Novel Bovine Plasma Protein Film Reinforced with Nanofibrillated Cellulose Fiber as Edible Food Packaging Material

Proteins, such as those in blood from slaughterhouses, are a good option for developing edible films. However, films made exclusively from proteins have low strength and high water solubility, which makes them difficult to use in the food industry. The use of cellulosic material, such as nanofibrillated cellulose (NFC), can improve the properties of these films. In the present work, bovine plasma was acidified and treated with ethanol to precipitate its proteins, and these proteins were used to prepare films reinforced with several concentrations of NFC. In addition, control films prepared with untreated bovine plasma and reinforced with NFC were prepared as well. These new edible films were characterized according to their mechanical properties, water vapor permeability, light transmittance, and microstructure. Furthermore, the film with the best properties was selected to be additivated with nisin to test its antimicrobial properties by wrapping meat previously contaminated with Staphylococcus aureus. In this sense, films prepared with the extracted proteins showed better properties than the films prepared with untreated plasma. In addition, the results showed that the reinforcement of the films with a 10% (w/w) of NFC decreased their water solubility and improved their puncture strength and water vapor barrier properties. Finally, the addition of nisin to the films prepared with extracted protein from bovine plasma and NFC gave them antimicrobial properties against S. aureus.

Establishment of a sandwich ELISA for bovine plasma PON-1 and its predictive capabilities for dairy cows with fatty liver

The aim of this study to improve the clinical diagnosis of fatty livers (FL) in dairy cows by using the paraoxonase-1 (PON-1) enzyme as a detection index. Prokaryotic expression technology was used to generate recombinant bovine PON-1 protein. Mice were immunized with this protein to generate hybridoma cells, stably secreting anti-PON-1. Cells were injected into the peritoneal cavity of mice, and ascites were purified to generate bovine PON-1 monoclonal antibody. Rabbits were then immunized with this antigen, and a polyclonal antibody against bovine PON-1 was obtained. Using monoclonal and polyclonal antibodies, a double-antibody sandwich ELISA for plasma PON-1 was constructed. Plasma samples were collected from healthy (n = 13) and FL (n = 13) cows, and plasma PON-1 levels were detected using the PON-1 ELISA. Receiver operating characteristic curve (ROC) analysis was used to analyze correlations between PON-1 levels and FL. Results showed that the ideal working concentration of the monoclonal antibody was 0.8 mg/mL, and the quantitative detection limit was 90 ng/mL. Plasma PON-1 levels were significantly lower in FL cows, when compared with healthy animals. It is concluded that PON-1 ELISA predicts risk factors for dairy cows with FL. PON-1 levels in plasma can be used as an early warning indicator for FL and concentration of 61.87 nmol/L was identified as warning index.

Analysis of Circulating microRNA During Early Gestation in Japanese Black Cattle

Background Circulating microRNAs (miRNAs) are biomarkers for various diseases and physiological conditions in humans and mice; studies in domestic animals, particularly cattle, are limited. The importance of early pregnancy diagnosis (especially within the 21 day cow estrous cycle) in the livestock industry is extremely high. This study compared the circulating miRNAs in non-pregnant and pregnant Japanese Black cows, explored miRNAs as biomarkers for early pregnancy diagnosis, and established a measurement system that includes reference miRNA selection and the effect of hemolysis. Methods miRNA was extracted from the plasma of Japanese Black cows on day 21 after artificial insemination and subjected to a customized bovine oligonucleotide microarray for expression analysis. Differentially expressed miRNAs and reference miRNA candidates were selected and validated by real-time quantification PCR (RT-qPCR). Their stability was evaluated using NormFinder software. Hemolyzed samples were prepared using plasma from five cows in estrous cycle and subjected to RT-qPCR. Results A total of 124 miRNAs were detected in bovine plasma by microarray analysis in non-pregnant and pregnant cows. The levels of five circulating miRNAs were significantly higher in pregnant cows than in non-pregnant cows, and 24 miRNAs were detected only in the pregnant group. NormFinder analysis and RT-qPCR validation showed that miR-2455 was an appropriate reference miRNA in the plasma of non-pregnant and pregnant Japanese Black cows and miR-19b, miR-25, miR-29a, and miR-148a were significantly higher in the pregnant group. These four circulating miRNAs did not change during the estrous cycle and were less affected by hemolysis. Conclusions In the current study, we found high levels of four miRNAs in the plasma of pregnant Japanese Black cows. Since these miRNAs are less affected by hemolysis, they may potentially be used as markers for early pregnancy diagnosis in cattle.

Scaling up the Enzymatic Hydrolysis of Bovine Plasma Protein to Produce an Antioxidant from a Biological Source

Background: modern society, there is a tendency to consume products with natural origins and minimum chemical additives. This has encouraged the replacement of synthetic antioxidants for the ones obtained from natural sources, such as the antioxidants acquired from enzymatic protein hydrolysates. Objective: In this study, the process of enzymatic hydrolysis of proteins from bovine plasma, which produces hydrolysates with an Antioxidant Capacity (AC), was scaled up from 1 to 5 L. Methods: An experimental design was developed in 1 L to evaluate the effect of the Substrate concentration (So) on the time needed to reach a Degree of Hydrolysis (DH) of 20% as well as the AC. Results: The best conditions in the 1 L reactor controlled by a Titrando 842 were transferred to 5L in a BioFlo310 reactor. These conditions were achieved at a ratio of 80g/L of the substrate and 0.89 AU of Alcalase 2.4L/g of the substrate in order to obtain a level of 16.36 ± 0.21min of the 20% of DH and antioxidant capacity of 58.98 ± 1.80%. Conclusion: The results showed that DH depends significantly on So, while the antioxidant capacity only depends on the DH. Additionally, the dimensional analysis using Re as a scaling criterion allowed us to obtain the same results in the model (1 L) and the prototype (5 L).

In Vitro Immunological Cross-Reactivity of Thai Polyvalent and Monovalent Antivenoms with Asian Viper Venoms

The intravenous administration of polyclonal antibodies known as antivenom is the only effective treatment for snakebite envenomed victims, but because of inter-specific variation in the toxic components of snake venoms, these therapies have variable efficacies against different snake species and/or different populations of the same species. In this study, we sought to characterize the in vitro venom binding capability and in vitro cross-neutralizing activity of antivenom, specifically the Hemato Polyvalent antivenom (HPAV; The Queen Saovabha Memorial Institute (QSMI) of the Thai Red Cross Society, Thailand) and three monovalent antivenoms (QSMI) specific to Daboia siamensis, Calloselasma rhodostoma, and Trimeresurus albolabris venoms, against a variety of South Asian and Southeast Asian viper venoms (Calloselasma rhodostoma, Daboia russelii, Hypnale hypnale, Trimeresurus albolabris, Trimeresurus purpureomaculatus, Trimeresurus hageni, and Trimeresurus fucatus). Using ELISA and immunoblotting approaches, we find that the majority of protein components in the viper venoms were recognized and bound by the HPAV polyvalent antivenom, while the monospecific antivenom made against T.albolabris extensively recognized toxins present in the venom of related species, T. purpureomaculatus, T. hageni, and T. fucatus. In vitro coagulation assays using bovine plasma revealed similar findings, with HPAV antivenom significantly inhibiting the coagulopathic activities of all tested viper venoms and T. albolabris antivenom inhibiting the venoms from Malaysian arboreal pit vipers. We also show that the monovalent C. rhodostoma antivenom exhibits highly comparable levels of immunological binding and in vitro venom neutralization to venom from both Thailand and Malaysia, despite previous reports of considerable intraspecific venom variation. Our findings suggest that Thai antivenoms from QSMI may by useful therapeutics for managing snake envenomings caused by a number of Southeast Asian viper species and populations for which no specific antivenom currently exists and thus should be explored further to assess their clinical utility in treating snakebite victims.

172 Increasing inclusion of fermentable fiber decreases nutrient digestibility in grower pigs

Fermentable fiber may increase endogenous losses of P and AA, thereby reducing apparent nutrient digestibility. Acacia gum fiber with medium-to-high fermentability and low viscosity was used to investigate its effect on apparent ileal digestibility (AID) and apparent total tract digestibility (ATTD) of nutrients, and standardized total tract digestibility (STTD) of P in grower pigs. A P-free basal diet (49% corn starch; 18% bovine plasma protein) was formulated to measure basal endogenous P losses (EPL). Three diets were formulated to include 2.5, 5.0, or 7.5% acacia gum fiber at the expense of corn starch in the P-free basal diet. Diets contained 16.1–17.4% CP and 0.31–0.33% total P (DM-basis). The 4 diets were fed to 8 ileal-cannulated barrows (initial BW, 54.6 kg) for four 9-d periods in a double 4 × 4 Latin square. Apparent hindgut fermentation (AHF) was ATTD minus AID. Increasing inclusion of acacia gum quadratically decreased (P < 0.01) AID of DM (∆ 11.1%), linearly decreased (P < 0.05) ATTD of DM (∆ 1.7%) CP (∆ 1.2%), and quadratically increased (P < 0.05) AHF of DM (∆ 9.4%). Basal EPL were 391 and 377 mg/kg DM intake (DMI) for ileum and total tract, respectively. Increasing inclusion of fiber linearly increased (P < 0.05) ileal EPL (∆ 184 mg/kg DMI), and tended to linearly increase (P < 0.10) EPL for total tract (∆ 243 mg/kg DMI). Dietary inclusion of acacia gum tended to linearly decrease (P < 0.10) AID of P, but did not affect (P > 0.10) ATTD, or STTD of P. In conclusion, increasing inclusion of fermentable, low viscous acacia gum fiber decreased diet digestibility of DM and ATTD of CP, but did not affect total tract P digestibility, indicating that increasing fermentable fiber did not increase specific endogenous losses of P in the total tract.