Unstructured kinetic model for reuterin and 1,3‐propanediol production by Lactobacillus reuteri from glycerol/glucose cofermentation

BACKGROUND: Lactobacillus reuteri is unable to grow on glycerol as sole carbon and energy source hence, glycerol is used as an alternative hydrogen acceptor during growth on available carbohydrates. Thus, glycerol is converted to reuterin and 1,3‐propanediol (1,3‐PDL), both products with interesting industrial applications. These compounds are commonly produced by using resting cells in two‐step fermentation processes. RESULTS: The glycerol/glucose cofermentation by L. reuteri yields reuterin and 1,3‐PDL at a glycerol concentration higher than 100 mmol L⁻¹. An increase of glycerol concentration from 200 to 400 mmol L⁻¹ showed no additional stimulatory effect on ethanol and acetate production but consistently reduced the lactate concentration. It was also found that reuterin concentration reached a maximum value and subsequently decreased due to its conversion to 1,3‐PDL. An unstructured kinetic model was proposed to describe simultaneously microbial growth, substrates consumption and products formation. A multi‐response nonlinear regression analysis based on Marquardt algorithm combined with a Runge‐Kutta integration method was used to obtain the values of the fitting parameters. CONCLUSIONS: The optimum concentration of glycerol for maximum reuterin and 1,3‐PDL production was 200 mmol L⁻¹. The complete process was satisfactorily described by the kinetic model proposed. Copyright © 2008 Society of Chemical Industry     

Antimicrobial activity of nisin, reuterin, and the lactoperoxidase system on Listeria monocytogenes and Staphylococcus aureus in cuajada, a semisolid dairy product manufactured in Spain

The inhibitory activity of nisin (N), reuterin (R), and the lactoperoxidase system (LPS), added individually or in combination, against Listeria monocytogenes and Staphylococcus aureus was investigated in “cuajada” (curdled milk), a semisolid dairy product manufactured in Spain. Cuajada was manufactured from UHT skim milk separately inoculated with L. monocytogenes and Staph. aureus, each at approximately 4 log cfu/mL, and held under conditions of temperature abuse (10°C). On d 3, a synergistic bactericidal activity was observed for the combinations of biopreservatives assayed, with L. monocytogenes counts of only 0.30 log cfu/mL in cuajada made with N + R + LPS vs. 8.31 log cfu/mL in control cuajada. After 12 d, L. monocytogenes could not be detected in cuajada made with added N + LPS or N + R + LPS. Staphylococcus aureus was more resistant than L. monocytogenes to biopreservatives added individually. On d 3, the synergistic effect of the 3 biopreservatives against Staph. aureus resulted in counts of 3.03 log cfu/mL in cuajada made with N + R + LPS vs. 6.40 in control cuajada. After 12 d, Staph. aureus counts were 2.61 log cfu/mL in cuajada made with N + R + LPS, whereas they ranged from 6.11 to 7.70 log cfu/mL in control cuajada and in cuajada made with other combinations of biopreservatives. The most pronounced decrease in pathogen counts was achieved by the triple combination N + R + LPS, which acted synergistically on the inactivation of L. monocytogenes and Staph. aureus in cuajada over 12 d at 10°C. The treatment combining these 3 natural biopreservatives at low concentrations, within the hurdle concept of food preservation, might be a useful tool to control the growth of pathogenic microorganisms in nonacidified dairy products.     

Efficacy of bacteriocin-based formula for reducing staphylococci,streptococci, and total bacterial counts on teat skin of dairy cows

The use of teat dips is one of the most effective strategies to control mastitis by preventing new intramammary infections. Reducing bacterial load on teat skin helps control the spread of pathogens and spoilage and improves the quality of milk. The objective of this study was to evaluate the reduction of bacterial populations through the application of bacteriocin-based teat formulas. Teats of 12 Holstein cows received 2 different concentrations of bactofencin A, nisin, and reuterin alone or in combination, as well as iodine (positive control) and saline (negative control). Teat swabs were collected before and after application of teat formulas and analyzed for staphylococci, streptococci, and total bacteria counts. There were no differences for staphylococci, streptococci, and total bacterial counts for samples collected before application throughout the entire experiment. Reuterin-low and reuterin-high treatments reduced total bacterial count by 0.47 and 0.36 logs, respectively, whereas bactofencin A had no effect on any tested bacterial groups. Nisin-low treatment reduced staphylococci, streptococci, and total bacterial counts by 0.47, 0.30 and 0.50 logs, respectively. Nisin-high treatment resulted in 0.50, 0.50, and 0.47 log reduction for staphylococci, streptococci, and total bacterial counts. The bacteriocin consortium showed the highest reduction rates with 0.91, 0.54, and 0.90 log reductions obtained for staphylococci, streptococci, and total bacteria counts, respectively, for the low-concentration consortium. Similarly, the high-concentration consortium showed reduction rates with 0.95, 0.60, and 0.82 log reductions obtained for staphylococci, streptococci, and total bacteria counts, respectively. Thus, nisin and the bacteriocin consortium showed the most promise as a teat disinfectant by reducing staphylococci, streptococci, and total bacteria counts.     

Microencapsulated Lactobacillus reuteri combined with modified atmosphere as a way to improve tuna burger shelf life

In this study, tuna burgers were subjected to a bio‐preservation treatment combined with modified atmosphere packaging (MAP) in order to prolong the shelf life. After L. reuteri production of reuterin, the antimicrobial compound was microencapsulated and then added to tuna burger formulation. Reuterin content was analysed before and after the spray drying treatment, thus assessing its final concentration. Then, the effects of spray‐dried reuterin on tuna burgers quality were investigated with success on samples packaged under both ordinary and MAP conditions. The efficacy of bio‐control was evident in all the samples. For burgers stored in air, a shelf life of about 7.63 days was recorded for the active samples with respect to the 5.78 days of the reuterin‐free samples. Under MAP, a similar reuterin effectiveness was recorded but a general shelf life improvement was reached, accounting for about 13 days in the active samples and 12 days in the reuterin‐free samples.     

Reuterin production by Lactobacillus reuteri NRRL B‐14171 immobilized in alginate

BACKGROUND: Lactobacillus reuteri NRRL B‐14171 was immobilized in spherical calcium alginate beads to study reuterin production. RESULTS: In relation to free cells (82 mmol L^?1), the immobilized cells showed a sharp increase in reaction rate and achieved similar conversions with a reduction in the required biomass. Tests for mass transfer limitations on the reaction rate indicated that neither intraparticle nor external constraints were present. Effectiveness factors near 1 were estimated. CONCLUSION: Immobilization induced increased production of reuterin in Lactobacillus reuteri and is a feasible option for increasing process productivity. Copyright © 2008 Society of Chemical Industry     

静息细胞生物转化累积罗伊氏素影响条件的研究(英文)

3-Hydroxypropionaldehyde (3-HPA) is a potential valuable chemical and new broad-spectrum antim-icrobial substance. In order to improve the conversion of 3-HPA/glycerol, our work studied the two-step process from glycerol to 3-HPA, and investigated the influence of cell harvest time, glycerol concentration, biomass con-centration, pH and temperature on the production of 3-HPA by Lactobacillus reuteri CG001, respectively. The re-sults showed that molar conversion yield of 3-HPA/glycerol reached 97.9% under the condition that 200 mmol·L-1 glycerol was converted by 25.3 g·L-1 resting cell for 1 h at 30 ℃. The cells could not be reused directly because the L. reuteri almost lost its bioconversion activity completely, but the ability of glycerol conversion could gradually recover if the fresh medium was added to the deactivated cell for 4 h.     

Short communication: Combined antimicrobial activity of reuterin and diacetyl against foodborne pathogens

Reuterin (β-hydroxypropionialdehyde) is a broad-spectrum antimicrobial substance produced by some strains of Lactobacillus reuteri during anaerobic fermentation of glycerol. Some of these strains are able to survive and produce reuterin in cheese and yogurt when added as adjuncts to the starter. Similarly, in fermented dairy foods, other inhibitory compounds such as lactic acid and diacetyl are produced during fermentation. In this work, we studied the combined effect of reuterin and diacetyl under different pH conditions against Escherichia coli O157:H7, Salmonella Enteritidis, and Listeria monocytogenes. Results from agar spot assays showed that the antimicrobial activity of reuterin-producing strains against the gram-negative bacteria tested was enhanced as the concentration of diacetyl increased to 50 mg/kg, and was higher under acidic conditions (pH 5.0) for the 3 pathogenic strains. The combination of reuterin and diacetyl had an additive effect against L. monocytogenes only at diacetyl concentrations of 50 mg/kg and pH 5.0. In addition, growth kinetics studies showed that the combination of 1 activity unit (AU)/mL of reuterin with 100 mg/kg diacetyl increased the lag time of the 3 pathogens. In milk, synergistic antimicrobial activity was observed with the combination of 1 AU/mL reuterin and 50 or 100 mg/kg of diacetyl on the gram-negative strains tested, and with 1 AU/mL reuterin and 100 mg/kg of diacetyl on L. monocytogenes. The greatest inhibition of the 3 pathogens was achieved in acidified milk at pH 5.0 with reuterin (1 AU/mL) and diacetyl (100 mg/kg). Based on these results, the combination of reuterin and diacetyl in acidified dairy products could be a promising strategy to control food pathogens in these products.     

Production of reuterin in a fermented milk product by Lactobacillus reuteri: Inhibition of pathogens,spoilage microorganisms,and lactic acid bacteria

We assessed the antimicrobial activity of reuterin produced in vitro in glycerol aqueous solutions in situ by Lactobacillus reuteri ATCC 53608 as part of a fermented milk product against starter (Lactobacillus delbrueckii ssp. bulgaricus and Streptococcus thermophilus), spoilage (Penicillium expansum), pathogenic (Staphylococcus aureus Salmonella enterica ssp. enterica, and Listeria monocytogenes), and pathogen surrogate (Escherichia coli DH5α) microorganisms. We also assayed the influence of cold storage (28 d at 4°C) and reuterin on the color and rheology of the fermented milk product. We obtained maximum reuterin concentrations of 107.5 and 33.97 mM in glycerol aqueous solution and fermented milk product, respectively. Reuterin was stable throughout its refrigerated shelf life. Gram-positive microorganisms were more resistant to reuterin than gram-negative microorganisms. Penicillium expansum and Lactobacillus reuteri ATCC 53608 survived at concentrations up to 10 and 8.5 mM, respectively. Escherichia coli DH5α was the most sensitive to reuterin (0.9 mM). The presence of reuterin did not cause relevant changes in the quality parameters of the fermented milk product, including pH, acidity, soluble solids, color, and rheological aspects (storage and loss moduli and viscosity). This study demonstrated the viability of using Lactobacillus reuteri ATCC 53608 as a biopreservative in a fermented milk product through reuterin synthesis, without drastically modifying its quality parameters.     

产Reuterin罗伊氏乳杆菌的定向筛选及其生物学特性

依据菌株宿主适应性特点，从健康家禽新鲜粪便中定向筛选具有开发潜力的罗伊氏乳杆菌。经16S rDNA序列分析和pduC基因筛查，获得3 株产抗菌物质Reuterin的罗伊氏乳杆菌，对其耐受模拟胃肠道环境、抗生素敏感性、甘油发酵上清液抑菌活性及肠上皮细胞黏附能力等生物学特性进行系统评价。结果表明，罗伊氏乳杆菌WLRE01、WLRE03、WLRE04均具有较强耐受极端酸（pH 2.5和pH 3.5）、胆盐（1.5 g/L）及模拟胃肠液的能力；经1 mmol/L过氧化氢处理6 h，WLRE03、WLRE04的活菌数能基本维持初始浓度；甘油发酵上清液对金黄色葡萄球菌具有极强的抑制作用，抑菌圈最高达（28.1±1.2）mm；3 株菌对HT-29细胞均具有良好的黏附特性，其中WLRE01黏附能力最强。综上所述，健康家禽肠道源的3 株产Reuterin的罗伊氏乳杆菌可作为益生菌候选菌株进行深入研究。