisión de la literatura.

BACKGROUND: Five cases of disseminated infection by Mycobacterium genavense in patients with HIV infection are reported with a review of the literature. MATERIAL AND METHODS: A description of the clinical, epidemiologic and therapeutic characteristics of five cases are presented. The initial isolation of the microorganism was performed in Bactec 13A from blood and bone marrow aspirates. Subcultures were made in different media and the growth characteristics evaluated. Sensitivity to NAP was determined by radiometric techniques and gas chromatography allowed a possible identification. Definitive identification was based on PCR amplification of the gene which codifies the 65kDa protein and the posterior restriction of the amplified fragments by using BstEII and HaeIII. RESULTS: All five patients were males with HIV infection and a lymphocyte count of less than 25 cells/mm3 with an non-specific clinical picture similar to that produced by M. avium complex (MAC). Empiric antiMAC treatment was administered in four of the patients with good clinical response. All five strains were sensitive to NAP. The greatest growth rate was obtained in the subcultures with acid pH in liquid medium. Gas chromatography was very similar to that of M. simiae. Amplification of the gene which codifies the 65 kDa protein and posterior restriction with BstEII resulted in 2 fragments of 325 and 125 bp, while restriction with HaeIII resulted in two fragments of 140 and 105 bp. CONCLUSIONS: Mycobacterium genavense represents 9% of the disseminated infections by mycobacteria in AIDS patients. The clinical manifestations, empiric treatment and response is similar to that of infection by M. avium complex. Growth is favored by acid pH in liquid medium. Susceptibility to NAP leads to possible identification which should be confirmed by molecular techniques.

[Anti-calcium effects of 4-hydroxypropiophenone derivatives--potential beta-adrenoreceptor blockers]. / Antikalciový efekt derivátov 4-hydroxypropiofenónu--potenciálnych blokátorov beta-adrenoreceptorov.

The present paper carries out the pharmacological evaluation of 4-(2-hydroxy-3-isopropylaminopropoxy)-3-(alkoxymethyl) propiophenones with an ethoxy, propoxy and butoxy-group, whose structures are typical of the blockers of beta-adrenergic receptors. In the above-mentioned compounds the anticalcium effect on the frequency and the amplitude and the negative chronotropic and the negative inotropic effects were evaluated by means of the method of spontaneously pulsing guinea-pig atria within a concentration range of 4-16 microgram.cm-3. The obtained results confirmed a significant anticalcium effect on the heart rate in the compounds with a propoxy and a butoxy group, and in the standard verapamil. In all three compounds as well as in the standard verapamil, no anticalcium effect on the amplitude was found. The results of this membrane efficacy are in agreement with the preceding evaluation of the antidysrhythmic and anti-isoprenaline activity with the most marked effect in the compound with a propoxy group. On the basis of these results it is possible to conclude that in these derivatives of 4-hydroxypropiophenone also the anticalcium effect can participate, besides the beta-adrenolytic and the membranostabilizing effects, in the antidysrhythmic activity.

[Changes in basic nitrogen in a group of potential new beta- adrenolytics--derivatives of p-hydroxyacetophenone and p-hydroxypropiofenone]. / Obmeny na bázickom dusíku v skupine nových potenciálnych betaadrenolytík--derivátov p-hydroxyacetofenónu a p-hydroxypropiofenónu.

Within the relationship of the structure and effect of new beta-adrenolytic agents derivatived from p-hydroxyacetophenone and p-hydroxypropiophenone with a propoxymethyl group in the lipophilic part of the molecule and with a propanamine, a butanamine and a pyrrolidine in the side-chain were studied. In order to prepare these substances, a procedure was selected from several tested ones, in which 4-hydroxy-3propoxymethylphenylketone were treated with chloromethyloxirane and subsequent reaction a hydrobromic acid were prepared 4-(3-brom-2-hydroxypropoxy)-3-propoxymethylalkylketone. Final substances were prepared reaction with amine. The structure of prepared compounds was confirmed on the basic interpretation of the IR, UV and 1H NMR spectra. The results of pharmacological evaluation of selected compounds showed a significant beta 1-blocking activity lower than acebutolol. Their local anesthetic activity is low according with their partition coefficients. The characteristic of the prepared compounds was supplemented by the determination of their partition coefficients, surface tension, dissociation constants and acute toxicity.

Acyloin formation by benzoylformate decarboxylase from Pseudomonas putida.

Whole cells and cell extracts of Pseudomonas putida grown in a medium containing ammonium mandelate have the capacity to produce the acyloin compound 2-hydroxypropiophenone when incubated with benzoylformate and acetaldehyde. Benzaldehyde and benzyl alcohol were formed as reaction by-products. The enantiomeric excess of the 2-hydroxypropiophenone product was found to be 91 to 92%. The absolute configuration of the enzymatically prepared product at the carbinol carbon was found to be S. The thiamine PPi-linked enzyme benzoylformate decarboxylase, purified to give a single protein band on polyacrylamide gel electrophoresis, was shown to be responsible for the catalysis of this novel condensation reaction.

Chemiexcitation in the peroxidative metabolism of diethylstilbestrol. Metabolic products.

In the presence of the surfactant hexadecyltrimethyl ammonium bromide (CTAB) a cascade of electronically excited states accompanies the successive steps in the peroxidative metabolization of the strong estrogenic and tumourogenic diethylstilbestrol. Reversing the order by necessity, we report in this first paper results with the metabolites. Exposure of 4-hydroxypropiophenone, Z,Z-dienestrol or E,E-dienestrol to horseradish peroxidase and H2O2 promotes oxygen uptake and spectral alterations. Light emission is observed provided that the surfactant CTAB is present. With the three substrates, 4-hydroxybenzoic acid and a new metabolite, p-benzoquinone, have been identified. With both dienestrol isomers, 1-(4'-hydroxyphenyl)-propan-1-on-2-ol has been identified. In all cases the emission spectrum indicates the presence of several emitters. Possible chemiexcitation routes are pointed out. From the dramatic increase of the emission by enhancers, values as high as 1 x 10(-5) are inferred for the product of the quantum yields of chemiexcitation and energy transfer.