Bacillus subtilis subsp. subtilis CBMDC3f with antimicrobial activity against Gram-positive foodborne pathogenic bacteria: UV-MALDI-TOF MS analysis of its bioactive compounds.

In this work a new Bacillus sp. strain, isolated from honey, was characterized phylogenetically. Its antibacterial activity against three relevant foodborne pathogenic bacteria was studied; the main bioactive metabolites were analyzed using ultraviolet matrix assisted laser desorption-ionization mass spectrometry (UV-MALDI MS). Bacillus CBMDC3f was phylogenetically characterized as Bacillus subtilis subsp. subtilis after rRNA analysis of the 16S subunit and the gyrA gene (access codes Genbank JX120508 and JX120516, respectively). Its antibacterial potential was evaluated against Listeria monocytogenes (9 strains), B. cereus (3 strains) and Staphylococcus aureus ATCC29213. Its cell suspension and cell-free supernatant (CFS) exerted significant anti-Listeria and anti-S. aureus activities, while the lipopeptides fraction (LF) also showed anti-B. cereus effect. The UV-MALDI-MS analysis revealed surfactin, iturin and fengycin in the CFS, whereas surfactin predominated in the LF. The CFS from CBMDC3f contained surfactin, iturin and fengycin with four, two and four homologues per family, respectively, whereas four surfactin, one iturin and one fengycin homologues were identified in the LF. For some surfactin homologues, their UV-MALDI-TOF/TOF (MS/MS; Laser Induced Decomposition method, LID) spectra were also obtained. Mass spectrometry analysis contributed with relevant information about the type of lipopeptides that Bacillus strains can synthesize. From our results, surfactin would be the main metabolite responsible for the antibacterial effect.

Antibacterial activity of naringin derivatives against pathogenic strains.

AIMS: To study the antimicrobial activity of naringin (NAR), a flavonoid extracted from citrus industry waste, and NAR derivatives [naringenin (NGE), prunin and alkyl prunin esters] against pathogenic bacteria such as L. monocytogenes, E. coli O157:H7 and S. aureus. The relationship between the structure of the chemical compounds and their antagonistic effect was also analysed. METHODS AND RESULTS: The agar dilution technique and direct contact assaying were applied. NGE, prunin and NAR showed no antimicrobial activity at a concentration of 0.25 mmol l(-1). Similarly, fatty acids with a chain length between C2 and C18 showed no antimicrobial activity at the same concentration. However, prunin-6″-O-acyl esters presented high antibacterial activity, mainly against Gram-positive strains. This activity increased with increasing chain length (up to 10-12 carbon atoms). Alkyl prunin esters with 10-12 carbon atoms diminished viability of L. monocytogenes by about 3 log orders and S. aureus by 6 log orders after 2 h of contact at 37°C and at a concentration of 0.25 mmol l(-1). The compounds examined were not effective against any of the Gram-negative strains assayed, even at the highest concentration. CONCLUSIONS: Addition of sugars to the aglycone did not enhance its antimicrobial activity. Attachment of a saturated aliphatic chain with 10-12 carbon atoms to the A ring of the flavonoid (or to sugars attached to this ring), seems to be the most promising modification. In conclusion, alkyl prunin esters with a chain length of C10-C12 have promising features as antimicrobial agents because of their high antilisterial and antistaphylococcal activity. SIGNIFICANCE AND IMPACT OF THE STUDY: This study shows that it is possible to obtain NAR derivatives with important antimicrobial activity, especially against Gram-positive pathogenic bacteria. It also provides guidelines on the structural modifications in similar molecules to enhance the antimicrobial activity.

Antimicrobial properties of prunin, a citric flavanone glucoside, and its prunin 6″-O-lauroyl ester.

AIMS: To determine the antimicrobial potential of prunin (P), a flavanone glucoside resulting from the hydrolysis of naringin present in grapefruit, and of its prunin 6″-O-lauroyl ester (PL), synthesized by enzymatic catalysis. METHODS AND RESULTS: P and its lauroyl ester were tested against Gram-negative and Gram-positive bacteria, yeasts and moulds. P showed no inhibitory effect against the micro-organisms assayed, but stimulated growth of Pseudomonas aeruginosa and different Bacilllus sp. However, 150 µg ml(-1) of PL inhibited Escherichia coli, Salmonella enterica serovar Enteritidis, Salmonella enterica serovar Typhimurium, many Bacillus sp., Staphylococcus aureus ATCC29213, Enterococcus avium DSMZ17511, and different Listeria monocytogenes strains. In the last case, L. monocytogenes, sensitive or bacteriocin-resistant cells, lost nearly 4-log reductions after 30 min of contact. A bactericidal mode of action was determined using both scanning and transmission electronic microscopies. CONCLUSIONS: PL could be used as a food additive, because at low concentration (150 µg ml(-1)) it exhibited antimicrobial activity against important food-borne pathogens. A bactericidal effect was also determined on L. monocytogenes sensitive and bacteriocin-resistant mutant strains. P did not show any antimicrobial property at all. SIGNIFICANCE AND IMPACT OF THE STUDY: PL is a potential antimicrobial compound with a high anti-Listeria property.

Enzymatic production of biodiesel from cotton seed oil using t-butanol as a solvent.

The enzymatic production of biodiesel by methanolysis of cottonseed oil was studied using immobilized Candida antarctica lipase as catalyst in t-butanol solvent. Methyl ester production and triacylglycerol disappearance were followed by HPLC chromatography. It was found, using a batch system, that enzyme inhibition caused by undissolved methanol was eliminated by adding t-butanol to the reaction medium, which also gave a noticeable increase of reaction rate and ester yield. The effect of t-butanol, methanol concentration and temperature on this system was determined. A methanolysis yield of 97% was observed after 24h at 50 degrees C with a reaction mixture containing 32.5% t-butanol, 13.5% methanol, 54% oil and 0.017 g enzyme (g oil)(-1). With the same mixture, a 95% ester yield was obtained using a one step fixed bed continuous reactor with a flow rate of 9.6 mlh(-1) (g enzyme)(-1). Experiments with the continuous reactor over 500 h did not show any appreciable decrease in ester yields.

Evolution of the Jahn-Teller distortion of MnO6 octahedra in RMnO3 perovskites (R = Pr, Nd, Dy, Tb, Ho, Er, Y): a neutron diffraction study.

Stoichiometric RMnO3 perovskites have been prepared in the widest range of R3+ ionic sizes, from PrMnO3 to ErMnO3. Soft-chemistry procedures have been employed; inert-atmosphere annealings were required to synthesize the materials with more basic R cations (R = Pr, Nd), in order to minimize the unwanted presence of Mn4+. On the contrary, annealings in O2 flow were necessary to stabilize the perovskite phases for the last terms of the series, HoMnO3, ErMnO3, and YMnO3, thus avoiding or minimizing the formation of competitive hexagonal phases with the same stoichiometry. The samples have been investigated at room temperature by high-resolution neutron powder diffraction to follow the evolution of the crystal structures along the series. The results are compared with reported data for LaMnO3. The distortion of the orthorhombic perovskite (space group Pbnm), characterized by the tilting angle of the MnO6 octahedra, progressively increases from Pr to Er due to simple steric factors. Additionally, all of the perovskites show a distortion of the MnO6 octahedra due to the orbital ordering characteristic of the Jahn-Teller effect of Mn3+ cations. The degree of orbital ordering slightly increases from La to Tb and then remains almost unchanged for the last terms of the series. The stability of the crystal structure is also discussed in light of bond-valence arguments.

Saccharification of native and degraded cotton cellulose and commercial microcrystalline cellulose by Trichoderma viride cellobiohydrolase I.

The degree of polymerization of samples of acid degraded cotton cellulose has no appreciable influence on the saccharification by cellobiohydrolase I from Trichoderma viride. The increase in the number of cellulose molecule ends, achieved by a 30-fold decrease in molecular weight, does not produce the effect which could be expected for a pure end-wise mode of action of this exoglucanase. Microcrystalline celluloses saccharified by the same enzyme yield considerably more reducing sugars than cotton cellulose, either with a similar degree of polymerization or one of about 7000. It appears, therefore, that the difference in the susceptibility of the commercial substrates is not a consequence of their low degree of polymerization.