Amino Acid Sequences of Lactoferrin from Red Deer (Cervus elaphus) Milk and Antimicrobial Activity of Its Derived Peptides Lactoferricin and Lactoferrampin.

Although the bioactivities of bovine lactoferrin have been extensively investigated, little is known about deer milk lactoferrin bioactivity and its amino acid sequence. This research investigated the amino acid sequence of deer lactoferrin and the antimicrobial activities of two lactoferrin-encrypted peptides; lactoferricin (Lfcin) and lactoferrampin (Lfampin). Deer lactoferrin was found to have a molecular weight of 77.1 kDa and an isoelectric point of 7.99, which are similar to that of bovine lactoferrin, 78 kDa and pI 7.9. Deer lactoferrin contains 707 amino acids, one amino acid less than bovine lactoferrin, and has 92% homology with bovine lactoferrin. Deer lactoferricin exhibited strong antimicrobial activity against E. coli American Type Culture Collection (ATCC) 25922 and L. acidophilus ATCC 4356. The antimicrobial activities of deer and bovine Lfcin and Lfampin were compared. Based on MIC, deer Lfcin was found to be a more effective inhibitor of L. acidophilus ATCC 4356 than bovine Lfcin, but bovine Lfcin and Lfampin were more effective against E. coli ATCC 25922 than deer Lfcin and Lfampin. The deer Lfcin sequence differed at seven amino acids from bovine Lfcin and this decreased the net positive charge and increased the hydrophobicity. Deer Lfampin contained two differences in amino acid sequence compared to bovine Lfampin which decreased the net positive charge. These amino acid sequence differences likely account for differences in antibacterial activity. Positive charge and hydrophobic residues provide the amphipathic character of these helical peptides, and are considered important for binding of antimicrobial peptides. In silico modelling of deer Lfcin indicated an identical α-helical structure compared to bovine Lfcin.

Lactoferrin Isolation and Hydrolysis from Red Deer (Cervus elaphus) Milk and the Antibacterial Activity of Deer Lactoferrin and Its Hydrolysates.

Lactoferrin (Lf) and other whey proteins have been isolated from red deer milk for the first time using a three-step anion and cation exchange chromatography protocol. The separated deer Lf was subject to in vitro gastric and duodenal digestions to generate peptides. The purity of the deer Lf and its hydrolysis products were analyzed by SDS-PAGE. The antibacterial activity of the deer Lf and its hydrolysates were investigated and was compared to cow counterpart. Gastric and duodenal digested deer Lf had strong bactericidal activity against E. coli ATCC 25922 with minimum inhibition concentration (MIC) of 280 µM and 402 µM, respectively. These results suggest that deer milk contains bioactive whey proteins and can generate bioactive peptides, which can benefit human health by inhibiting food-borne pathogenic bacteria.

Sous-vide cooking improves the quality and in-vitro digestibility of Semitendinosus from culled dairy cows.

Effect of sous-vide processing was evaluated on in-vitro simulated protein digestion, myofibrillar protein profile, lipid oxidation and physicochemical properties of cold-boned beef Semitendinosus. Sous-vide processed [T1, 60 °C (4.5 h); T2, 60 °C (10 h)] samples along with control beef (cooked at 80 °C until a core temperature of 75 °C was attained) were subjected to in-vitro simulated gastrointestinal protein digestion. Samples collected at 0, 30 and 60 min of gastric digestion and 120 and 180 min of intestinal digestion were analysed for protein profile (SDS-PAGE), protein digestibility (%), soluble protein (%), free amino acid analysis and mineral profile. A significant (P < 0.05) decrease was observed in shear force (N) and cooking loss (%) of sous-vide processed samples in comparison to control whereas a significant (P < 0.05) increase was observed in colour (L*, a*, b*). A significant (P < 0.05) increase was also observed in protein digestibility (%), soluble protein (%) and release of free amino acids and minerals from sous-vide processed samples during in-vitro digestion. By modifying the protein profile of the meat, sous-vide processing had a positive influence on in-vitro digestion kinetics that led to a greater and faster digestion of proteins during simulated digestion.

The application of pulsed electric field as a sodium reducing strategy for meat products.

This study investigated the potential of pulsed electric field (PEF) as a sodium-reduction strategy for processed meat. Beef jerky was used as a model and prepared using different levels of NaCl, viz. 2.0% (control), 1.2% (T1) and 1.2% along with PEF processing (T2). A significant (p < 0.05) effect of PEF was observed on shear force (N) and toughness (N/mm s) of the products, which was also reflected in sensory scores. No effects for PEF were observed on colour, yield (%) and oxidative and microbial stability. PEF-treated samples (T2) had significantly (p < 0.05) lower sodium content than the control, however, the sensory scores were comparable (p > 0.05) with control and >84% of the panellists preferred T2 samples over T1 for saltiness. Results suggest that PEF treatment improved saltiness by influencing the salt diffusion and sodium delivery that led to better perception during chewing. PEF could be a novel method to produce healthier reduced-sodium meat products.

The Potential of Modulating the Reducing Sugar Released (and the Potential Glycemic Response) of Muffins Using a Combination of a Stevia Sweetener and Cocoa Powder.

Muffins are popular bakery products. However, they generally contain high amounts of sugar. The over-consumption of muffins may therefore result in a high calorie intake and could lead to increased health risks. For this reason, muffins were prepared substituting sucrose with two levels of a base of stevia (Stevianna®). In addition, cocoa powder and vanilla were added to the muffin formulation with and without Stevianna® to mask any potential off flavors. Results illustrate that muffins with 50% Stevianna® replacement of sucrose were similar to the control samples in terms of volume, density and texture. However, replacement of sugar with 100% Stevianna® resulted in reductions in height (from 41 to 28 mm), volume (from 63 to 51 mL), and increased firmness (by four-fold) compared to the control sample. Sugar replacement significantly reduced the in vitro predictive glycemic response of muffins (by up to 55% of the control sample). This work illustrates the importance of sugar in maintaining muffin structure as well as controlling the rate of glucose release during simulated digestions.

Viability assessment of Bifidobacterium longum ATCC 15707 on non-dairy foods using quantitative fluorescence microscopy.

This study demonstrates an effective technique for separating and purifying viable bacteria from samples that interfere with viability staining. The viability of Bifidobacterium longum ATCC 15707 was assessed using Percoll Buoyant Density Gradient Centrifugation (PBDC) to separate bacteria from complex non-dairy food matrices and Quantitative Fluorescence Microscopy (QFM) to determine individual cells using LIVE/DEAD BacLight bacterial viability staining. Water agar (3%) was used to retain cells of B. longum and offered a lower fluorescence background with BacLight viability staining, compared with fixation on polycarbonate (PC) black membrane. The effect of drying temperatures and non-dairy foods on viability of B. longum was assessed. B. longum coated on oat, peanut or raisin was separated by filtration, low- and high-speed centrifugation, flotation and sedimentation buoyant density centrifugation. Purified cells were subsequently deposited on water agar for rehydration followed by LIVE/DEAD BacLight viability staining and enumeration. Conventional plate counting was also conducted to compare viability results. Finally, this method was applied to assess cell membrane damages of B. longum incorporated onto non-dairy foods during 24 h drying. Furthermore, viability assessment of B. longum coated onto oat, peanut, or raisin was much lower by plate counting compared to viability staining. Drying appeared to have a greater impact when viability was assessed by plate counting compared to viability staining. IMPORTANCE: Enumeration of viable beneficial bacteria from function foods presents a significant bottleneck for product development and quality control. Interference with microscopic and/or fluorescent techniques by ingredients, time required to incubate plated microbes, and the transient nature of the colony forming unit make rapid assessment of viable bacteria difficult. Viability assessment of Bifidobacterium longum ATCC 15707 by Percoll Buoyant Density Gradient Centrifugation with LIVE/DEAD BacLight viability staining on water agar (3%) was in agreement with serial dilution enumeration. Without the need for incubation viability assessment by staining provided a more rapid means to assess the impact of drying on the viability of B. longum coated onto oat, peanut or raisin.

Pulsed electric field: A new way to improve digestibility of cooked beef.

The effect of pulsed electric field (PEF) on in-vitro simulated gastrointestinal protein digestion of cooked beef Semimembranosus was elucidated. PEF treated (T1, 10 kV, 20 Hz, 20 µs) samples along with control were cooked (core temperature of 75 °C) and subjected to in-vitro simulated gastrointestinal digestion. Samples were analysed for protein digestibility (%), protein profile (SDS-PAGE), soluble protein (%), free amino acid analysis and mineral profile. PEF treatment led to significantly higher (P < .05) values for protein digestibility (%) and soluble protein (%). No significant (P > .05) effect of PEF was recorded on the release of free amino acids and various minerals during gastrointestinal digestion. PEF treatment modified the protein profile of the meat digests and had a positive impact on in-vitro digestion kinetics causing greater and faster digestion of proteins during in-vitro gastrointestinal simulation.

Pulsed electric field operates enzymatically by causing early activation of calpains in beef during ageing.

A study was conducted to elucidate the impact of pulsed electric field (PEF) on the activity of calpains, proteolytic activity of desmin and troponin-T and physicochemical properties of beef Biceps femoris during ageing. The meat samples (meat blocks) were subjected to two PEF treatments; T1 (5 kV, 90 Hz, 0.38 kV/cm) and T2 (10 kV, 20 Hz, 0.61 kV/cm) and a non-treated control was run in parallel. The samples were vacuum packaged and aged for 1, 7 and 14 days at 4 ±â€¯1 °C. This study reports for the first time the impact of PEF-processing on the calpain activity in beef. Early post-mortem activation of calpain 2 was observed in PEF-treated samples. An increase in the calpain activity and proteolysis of desmin and troponin-T was observed. No significant effect of PEF was observed on the shear force of tough muscles from culled dairy animals during the entire ageing period.

Pulsed electric field: Effect on in-vitro simulated gastrointestinal protein digestion of deer Longissimus dorsi.

The effect of pulsed electric field (PEF) on in-vitro simulated gastrointestinal protein digestion of cold-boned deer Longissimus dorsi was elucidated. PEF treated samples viz. T1 (2.5 kV, 50 Hz) and T2 (10 kV, 90 Hz) along with a control were subjected to in-vitro simulated gastrointestinal protein digestion. Samples were collected after 0, 30, and 60 min of gastric digestion and 120 and 180 min of intestinal digestion. A significant (P < 0.05) increase was observed in the protein digestibility and soluble protein (%) of PEF treated samples, highlighting a positive influence of PEF processing on the digestion process. Higher concentrations of almost all the free amino acids were observed for the PEF treated samples. No significant effect of PEF was observed on the release of various minerals. PEF may be utilized for the development of novel muscle foods with improved protein digestibility.

Non-dairy probiotic food products: An emerging group of functional foods.

The functional food sector has shown tremendous growth in recent years with the application of probiotic bacteria as "food additives". The utilization of probiotic bacteria in food presents many challenges related to their growth, survival, viability, stability and functionality in food processing, storage and consumption as well as changes of sensory characteristics of probiotic foods. Although dairy foods are currently the most common food carrier to deliver probiotics, an increasing number of non-dairy food matrices exhibit potential for delivery of probiotics. This review provides more recent insight into the emergence of non-dairy probiotics products, the interactions between probiotics and different food matrices and the challenges in developing such products. Some of the technical issues are also reviewed and discussed. These issues include the efficacy of probiotic bacteria in non-chilled, low pH or high water activity foods; the potential loss of bacterial viability, additionally unwanted fermentation and changes of the sensory characteristics of food products which may result in poor microbiological quality and low acceptability to consumers.

Current and future prospects for the use of pulsed electric field in the meat industry.

Pulsed electric field (PEF) is a novel non-thermal technology that has recently attracted the attention of meat scientists and technologists due to its ability to modify membrane structure and enhance mass transfer. Several studies have confirmed the potential of pulsed electric field for improving meat tenderness in both pre-rigor and post-rigor muscles during aging. However, there is a high degree of variability between studies and the underlying mechanisms are not clearly understood. While some studies have suggested physical disruption as the main cause of PEF induced tenderness, enzymatic nature of the tenderization seems to be the most plausible mechanism. Several studies have suggested the potential of PEF to mediate the tenderization process due to its membrane altering properties causing early release of calcium ions and early activation of the calpain proteases. However, experimental research is yet to confirm this postulation. Recent studies have also reported increased post-mortem proteolysis in PEF treated muscles during aging. PEF has also been reported to accelerate curing, enhance drying and reduce the numbers of both pathogens and spoilage organisms in meat, although that demands intense processing conditions. While tenderization, meat safety and accelerated curing appears to be the areas where PEF could provide attractive options in meat processing, further research is required before the application of PEF becomes a commercial reality in the meat industry. It needs to deal with carcasses which vary biochemically and in composition (muscle, fat, and bones). This review critically evaluates the published reports on the topic with the aim of reaching a clear understanding of the possible applications of PEF in the meat sector in addition to providing some insight on critical issues that need to be addressed for the technology to be a practical option for the meat industry.

Technological, Regulatory, and Ethical Aspects of In Vitro Meat: A Future Slaughter-Free Harvest.

Defined as meat cultured in a laboratory within a bioreactor under controlled artificial conditions, in vitro meat is a relatively recent area that has opened a whole universe of possibilities and opportunities for the meat sector. With improved chemical and microbial safety and varied options, in vitro meat has been proposed as a green, healthy, environmentally friendly, and nutritionally better product that is free from animal suffering and death. Cell culture and tissue culture are the most probable technologies for the development of this futuristic muscle product. However, there are many challenges in the production of a suitable product at an industrial scale under a sustainable production system and a great body of research is required to fill the gaps in our knowledge. Many materials used in the product development are novel and untested within the food industry and demand urgent regulatory and safety assessment systems capable of managing any risks associated with the development of cultured meat. The future of this product will depend on the actions of governments and regulatory agencies. This article highlights emerging biotechnological options for the development of cultured meat and suggests ways to integrate these emerging technologies into meat research. It considers the problems and possibilities of developing cultured meat, opportunities, ethical issues as well as emerging safety and regulatory issues in this area.

Applied and Emerging Methods for Meat Tenderization: A Comparative Perspective.

The tenderization process, which can be influenced by both pre- and post-slaughter interventions, begins immediately after an animal's death and is followed with the disruption of the muscle structure by endogenous proteolytic systems. The post-slaughter technological interventions like electrical stimulation, suspension methods, blade tenderization, tumbling, use of exogenous enzymes, and traditional aging are some of the methods currently employed by the meat industry for improving tenderness. Over the time, technological advancement resulted in development of several novel methods, for maximizing the tenderness, which are being projected as quick, economical, nonthermal, green, and energy-efficient technologies. Comparison of these advanced technological methods with the current applied industrial methods is necessary to understand the feasibility and benefits of the novel technology. This review discusses the benefits and advantages of different emerging tenderization techniques such as hydrodynamic-pressure processing, high-pressure processing, pulsed electric field, ultrasound, SmartStretch™ , Pi-Vac Elasto-Pack® system, and some of the current applied methods used in the meat industry.

The Effect of Astaxanthin-Rich Microalgae "Haematococcus pluvialis" and Wholemeal Flours Incorporation in Improving the Physical and Functional Properties of Cookies.

Marine-based food supplements can improve human nutrition. In an effort to modulate glycaemic response and enhance nutritional aspects, marine-derived algal food rich in astaxanthin was used in the formulation of a model food (wholemeal cookie). Astaxanthin substitution of cookies made from three flours (wheat, barley and oat) demonstrated a significant reduction in the rate of glucose released during in vitro digestion together with an increase in the total phenolic content (TPC) and antioxidant capacity of the food. The significantly (p < 0.005) lower free glucose release was observed from cookies with 15% astaxanthin, followed by 10% and then 5% astaxanthin in comparison with control cookies of each flour. Total phenolic content, DPPH radical scavenging and Oxygen Radical Absorbance Capacity (ORAC) value also notably increased with increase in astaxanthin content. The results evidence the potential use of microalgae to enhance the bioactive compounds and lower the glycaemic response of wholemeal flour cookie.

Effect of Non-Dairy Food Matrices on the Survival of Probiotic Bacteria during Storage.

The viability of probiotics in non-dairy food products during storage is required to meet content criteria for probiotic products. This study investigated whether non-dairy foods could be matrices for probiotics. Selected probiotic bacteria were coated on non-dairy foods under two storage conditions, and viabilities were assessed. The non-dairy foods were coated with 5-7 log cfu g-1 of Lactobacillus acidophilus ATCC4356T, Lactobacillus plantarum RC30, and Bifidobacterium longum ATCC15707T. The coated non-dairy foods were stored at 20 °C and 20% relative humidity (RH) or 30 °C and 50% RH. Viability of probiotic bacteria was determined after 0, 2, and 4 weeks of storage. B. longum showed the highest survival at week 4 of 6.5-6.7 log cfu g-1 on wheat bran and oat, compared with 3.7-3.9 log cfu g-1 of L. acidophilus and 4.2-4.8 log cfu g-1 of L. plantarum at 20 °C 20% RH. Under the storage conditions of 30 °C 50% RH, survival of 4.5 log cfu g-1 of B. longum was also found on oat and peanut. This was two and four times higher than the population of L. acidophilus and L. plantarum, respectively. The results suggest that probiotics can survive on non-dairy foods under ambient storage conditions. However, the storage conditions, food matrices, and probiotic strains should be carefully chosen to maximize probiotic bacteria survival.

The Combined Effect of Blackcurrant Powder and Wholemeal Flours to Improve Health Promoting Properties of Cookies.

A diet with high glycaemic index, which causes rapid spikes in blood sugar level, can lead to disorders such as significantly increased risk for type 2 diabetes, cardiovascular disease and obesity. These conditions are also linked to the progression of cognitive decline and neurodegenerative diseases including Alzheimer's disease. Blackcurrant powder (BC) is a rich source of dietary fibre and bioactive compounds. Wholemeal wheat, barley and oat flours contain high amount of fibre. In this study, a model food (cookie) was developed and used to investigate the in vitro glycaemic glucose equivalent and antioxidant activities of the cookies made with three different wholemeal flours (wheat, barley and oat) with different replacement levels (5, 10 and 15%) of blackcurrant powder. Increasing the proportion of blackcurrant powder in the cookie resulted in a significant (P < 0.05) decrease in glucose release after in vitro digestion compared to the control. In addition, incorporation of blackcurrant powder in cookies up to 15% increased the antioxidant capacity. The combination of wholemeal flour and the bioactive compound rich blackcurrant has the potential to improve the nutritional value and reduce the glycaemic index of such foods.

High pressure processing improves the tenderness and quality of hot-boned beef.

Strip loins from different grades of cattle were subjected to two levels of high pressure processing (HPP) within 1h of slaughter at a commercial meat processing plant and chilled for 1day before freezing. The physical and eating quality characteristics of longissimus thoracis (strip loin) steaks from HPP were compared to meat that was chill aged for 1 or 28days. HPP produced meat after 1day with 60% lower shear force and higher sensory eating quality scores than 1day chill aged meat. Extended chill storage for 28days produced steaks of similar tenderness to HPP meat. HPP also increased the ultimate pH and decreased the cooking loss. Chilled storage of the gluteus medius from prime cattle for 28days significantly improved the shear force by 18%, whilst HPP improved both the shear force by 43% and the sensory eating quality scores. HPP can produce high eating quality eye of rump medallions within 1day of slaughter.

The Effects of Fortification of Legumes and Extrusion on the Protein Digestibility of Wheat Based Snack.

Cereal food products are an important part of the human diet with wheat being the most commonly consumed cereal in many parts of the world. Extruded snack products are increasing in consumer interest due to their texture and ease of use. However, wheat based foods are rich in starch and are associated with high glycaemic impact products. Although legume materials are generally rich in fibre and protein and may be of high nutritive value, there is a paucity of research regarding their use in extruded snack food products. The aim of this study was to prepare wheat-based extrudates using four different legume flours: lentil, chickpea, green pea, and yellow pea flour. The effects of adding legumes to wheat-based snacks at different levels (0%, 5%, 10%, and 15%) during extrusion were investigated in terms of protein digestibility. It was observed that fortification of snacks with legumes caused a slight increase in the protein content by 1%-1.5% w/w, and the extrusion technique increased the protein digestibility by 37%-62% w/v. The product developed by extrusion was found to be low in fat and moisture content.

Proteomic Differences between Listeria monocytogenes Isolates from Food and Clinical Environments.

Listeria monocytogenes is an organism associated with a wide range of foods. It causes listeriosis, a severe illness that mainly affects people with weakened immune systems. Proteomic profiles of three different L. monocytogenes isolates were studied using 1D SDS PAGE, 2DE and mass spectrometry. The protein banding patterns generated by 1D SDS PAGE of three strains of L. monocytogenes were found to be similar. Visual observations from 2DE gel maps revealed that certain spots appeared to have intensity differences. Key differences in proteins synthesis of three strains of L. monocytogenes were found using the PDQest TM 2DE Analysis software. Comparison showed that the clinical isolate (strain SB92/844) had 53.4% and 53.9% protein profile similarity with dairy isolate (strain V7) and seafood isolate (SB92/870), respectively. The identity of selected protein spots was achieved using MALDI-TOF and ion trap mass spectrometry. It was found that certain identified proteins (i.e., a major cold shock protein and superoxide dismutase) were expressed differently between two local strains of L. monocytogenes (SB92/844, SB92/870) and one strain from overseas (V7).

Concentrations of trace elements [corrected] and organochlorines in Mutton bird (Puffinus griseus).

The Sooty Shearwater (Puffinus griseus, commonly known as Mutton bird) is a migratory wild seabird, annually harvested for food by certain native groups in New Zealand and Australia and in many parts of the world. The concentrations of 22 elements and several organochlorine pesticides [2,2-bis(chlorophenyl)-1,1,1-trichloroethane (DDT), its derivatives dichlorodiphenyldichloroethene (DDE) and dichlorodiphenyldichloroethane (DDD), aldrin, chlordane, dicofol, lindane, and methoxychlor] in Mutton bird were determined over two consecutive years to evaluate its safety for human consumption. Twenty bird carcasses were purchased in each of 2007 and 2008 from a local source. No significant year effect (P>0.05) was found in the following nine trace elements: Al, As, Cd, Co, Cr, Cu, Ni, Pb and Se. The concentrations of Hg, Li and Na were higher (P<0.05) in 2008 samples compared to 2007. The toxic trace elements (mg/kg wet weight) in all the samples were below the maximum residual level (MRL). The concentration of Fe, Ca and Se in Mutton bird was higher than that in domestic land animal meats reported in literature. The residual organochlorine concentrations were all below the recommended MRL. Thus Mutton bird meat is high in essential nutrient elements and of low toxicological risk. Due to active use of agrochemical in New Zealand, a monitoring program for contaminants in Mutton birds is recommended.