Critical water activity for the preservation of Lactobacillus bulgaricus by vacuum drying

Lactobacillus delbrueckii subsp. bulgaricus was dried under vacuum at different temperatures and its preservation evaluated analyzing the evolution of three parameters throughout the process: lag time, percentage of membrane damage and zeta potential. Microorganisms were dehydrated at 30, 45 and 70 degrees C in a vacuum centrifuge for different times. The aw achieved for each time of drying was correlated with the cell recovery at all the temperatures assayed. The recovery of microorganisms was evaluated by means of: a) kinetics of growth in milk after drying, as a measure of the global damage; b) quantification of the membrane damage using the fluorescent dyes SYTO 9 and PI; c) determination of changes in the superficial charges (zeta potential) as measured of the increase in the hydrophobic residues exposed in the bacterial surface after dehydration. These changes correlate well with the bacterial damage occurred during the dehydration process. The Page's equation allowed fitting of aw and time of drying, thus making possible the determination of the appropriate dehydration conditions (time-temperature ratios) for which no cell damage occurs. The evaluation of three parameters (lag time, percentage of membrane damage and zeta potential) allowed us to conclude that at the lowest temperature of dehydration, the first target of damage is the cell membrane. However, this damage is not decisive for the bacterial recovery after rehydration, as are the increase in the lag time and the changes in the zeta potential, as was observed for L. bulgaricus dehydrated at 45 and 70 degrees C for larger times.     

Electrostatic spraying of food-grade organic and inorganic acids and plant extracts to decontaminate Escherichia coli O157:H7 on spinach and iceberg lettuce

The prevalence of foodborne illnesses is continually on rise. In the U.S.A., Escherichia coli O157:H7 (E. coli) has been associated with several outbreaks in minimally processed foods. Spinach and lettuce pose higher food safety risks and recurring food recalls suggest the insufficiency of current disinfection strategies. We aimed at offering a natural antimicrobial alternative using organic acids (malic, tartaric, and lactic acids [MA, TA, and LA, respectively]) and grape seed extract (GSE) and a novel application method using electrostatic spraying to evenly distribute the antimicrobials onto produce. Spinach and lettuce samples were washed, sanitized with sodium hypochlorite solution (6.25 mL/L), dip inoculated in water containing E. coli (7.0 log CFU/mL) for 24 h, and rewashed with sterile water to remove nonadhered pathogens. The samples were sprayed electrostatically with MA, LA, and GSE alone and in combinations and for comparison, with phosphoric acid (PA) and pH controls with deionized water adjusted to 1.5/2.3/3.6 and stored at 4 °C. When combined with LA (3%), MA (3%) showed 2.1 to 4.0 log CFU/g reduction of E. coli between the days 1 and 14 on spinach and 1.1 to 2.5 log CFU/g reduction on lettuce. Treatment with PA (1.5%) and PA (1.5%)-GSE (2%) exhibited 1.1 to 2.1 log CFU/g inhibition of E. coli on spinach during the 14-d storage. Our findings demonstrated the efficacy of electrostatic spraying of MA, LA, and GSE on fresh produce to improve the safety and lower the public health burden linked to produce contamination. PRACTICAL APPLICATION: Electrostatic spraying is an emerging technique that can be adopted to improve the distribution and application of antimicrobials during fresh produce sanitation. Relatively simple and quick, the process can access most/all parts of produce surface and offer protection from food pathogens. The use of malic and lactic acids with or without grape seed extract can serve as effective antimicrobials when sprayed electrostatically, lowering the risk from postcontamination issues with spinach and iceberg lettuce. This application technology can be extended to improve the commercial food safety of other produce, fruits, poultry, and meat.     

Estimating postprandial glucose fluxes using hierarchical Bayes modelling

A new stochastic computational method was developed to estimate the endogenous glucose production, the meal-related glucose appearance rate (R(a meal)), and the glucose disposal (R(d)) during the meal tolerance test. A prior probability distribution was adopted which assumes smooth glucose fluxes with individualized smoothness level within the context of a Bayes hierarchical model. The new method was contrasted with the maximum likelihood method using data collected in 18 subjects with type 2 diabetes who ingested a mixed meal containing [U-(13)C]glucose. Primed [6,6-(2)H(2)]glucose was infused in a manner that mimicked the expected endogenous glucose production. The mean endogenous glucose production, R(a meal), and R(d) calculated by the new method and maximum likelihood method were nearly identical. However, the maximum likelihood gave constant, nonphysiological postprandial endogenous glucose production in two subjects whilst the new method gave plausible estimates of endogenous glucose production in all subjects. Additionally, the two methods were compared using a simulated triple-tracer experiment in 12 virtual subjects. The accuracy of the estimates of the endogenous glucose production and R(a meal) profiles was similar [root mean square error (RMSE) 1.0±0.3 vs. 1.4±0.7μmol/kg/min for EGP and 2.6±1.0 vs. 2.9±0.9μmol/kg/min for R(a meal); new method vs. maximum likelihood method; P=NS, paired t-test]. The accuracy of R(d) estimates was significantly increased by the new method (RMSE 5.3±1.9 vs. 4.2±1.3; new method vs. ML method; P<0.01, paired t-test). We conclude that the new method increases plausibility of the endogenous glucose production and improves accuracy of glucose disposal compared to the maximum likelihood method.     

Factors influencing the national distribution of polycyclic aromatic hydrocarbons and polychlorinated biphenyls in British soils

The polycyclic aromatic hydrocarbons (PAHs) naphthalene, acenaphthylene, acenaphthene, fluorene, fluoranthene, pyrene, benzo[a]anthracene, chrysene, benzo[b]fluoranthene, benzo[k]fluoranthene, benzo[a]pyrene, ideno[1,2,3,-cd]-pyrene, dibenz[a,h]anthracene, benzo[g,h,i]perylene and the polychlorinated biphenyls (PCBs) 8, 18, 28, 29, 31, 52, 77, 101, 105, 114, 118, 123, 126, 128, 138, 141, 149, 153, 156, 157, 163, 169, 170, 171, 180, 183, 187, 189, 194, 199, 201, 206, and 209 were measured in -200 rural soils across Great Britain (GB). Dominance of soil PAH profiles by heavier compounds (4-6 rings) provided initial evidence for the importance of source in governing soil PAH concentrations. No relationship was found between soil organic matter (SOM) and sum concentration of total and "heavy" PAHs, although there was a weak positive relationship with lighter compounds. A spatial statistical technique showed that highest soil PAH concentrations were usually found close to industrial/urban centers where presumably source intensity is highest. PCBs clustered into seven groups, five of which contained a single "dioxin like" PCB, one contained lighter congeners (2-4 chlorines), and one contained heavy congeners (5-10 chlorines). Linear regressions with SOM explained up to 24.3% of variation for the sum concentration of penta- to deca- congeners, but <1% for the lighter congener groups. No significant relationships were found with latitude. Spatial statistical techniques showed clusters of high soil PCB concentrations predominantly in west and south east GB, either associated with urbanized areas or on the West coast.     

Nitrogen budget for fescue pastures fertilized with broiler litter in Major Land Resource Areas of the southeastern US

The southeast US produces a tremendous number of broiler chickens (Gallus gallus), which in turn produce massive quantities of litter (manure and bedding materials). In the Southeast, litter is most often disposed of via land application to pastures, however, the ultimate fate of much of the applied nitrogen (N) is not known. We have constructed N budgets for three sites across the southeastern U.S. in an effort to determine how much of the applied N is useful for plant production and how much is left to be absorbed by the environment. Study sites were located in the Coastal Plain (Alabama), Piedmont (Georgia), and Cumberland Plateau (Tennessee) Major Land Resource Areas (MLRA) of the southeastern US. Litter was applied in the Spring of two consecutive years at a rate to supply 70 kg of available N ha^–1. The total amount of N applied ranged from 103 to 252 kg N ha^–1 depending on site and year. Nitrogen fluxes monitored in this study were broiler litter N, ammonia (NH₃) volatilization, denitrification, plant uptake, and leaching. Plant uptake represented the largest flux of applied N, averaging 43% of applied N. Losses due to NH₃ volatilization and denitrification combined were only 6% of applied N on average. Loss of N due to NO₃-N leaching appeared to be significant only at the Coastal Plain site where NO₃-N concentrations in the groundwater peaked at 38 mg N l^–1. We believe the majority of excess N shown in these budgets is likely accounted for by leaching losses and soil accumulation. Regardless of these assumptions and low gaseous losses, it is apparent that on average, 57% of applied N is destined for a fate other than plant uptake. The results of this study indicate that land-application of broiler litter at currently recommended rates has the potential for negative impacts on the environment of the southeastern U.S. in the long-term.     

Amphiphilic Pentablock Copolymers Prepared from Pluronic and ε-Caprolactone by Enzymatic Ring Opening Polymerization

Amphiphilic copolymers are appealing materials because of their interesting architecture and tunable properties. In view of their application in the biomedical field, the preparation of these materials should avoid the use of toxic compounds as catalysts. Therefore, enzymatic catalysis is a suitable alternative to common synthetic routes. Pentablock copolymers (CUC) were synthesized with high yields by ring-opening polymerization of ε-caprolactone (ε-CL) initiated by Pluronic (EPE) and catalyzed by Candida antarctica lipase B enzyme. The variables to study the structure–property relationship were EPEs’ molecular weight and molar ratios between ε-CL monomer and EPE macro-initiator (M/In). The obtained copolymers were chemically characterized, the molecular weight determined, and morphologies evaluated. The results suggest an interaction between the reaction time and M/In variables. There was a correlation between the differential scanning calorimetry data with those of X-ray diffraction (WAXD). The length of the central block of CUC copolymers may have an important role in the crystal formation. WAXD analyses indicated that a micro-phase separation takes place in all the prepared copolymers. Preliminary cytotoxicity experiments on the extracts of the polymer confirmed that these materials are nontoxic.     

Dual-Function Semaphorin 4D Released by Platelets: Suppression of Osteoblastogenesis and Promotion of Osteoclastogenesis

Effects of the antiosteoblastogenesis factor Semaphorin 4D (Sema4D), expressed by thrombin-activated platelets (TPs), on osteoblastogenesis, as well as osteoclastogenesis, were investigated in vitro. Intact platelets released both Sema4D and IGF-1. However, in response to stimulation with thrombin, platelets upregulated the release of Sema4D, but not IGF-1. Anti-Sema4D-neutralizing monoclonal antibody (mAb) upregulated TP-mediated osteoblastogenesis in MC3T3-E1 osteoblast precursors. MC3T3-E1 cells exposed to TPs induced phosphorylation of Akt and ERK further upregulated by the addition of anti-sema4D-mAb, suggesting the suppressive effects of TP-expressing Sema4D on osteoblastogenesis. On the other hand, TPs promoted RANKL-mediated osteoclastogenesis in the primary culture of bone-marrow-derived mononuclear cells (BMMCs). Among the known three receptors of Sema4D, including Plexin B1, Plexin B2 and CD72, little Plexin B2 was detected, and no Plexin B1 was detected, but a high level of CD72 mRNA was detected in RANKL-stimulated BMMCs by qPCR. Both anti-Sema4D-mAb and anti-CD72-mAb suppressed RANKL-induced osteoclast formation and bone resorptive activity, suggesting that Sema4D released by TPs promotes osteoclastogenesis via ligation to a CD72 receptor. This study demonstrated that Sema4D released by TPs suppresses osteogenic activity and promotes osteoclastogenesis, suggesting the novel property of platelets in bone-remodeling processes.     

Increased HDL Size and Enhanced Apo A-I Catabolic Rates Are Associated With Doxorubicin-Induced Proteinuria in New Zealand White Rabbits

The catabolism and structure of high‐density lipoproteins (HDL) may be the determining factor of their atheroprotective properties. To better understand the role of the kidney in HDL catabolism, here we characterized HDL subclasses and the catabolic rates of apo A‐I in a rabbit model of proteinuria. Proteinuria was induced by intravenous administration of doxorubicin in New Zealand white rabbits (n = 10). HDL size and HDL subclass lipids were assessed by electrophoresis of the isolated lipoproteins. The catabolic rate of HDL‐apo A‐I was evaluated by exogenous radiolabelling with iodine‐131. Doxorubicin induced significant proteinuria after 4 weeks (4.47 ± 0.55 vs. 0.30 ± 0.02 g/L of protein in urine, P < 0.001) associated with increased uremia, creatininemia, and cardiotoxicity. Large HDL2b augmented significantly during proteinuria, whereas small HDL3b and HDL3c decreased compared to basal conditions. HDL2b, HDL2a, and HDL3a subclasses were enriched with triacylglycerols in proteinuric animals as determined by the triacylglycerol‐to‐phospholipid ratio; the cholesterol content in HDL subclasses remained unchanged. The fractional catabolic rate (FCR) of [¹³¹I]‐apo A‐I in the proteinuric rabbits was faster (FCR = 0.036 h^?1) compared to control rabbits group (FCR = 0.026 h^?1, P < 0.05). Apo E increased and apo A‐I decreased in HDL, whereas PON‐1 activity increased in proteinuric rabbits. Proteinuria was associated with an increased number of large HDL2b particles and a decreased number of small HDL3b and 3c. Proteinuria was also connected to an alteration in HDL subclass lipids, apolipoprotein content of HDL, high paraoxonase‐1 activity, and a rise in the fractional catabolic rate of the [¹³¹I]‐apo A‐I.