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Pharmaceutical industry has been encountering antimicrobial activity of non-antibiotics during suitability tests carried out prior to routine pharmacopoeial microbiological purity ...analysis of finished dosage forms. These properties are usually ignored or perceived as a nuisance during pharmaceutical analysis.
The aim of this study was: (i) to compare the available data to our method suitability test results carried out on products containing ibuprofen, i.e. to demonstrate that method suitability can be a valuable tool in identifying new antimicrobials, (ii) to demonstrate the antimicrobial activity of ibuprofen and ibuprofen lysine.
Microbiological purity method suitability testing was carried out according to European Pharmacopoeia (EP), chapters 2.6.12. and 2.6.13. Antimicrobial activity of ibuprofen and ibuprofen lysine was demonstrated by a disk diffusion method, a modification of the European Committee for Antimicrobial Susceptibility Testing method (EUCAST), against test microorganisms recommended in the EP.
It was confirmed that ibuprofen may be responsible for the broad spectrum of antimicrobial activity of the tested products, and that method suitability tests according to the EP can indeed be exploited by the scientific community in setting guidelines towards future research of new antimicrobials. In the disk diffusion assay, inhibition zones were obtained with more than 62.5μg and 250μg for Staphylococcus aureus, 125μg and 250μg for Bacillus subtilis, 31.3μg and 125μg for Candidaalbicans, 31.3μg and 62.5μg for Aspergillusbrasiliensis, of ibuprofen/disk, and ibuprofen lysine/disk, respectively. For Escherichiacoli, Pseudomonasaeruginosa and Salmonellatyphimurium inhibition zones were not obtained.
Antimicrobial activity of ibuprofen is considered merely as a side effect, and it is not mentioned in the patient information leaflets of ibuprofen drugs. As such, for the patient, it could represent an advantage, but, it could also introduce additional risks during usage. Further microbiological, pharmacological and clinical trials are of great importance.
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Bioactive peptides (BPs) are molecules of paramount importance with great potential for the development of functional foods, nutraceuticals or therapeutics for the prevention or treatment of various ...diseases. A functional BP-rich dairy product was produced by lyophilisation of bovine milk fermented by the autochthonous strains
subsp.
ZGBP5-51,
ZGBP5-52 and
ZGBP5-53 isolated from the same artisanal fresh cheese. The efficiency of the proteolytic system of the implemented strains in the production of BPs was confirmed by a combined high-throughput mass spectrometry (MS)-based peptidome profiling and an in silico approach. First, peptides released by microbial fermentation were identified via a non-targeted peptide analysis (NTA) comprising reversed-phase nano-liquid chromatography (RP nano-LC) coupled with matrix-assisted laser desorption/ionisation-time-of-flight/time-of-flight (MALDI-TOF/TOF) MS, and then quantified by targeted peptide analysis (TA) involving RP ultrahigh-performance LC (RP-UHPLC) coupled with triple-quadrupole MS (QQQ-MS). A combined database and literature search revealed that 10 of the 25 peptides identified in this work have bioactive properties described in the literature. Finally, by combining the output of MS-based peptidome profiling with in silico bioactivity prediction tools, three peptides (
QFLPYPYYAKPA
,
VAPFPEVFGK
,
ARHPHPHLSF
), whose bioactive properties have not been previously reported in the literature, were identified as potential BP candidates.
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Lactiplantibacillus plantarum D13 shows antistaphylococcal and antilisterial activity, probably due to the synthesis of a presumptive bacteriocin with antibiofilm capacity released in the cell-free ...supernatant (CFS), whose inhibitory effect is enhanced by cocultivation with susceptible strains. An in silico analysis of the genome of strain D13 confirmed the pln gene cluster. Genes associated with plantaricin biosynthesis, structure, transport, antimicrobial activity, and immunity of strain D13 were identified. Furthermore, the predicted homology-based 3D structures of the cyclic conformation of PlnE, PlnF, PlnJ, and PlnK revealed that PlnE and PlnK contain two helices, while PlnF and PlnJ contain one and two helices, respectively. The potential of the strain to modulate the intestinal microbiota in healthy or dextran sulphate sodium (DSS)-induced colitis mouse models was also investigated. Strain D13 decreased the disease activity index (DAI) and altered the gut microbiota of mice with DSS-induced colitis by increasing the ratio of beneficial microbial species (Allobaculum, Barnesiella) and decreasing those associated with inflammatory bowel disease (Candidatus Saccharimonas). This suggests that strain D13 helps to restore the gut microbiota after DSS-induced colitis, indicating its potential for further investigation as a probiotic strain for the prevention and treatment of colitis.
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The antimicrobial activity of industrially important lactic acid bacteria as starter cultures and probiotic bacteria is the main subject of this review. This activity has been attributed to the ...production of metabolites such as organic acids (lactic and acetic acid), hydrogen peroxide, ethanol, diacetyl, acetaldehyde, acetoine, carbon dioxide, reuterin, reutericyclin and bacteriocins. The potential of using bacteriocins of lactic acid bacteria, primarily used as biopreservatives, represents a perspective, alternative antimicrobial strategy for continuously increasing problem with antibiotic resistance. Another strategy in resolving this problem is an application of probiotics for different gastrointestinal and urogenital infection therapies. Key words: antimicrobial activity, bacteriocins, lactic acid bacteria, probiotics, starter cultures
Human milk not only provides a perfect balance of nutrients to meet all the needs of the infant in the first months of life but also contains a variety of bacteria that play a key role in tailoring ...the neonatal faecal microbiome. Microbiome analysis of human milk and infant faeces from mother-breastfed infant pairs was performed by sequencing the V1-V3 region of the 16S rRNA gene using the Illumina MiSeq platform. According to the results, there is a connection in the composition of the microbiome in each mother-breastfed infant pair, supporting the hypothesis that the infant's gut is colonised with bacteria from human milk. MiSeq sequencing also revealed high biodiversity of the human milk microbiome and the infant faecal microbiome, whose composition changes during lactation and infant development, respectively. A total of 28 genetically distinct strains were selected by hierarchical cluster analysis of RAPD-PCR (Random Amplified Polymorphic DNA-Polymerase Chain Reaction) electrophoresis profiles of 100 strains isolated from human milk and identified by 16S RNA sequencing. Since certain cellular molecules may support their use as probiotics, the next focus was to detect (S)-layer proteins, bacteriocins and exopolysaccharides (EPSs) that have potential as therapeutic biomolecules. SDS-PAGE (Sodium Dodecyl-Sulfate Polyacrylamide Gel Electrophoresis) coupled with LC-MS (liquid chromatography-mass spectrometry) analysis revealed that four
strains expressed S-layer proteins, which were identified for the first time in strains isolated from human milk. The potential biosynthesis of plantaricin was detected in six
strains by PCR analysis and
antibacterial studies.
H NMR (Proton Nuclear Magnetic Resonance) analysis confirmed EPS production in only one strain,
MC1. The overall microbiome analysis suggests that human milk contributes to the establishment of the intestinal microbiota of infants. In addition, it is a promising source of novel
strains expressing specific functional biomolecules.
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This study aimed to define a consortium of lactic acid bacteria (LAB) that will bring added value to dried fresh cheese through specific probiotic properties and the synthesis of bioactive peptides ...(biopeptides). The designed LAB consortium consisted of three
strains: S-layer carrying
D6, exopolysaccharides producing
D12 and plantaricin expressing
D13, and one
strain,
ZGZA7-10. Chosen autochthonous LAB strains exhibited efficient adherence to the Caco-2 cell line and impacted faecal microbiota biodiversity. The cheese produced by the LAB consortium showed better physicochemical, textural and sensory properties than the cheese produced by a commercial starter culture. Liquid chromatography coupled with matrix-assisted laser desorption/ionization-time of flight tandem mass spectrometry (LC-MALDI-TOF/TOF) showed the presence of 18 specific biopeptides in dried fresh cheeses. Their identification and relative quantification was confirmed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) using multiple reaction monitoring (MRM). The results also showed that their synthesis resulted mainly from β-casein and also α-S1 casein degradation by proteolytic activities of the LAB consortium. The designed LAB consortium enhanced the functional value of the final product through impact on biopeptide concentrations and specific probiotic properties.
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Increasing awareness of the importance of nutrition in health promotion and disease prevention has driven to the development of foods for special medical purposes (FSMPs). In this study, the ...probiotic strain
ssp.
(
431
) was incorporated into FSMPs to develop an innovative product. The aim was to investigate the influence of the FSMP matrix on the specific probiotic properties of
431
.
A series of
experiments were performed as part of the probiotic approach. After evaluation of antibiotic susceptibility profiles, functional properties such as survival under simulated gastrointestinal tract (GIT) conditions, bile salt deconjugation activities, cholesterol assimilation, antagonistic activity against spoilage bacteria and adhesion to Caco-2 cell line monolayers and extracellular matrix proteins were investigated.
The
431
strain, both the lyophilised strain and the strain isolated from the FSMP matrix, effectively survived the simulated adverse gastrointestinal conditions without significant effects of the food matrix. The effect of the FSMP matrix on the deconjugation activity of the bile salts of
431
was minimal; however, cholesterol assimilation was increased by 16.4 %.
431
had antibacterial activity against related lactic acid bacteria regardless of whether it was used in FSMPs or not. Conversely, the probiotic strain isolated from FSMP matrix had significantly higher inhibitory activity against six potential pathogens than the lyophilised culture. The autoaggregation ability of the
431
cells was not affected by the FSMP matrix. The adhesion of
431
bacterial cells to the extracellular matrix proteins was reduced after treatment with proteinase K, with the highest adhesion observed to laminin. The adhesion of
431
reduced the binding of
3014 by 1.81 log units and the binding of
. Typhimurium FP1 to Caco-2 cell lines by 1.85 log units, suggesting the potential for competitive exclusion of these pathogens.
The results support the positive effect of the FSMP matrix on the specific probiotic properties of
431
, such as antibacterial activity, bile salt deconjugation and cholesterol assimilation, while the incorporation of this probiotic strain adds functional value to the FSMPs. The synergistic effect achieved by the joint application of
431
and innovative FSMP matrix contributed to the development of the novel formulation of an improved functional food product with added value.
Lactobacillus (Limosilactobacillus) fermentum D12 is an exopolysaccharide (EPS) producing strain whose genome contains a putative eps operon. Whole-genome analysis of D12 was performed to disclose ...the essential genes correlated with activation of precursor molecules, elongation and export of the polysaccharide chain, and regulation of EPS synthesis. These included the genes required for EPS biosynthesis such as epsA, B, C, D and E, also gt, wzx, and wzy and those involved in the activation of the precursor molecules galE, galT and galU. Both the biosynthesis and export mechanism of EPS were proposed based on functional annotation. When grown on MRS broth with an additional 2% w/v glucose, L. fermentum D12 secreted up to 200 mg/L of a mixture of EPSs, whose porous structure was visualized by scanning electron microscopy (SEM). Structural information obtained by
HNMR spectroscopy together with composition and linkage analyses, suggested the presence of at least two different EPSs, a branched heteropolysaccharide containing t-Glcp and 2,6-linked Galf, and glycogen. Since recent reports showed that polysaccharides facilitate the probiotic-host interactions, we at first sought to evaluate the functional potential of L. fermentum D12. Strain D12 survived simulated gastrointestinal tract (GIT) conditions, exhibited antibacterial activity against enteropathogenic bacteria, adhered to Caco-2 cells in vitro, and as such showed potential for in vivo functionality. The EPS crude extract positively influenced D12 strain capacity to survive during freeze-drying and to adhere to extracellular matrix (ECM) proteins but did not interfere Caco-2 and mucin adherence when added at concentrations of 0.2, 0.5, and 1.0 mg/mL. Since the viable bacterial count of free D12 cells was 3 logarithmic units lower after the exposure to simulated GIT conditions than the initial count, the bacterial cells had been loaded into alginate for viability improvement. Microspheres of D12 cells, which were previously analyzed at SEM, significantly influenced their survival during freeze-drying and in simulated GIT conditions. Furthermore, the addition of the prebiotic substrates mannitol and lactulose improved the viability of L. fermentum D12 in freeze-dried alginate microspheres during 1-year storage at 4 °C compared to the control.
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Prof Dr Sc Marijan Bosnjak Suskovic, Jagoda; Kos, Blazenka
Food technology and biotechnology,
01/2020, Volume:
58, Issue:
1
Journal Article
Peer reviewed
Open access
An obituary for professor Dr Sc Marijan Bosnjak, professor at the Faculty of Food Technology and Biotechnology at University of Zagreb, who died on Feb 11, 2020, at the age of 86 is presented. He is ...the author of about 100 scientific and professional papers published in the national and international journals and proceedings, one university textbook, three patents and a scientific book 'Introduction to the Kinetics of Microbial Processes' published in 2009.
We evaluated the functional capacity of plantaricin-producing Lactobacillus plantarum SF9C and S-layer-carrying Lactobacillus brevis SF9B to withstand gastrointestinal transit and to compete among ...the gut microbiota in vivo. Considering the probiotic potential of Lb. brevis SF9B, this study aims to investigate the antibacterial activity of Lb. plantarum SF9C and their potential for in vivo colonisation in rats, which could be the basis for the investigation of their synergistic functionality.
A plantaricin-encoding cluster was identified in Lb. plantarum SF9C, a strain which efficiently inhibited the growth of Listeria monocytogenes ATCC
19111™ and Staphylococcus aureus 3048. Homology-based three-dimensional (3D) structures of SF9C plantaricins PlnJK and PlnEF were predicted using SWISS-MODEL workspace and the helical wheel representations of the plantaricin peptide helices were generated by HELIQUEST. Contrary to the plantaricin-producing SF9C strain, the S-layer-carrying SF9B strain excluded Escherichia coli 3014 and Salmonella enterica serovar Typhimurium FP1 from the adhesion to Caco-2 cells. Finally, PCR-DGGE analysis of the V2-V3 regions of the 16S rRNA gene confirmed the transit of the two selected lactobacilli through the gastrointestinal tract (GIT). Microbiome profiling via the Illumina MiSeq platform revealed the prevalence of Lactobacillus spp. in the gut microbiota of the Lactobacillus-treated rats, even on the 10th day after the Lactobacillus application, compared to the microbiota of the healthy and AlCl
-exposed rats before Lactobacillus treatment.
The combined application of Lb. plantarum SF9C and Lb. brevis SF9B was able to influence the intestinal microbiota composition in rats, which was reflected in the increased abundance of Lactobacillus genus, but also in the altered abundances of other bacterial genera, either in the model of healthy or aberrant gut microbiota of rats. The antibacterial activity and capacity to withstand in GIT conditions contributed to the functional aspects of SF9C and SF9B strains that could be incorporated in the probiotic-containing functional foods with a possibility to positively modulate the gut microbiota composition.
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