Donkey Milk for Manufacture of Novel Functional Fermented Beverages

The aim of this work was to investigate on the functional features of a donkey milk probiotic berevage as a novel food. Particularly, it was to study the decrease of lactose content and the antioxidant activity of standard yogurt (YC) and probiotic yogurt (YP; Lactobacillus acidophilus, Lactobacillus casei) from donkey milk during the storage up to 30 d at 4 ºC. The evolution of lactose content using enzymatic‐spectrophotometric kits was analyzed. Antioxidant activity of yogurt was measured using 2,2’‐azino‐bis‐3‐ethylbenzothiazoline‐6‐sulfonic acid (ABTS), ferric reducing antioxidant power (FRAP), and thiol assays. Parallel consumer sensory studies were carried out as consumer test in order to gain information about the impact of these novel fermented beverages on sensory perceptions. The statistical analysis has shown significant effect of studied factors. The results showed that the lactose content gradually decreased during storage in both yogurts, reaching values of 2.36% and 2.10% in YC and YP, respectively, at 30 d (P < 0.05). During storage of both yogurt types, the antioxidant activity increased, but YP showed a higher antioxidant activity than YC. The results suggest that the antioxidant activity of yogurt samples was affected by cultures of lactic acid bacteria (LAB). We conclude that the fermented donkey milk could be configured as health and nutraceutical food, which aims to meet nutritional requirements of certain consumers groups with lactose or cow milk protein intolerance.     

Production and purification of antioxidant peptides from a mungbean meal hydrolysate by Virgibacillus sp. SK37 proteinase

Antioxidant peptides of mungbean meal hydrolysed by Virgibacillus sp. SK37 proteinases (VH), Alcalase (AH) and Neutrase (NH) were investigated. The antioxidant activities based on 2,2′-azinobis (3-ethyl-benzothiazoline-6-sulphonate) (ABTS) radical-scavenging, ferric-reducing antioxidant power (FRAP) and metal chelation of VH were comparable to those of NH. VH was purified using ultrafiltration, ion exchange and gel filtration chromatography. The purified peptides (F37) from VH, which had the highest specific antioxidant activity, consisted of four peptides containing an arginine residue at their C-termini. In addition, the ABTS radical-scavenging activity of the purified peptides (F42) at 0.148 mg/ml was comparable to that of 1 mM of butylated hydroxytoluene (BHT). These two fractions were stable over a wide pH (4–10) and temperature (25–121 °C) range. Virgibacillus sp. SK37 proteinase is a potential processing-aid for the production of a mungbean meal hydrolyzate with antioxidant properties.     

Casein hydrolysis by Bifidobacterium longum KACC91563 and antioxidant activities of peptides derived therefrom

Milk protein is a well-known precursor protein for the generation of bioactive peptides using lactic acid bacteria. This study investigated the antioxidant activity of bovine casein hydrolysate after fermentation with Bifidobacterium longum KACC91563 using the 2,2′-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) assay and total phenolic content (TPC). The antioxidant activities of the 24-h and 48-h hydrolysates were higher than that of the 4-h hydrolysate (2,045.5 and 1,629.3 μM gallic acid equivalents, respectively, vs. 40.3 μM) in the ABTS assay. In contrast, TPC values showed activities of 43.2 and 52.4 μM gallic acid equivalents for the 4-h and 24-h hydrolysates, respectively. Three fractions (≥10 kDa, ≥3 but <10 kDa, and <3 kDa) were separated from the 24-h hydrolysate by ultrafiltration. Among these fractions, the <3 kDa fraction exhibited the highest antioxidant activity (936.7 μM) compared with the other fractions (42.1 and 34.2 μM for >10 kDa and 3–10 kDa fractions, respectively). Through liquid chromatography-electrospray ionization-tandem mass spectrometry analysis, 2 peptides, VLSLSQSKVLPVPQK and VLSLSQSKVLPVPQKAVPYPQRDMPIQA, containing the fragment VLPVPQ that has antioxidant properties, were identified in the <3 kDa fraction after 24 h of hydrolysis. The present study demonstrates the possibility of antioxidant peptide production from bovine casein using Bifidobacterium longum.     

Consumer acceptability and antioxidant potential of probiotic-yogurt with spices

The study focuses on developing novel probiotic yogurts containing spices with acceptable sensory properties, therapeutic levels of probiotics and with beneficial antioxidant capacity. Eight types of yogurts with added spice oleoresins (cardamom, cinnamon and nutmeg) and probiotics [Lactobacillus acidophilus strain 5(LA5), or Bifidobacterium animalisssp.Lactis (Bb12)] were produced. Two successive consumer sensory taste panels (n = 54) using a nine point hedonic scale were conducted to evaluate the acceptability of the yogurts. Viable counts of probiotics and antioxidant capacity of yogurt samples at 1, 7, 14 and 28 days of storage at 4 °C were monitored. The probiotic-yogurt products containing spices showed good sensory properties, with the best results obtained with cardamom oleoresin (with LA5 or Bb12). The presence of spice oleoresins (cardamom, nutmeg and cinnamon) did not affect the probiotic population (LA5or Bb12) in yogurt during 4 weeks of refrigerated storage. The antioxidant capacity (with LA5or Bb12) over the storage period was also maintained.     

Comparison of antioxidant capacity of cow and ewe milk kefirs

This research aimed to evaluate the effects of using either grain or commercial starter culture on the antioxidative capacity of cow and ewe milk kefirs. The antioxidant capacity of kefir samples during fermentation and 21 d of storage was assessed by using 3 assays: 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS) radical cation decolorization; 2,2-diphenyl-1-picrylhydrazyl (DPPH^?) radical scavenging activity assay; and Fe⁺³-reducing power (ferric reducing antioxidant power assay, FRAP). Vitamin E and β-carotene contents were also quantified. All kefir samples exhibited varying values for DPPH, ABTS, and FRAP assays depending on the starter culture and milk type. Vitamin E and β-carotene contents were similar in all kefir samples during storage. The results of this study suggest that milk type (cow or ewe) and culture type (kefir grains or commercial starter) were the significant parameters for the antioxidative activity of kefir.     

Consumer-perception,nutritional, and functional studies of a yogurt with restructured elderberry juice

We investigated the effects of supplementation of yogurt with elderberry juice (Sambucus nigra L.), in both natural and restructured forms, on certain technological, physicochemical, sensory, and health-promoting properties of yogurt, including antioxidant activity and inhibition of angiotensin-converting enzyme (ACE), α-amylase, and α-glucosidase activity. Consumer acceptance of the yogurt-juice products was assessed. Gel-strength restructured elderberry juice retained a spherical shape and most of the juice, despite decreasing in mass from 57.2 to 50.9 g during storage. As a result, yogurt supplemented with 10 and 25% restructured elderberry juice appeared to be more desirable from a sensory and technological perspective than yogurt with natural juice. Yogurt supplemented with restructured elderberry juice had a high water-holding capacity (94.4–96.4%), exhibited no spontaneous whey syneresis, and maintained a dense consistency (up to 5,626 g). Consumer penalty analysis of the just-about-right diagnostic attributes indicated that the flavor of these yogurts may not be sufficiently refreshing. High correlation was demonstrated between ACE inhibition, ABTS, α-amylase, and α-glucosidase in yogurt supplemented with restructured juice. An in vitro gastrointestinal simulation estimated bioaccessibility of antioxidants to be in the range of 62 to 66%. This model fermented yogurt supplemented with restructured elderberry juice is a novel dairy-juice beverage that represents a new approach for the development of functional fruit yogurt beverages.     

Assessment of antioxidant capacity by method comparison and amino acid characterisation in buffalo milk kefir

This study aimed to analyse the antioxidant capacity and total phenolic content of buffalo milk kefir fermented with grains or a commercial starter culture, as well as characterisation of the amino acid profile. During fermentation, the ferrous‐reducing activity of samples increased, while the 2,2‐diphenyl‐1‐picrylhydrazyl (DPPH) and 2,2′‐azino‐bis‐(3‐ethylbenzothiazoline‐6‐sulphonic acid) (ABTS) scavenging activity decreased. The highest value of the ABTS and DPPH scavenging activity was recorded for buffalo milk kefir with grain, which also showed a lower ferric‐reducing antioxidant power during storage. Statistical analyses showed a significant effect of starter culture type and fermentation/storage time on the antioxidant capacity of buffalo kefir.     

Fate of Escherichia coli and Kluyveromyces marxianus contaminants during storage of Greek-style yogurt produced by centrifugation or ultrafiltration

The production of Greek-style yogurts requires more processing steps than traditional yogurt, which increases the possibility of microbial contamination by pathogens or spoilage organisms. The growth and survival during storage of two microbial contaminants (Escherichia coli and Kluyveromyces marxianus) in Greek-style yogurt, produced by centrifugation or ultrafiltration, was compared with that in regular stirred yogurt. E. coli strain ATCC^® BAA-1430™ was shown to be a suitable surrogate for pathogenic O157:H7 in yogurt. The increased buffering capacity of the Greek-style yogurts produced from ultrafiltered milk led to lower E. coli viability during storage. On the other hand, the Greek-style yogurt seems to promote faster growth of the dairy yeast K. marxianus at a storage temperature of 4 °C.     

Phenolic fortification of yogurt using grape and callus extracts

In this study, acidified ethanol extracts of four different grape varieties (Cabernet sauvignon, Chardonnay, Shyrah, and Merlot) and grape callus were supplemented into yogurt as functional ingredients. The total phenolic substances, total anthocyanin content, and antioxidant activity of the yogurts were measured using chemical analyses. Yogurts inoculated with red grape and callus extracts displayed high phenolic - anthocyanin content and thus exhibited higher antioxidant power compared to yogurts containing chardonnay extracts and control samples. The storage time significantly affected the free radical scavenging capacity of the yogurts. The yogurts supplied with grape callus extract displayed the greatest antioxidant power on the first day of storage compared to all the assayed samples. The Gas Chromatography (GC) analysis revealed the presence of at least 10 individual bioactive phenolic compounds in the callus yogurt. The results obtained from this investigation shows that grape callus culture has a potential to be used as a food supplement to play a role in reducing the risk of developing chronic diseases such as cancer and cardiovascular disease.     

铁离子螯合亲和层析分离抗氧化活性核桃肽

采用铁离子螯合亲和层析方法分离出具有不同抗氧化活性的核桃肽,并探讨核桃肽的抗氧化活性与铁结合能力的关系。结果表明,核桃肽在酸性条件(pH 5.5)下与铁的结合能力最强,随着pH逐渐升高,结合能力下降。磷酸氢二钠对吸附到铁亲和层析柱上核桃肽的洗脱效果最好,通过阶段洗脱,得到铁结合能力逐渐增强的核桃肽组分(F1、F2和F3)。对其总还原能力和2,2-联氮-二(3-乙基-苯并噻唑-6-磺酸)二铵盐(ABTS)自由基清除能力进行测定,发现其抗氧化能力为F3>F2>F1(P<0.05)。结果表明,核桃肽的铁结合能力越强,其抗氧化活性越高。     

Antioxidant activity of yogurt made from milk characterized by different casein haplotypes and fortified with chestnut and sulla honeys

The aim of this work was to evaluate the antioxidant activity of yogurt made from milk characterized by different casein (CN) haplotypes (α_s1-, β-, κ-CN) and fortified with chestnut and sulla honeys. The CN haplotype was determined by isoelectric focusing, whereas antioxidant activity of yogurt was measured using 2,2’-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid and ferric-reducing antioxidant power. The statistical analysis showed a significant effect of the studied factors. The results showed that chestnut honey presented the highest phenolic acid and flavonoid contents, which are closely associated with its high antioxidant activity. The antioxidant activity of fortified yogurt samples was affected both by different CN haplotypes and by type of honey added. Yogurts fortified with chestnut honey showed higher antioxidant activity than those fortified with sulla honey. The different behavior observed among the fortified yogurts led us to hypothesize that the effects of protein-polyphenol complex on antioxidant activity are interactive. The results suggest that milk proteins polymorphism and polyphenols play different roles in affecting the bioavailability and the antioxidant activity of yogurt.     

Angiotensin I-converting (ACE)-inhibitory and anti-inflammatory properties of commercially available Greek yoghurt made from bovine or ovine milk: A comparative study

The objective of this study was to compare the presence of peptides with angiotensin I-converting (ACE)-inhibitory and immunomodulatory activities in commercial Greek yoghurt made from bovine or ovine milk. Water soluble extracts (WSE) in yoghurts made from ovine milk exhibited higher ACE-inhibitory activity than their bovine counterparts. Similarly, ovine yoghurt WSE inhibited more effectively the expression of two pro-inflammatory genes (inducible NO synthase and cyclooxygenase-2) by ovine monocytes. Following gel filtration of WSE by Sephadex G25, one fraction exhibited only ACE-inhibitory activity, two only immune inhibitory while one possessed both activities. Thus, selected peptidic fractions mimicked the original effects observed in WSE prior to fractionation.     

分子量对酪蛋白多肽抗氧化活性的影响

在对酪蛋白酶解产物制备工艺进行优化的基础上,对不同分子量抗氧化肽(>3ku,1～3ku,<1ku)的抗氧化活性进行了评价。首先对酶的种类、酶底物比及水解时间进行了单因素实验,最终确定采用碱性蛋白酶,在pH8.0,55℃,底物浓度5%,酶底物比0.192AU/g的条件下,酶解4h所得水解物抗氧化活性最高。经过超滤和凝胶过滤层析分离获得不同分子量的抗氧化肽,并采用2,2′-连氮基-双-(3-乙基苯并二氢噻唑啉-6-磺酸)二铵盐(ABTS+.)、羟自由基和超氧自由基清除活性评价其抗氧化性。结果表明,ABTS+.清除活性与分子量呈负相关(r=-0.898,p<0.01),分子量低于1ku组分活性最强(2mg/mL,Trolox当量为2.08±0.05mmol/L);分子量低于3ku的抗氧化肽羟自由基清除活性较高(IC50:1～3ku,4.43±0.03mg/mL;<1ku,4.35±0.06mg/mL);分子量高于3ku组分主要分布在3～5ku,超氧自由基清除活最强(10mg/mL,66.1%±1.0%)。     

Physicochemical and sensory properties of yogurts containing sacha inchi (Plukenetia volubilis L.) seeds and β-glucans from Ganoderma lucidum

Dairy products have been widely used for adding various biomolecules with the aim of improving their functional properties and health benefits. In this study, the physicochemical properties and sensory acceptance of yogurts enriched with sacha inchi (Plukenetia volubilis) seeds (SIS) and β-glucans from Ganoderma lucidum (BGGL) were investigated. The angiotensin-converting enzyme–inhibitory activity of some yogurt samples was also evaluated. Yogurts were produced from reconstituted skim milk powder, and SIS (4% wt/wt) and BGGL were added at different concentrations (0–1.5% wt/wt). The fermentation kinetics were not affected by the enrichment process. The addition of SIS and BGGL significantly increased the contents of protein, fat, carbohydrates, ash, total solids, aspartic acid, serine, arginine, glycine, threonine, tyrosine, and alanine. α-Linolenic (49.3%) and linoleic (32.2%) acids were the main fatty acids found in the enriched samples, whose values were about 50- and 25-fold higher than those of the control yogurt. The textural parameters (firmness, consistency, cohesiveness, and index of viscosity) of the enriched yogurts were significantly lower than those of the control samples during the whole storage period. All enriched yogurts showed a sensorial acceptance higher than 70% by untrained panelists. The angiotensin-converting enzyme–inhibitory activity of some selected yogurt samples ranged between 36 and 59%. These results indicate that SIS and BGGL could be used as natural ingredients for improving the nutritional value of yogurt and fermented milks.     

Survival of free and encapsulated probiotic bacteria and their effect on the sensory properties of yoghurt

The survival and effect of free and calcium-induced alginate-starch encapsulated probiotic bacteria (Lactobacillus acidophilus and Bifidobacterium lactis) on pH, exopolysaccharide production and influence on the sensory attributes of yogurt were studied over 7 weeks storage. Addition of probiotic bacteria (free or encapsulated) reduced acid development in yogurt during storage. Post-acidification in yogurt with encapsulated probiotic bacteria was slower compared to yogurt with free probiotic bacteria. More exopolysaccharides were observed in yogurts with probiotic cultures compared to those without probiotic cultures. The results showed that there was an increased survival of 2 and 1 log cell numbers of L. acidophilus and B. lactis, respectively due to protection of cells by microencapsulation. The addition of probiotic cultures either in the free or encapsulated states did not significantly affect appearance and colour, acidity, flavour and after taste of the yogurts over the storage period. There were, however, significant differences (P<0.05) in the texture (smoothness) of the yogurts. This study has shown that incorporation of free and encapsulated probiotic bacteria do not substantially alter the overall sensory characteristics of yogurts and microencapsulation helps to enhance the survival of probiotic bacteria in yogurts during storage.     

Bioavailability of peptides from casein hydrolysate in vitro: Amino acid compositions of peptides affect the antioxidant efficacy and resistance to intestinal peptidases

Measurements of in vitro bioactivity to support health benefits of bioactive compounds should be accomplished with estimates of their bioavailability to bolster nutritional significance to health claims. In vitro bioavailability of casein and casein peptides (casein hydrolysate and four peptide fractions) measured by the amount of peptide nitrogen is discussed. Antioxidant activities during gastrointestinal digestion and Caco-2 cell absorption were investigated as indices of peptide degradation. The antioxidant capacities of Trolox equivalent and oxygen radicals were used for assessing antioxidant efficacy of surviving peptides. Results showed that casein hydrolysate improved bioavailability compared to casein. Amino acid composition of peptides affected the resistance of peptides to digestive enzymes and intestinal peptidases. The acidic peptide fractions had higher bioavailability and a higher residual ratio of antioxidant activity. The peptides in the digest and absorbate of acidic fraction F1 with the highest bioavailability (23.14%) and the residual ratio of antioxidant activity were identified, and 12 intact, absorbed peptides (IAP) were obtained. Eleven of twelve of the IAPs were from β-Casein, and their amino acid components were rich in acidic and hydrophobic amino acids. Identification of IAPs might provide insight into the mechanism of how peptide structure provides resistance against peptidases by Caco-2 cells.     

Effect of addition of Versagel on microbial, chemical, and physical properties of low-fat yogurt

The objective of this study was to examine the effect of Versagel on the growth and proteolytic activity of Streptococcus thermophilus 1275 and Lactobacillus delbrueckii ssp. bulgaricus 1368 and angiotensin-I converting enzyme inhibitory activity of the peptides generated thereby as well as on the physical properties of low-fat yogurt during a storage period of 28 d at 4 degrees C. Three different types of low-fat yogurts, YV0, YV1, and YV2, were prepared using Versagel as a fat replacer. The fermentation time of the low-fat yogurts containing Versagel was less than that of the control yogurt (YV0). The starter cultures maintained their viability (8.68 to 8.81 log CFU/g of S. thermophilus and 8.51 to 8.81 log CFU/g of L. delbrueckii ssp. bulgaricus) in all the yogurts throughout the storage period. There was some decrease in the pH of the yogurts during storage and an increase in the concentration of lactic acid. However, the proteolytic and ACE-inhibitory potential of the starter cultures was suppressed in the presence of Versagel. On the other hand, the addition of Versagel had a positive impact on the physical properties of the low-fat yogurt, namely, spontaneous whey separation, firmness, and pseudoplastic properties.     

Identification of bioactive peptides in commercial Cheddar cheese

This study examined the presence of antimicrobial, antioxidant and antihypertensive peptides in three commercially available Australian Cheddar cheeses. Peptide extracts as well as fractionated peptide extracts were examined. Commercial cheese A peptides exhibited the greatest inhibition against Bacillus cereus and also commercial cheese A fractionated peptides greater than 10 kDa showed the highest inhibition against B. cereus. Commercial cheese A peptides also showed the highest inhibition of 2,2-diphenyl-1-picrylhydrazyl (DPPH), a free radical used to measure antioxidant activity. All cheese fractionated peptides greater than 10 kDa demonstrated higher inhibition of DPPH after fractionation. Antihypertensive peptides were determined by inhibition of the angiotensin-converting enzyme (ACE). Overall, commercial cheese A had the lowest concentration required to inhibit ACE and commercial cheese A fractionated peptides lower than 5 kDa had the lowest inhibition after fractionation. These preliminary findings suggest that peptide extracts of three commercial Australian Cheddar cheeses exhibit antimicrobial, antihypertensive and antioxidant properties.     

Assessment of the Antioxidant Properties During Storage of a Dessert Made from Grape, Cherry, and Berries

A 1‐y storage trial was designed involving different temperatures (8, 21, and 30 °C) to assess the antioxidant properties of a dessert formulated with concentrated juices of grape, blackberry, blackcurrant, raspberry, and cherry. The total antioxidant activity (TAA) of the dessert was measured as the scavenging capacity of the 2,2′‐azino‐di‐[3‐ethylbenzothiazoline‐6‐sulphonate] radical cation (ABTS) and as its iron‐reducing ability (FRAP). Color was spectrophotometrically measured, and vitamin C and phenolics were analyzed by HPLC. The color parameters and the concentrations of different antioxidant compounds were affected by storage conditions, especially in samples stored at 30 °C, although the TAA remained practically invariable during storage.     

羊肉火腿加工过程中粗肽抗氧化活性

为明确羊肉火腿加工过程中内源抗氧化肽的抗氧化能力变化,本实验以不同加工时期干腌羊肉火腿为研究对象,利用磷酸盐缓冲液提取粗肽并测定多肽质量分数,对粗肽还原力、1,1-二苯基-2-三硝基苯肼（1,1-diphenyl-2-picrylhydrazyl,DPPH）自由基清除率、2,2’-联氮-二(3-乙基-苯并噻唑-6-磺酸)（2,2’-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid),ABTS）阳离子自由基清除率、羟自由基清除率、脂质过氧化抑制率进行评价。结果表明：羊肉火腿加工过程中多肽质量分数显著升高,抗氧化活性显著增强,具体表现为加工过程中还原力显著增强（P＜0.05）,ABTS阳离子自由基和DPPH自由基清除率总体呈上升趋势,分别在成熟结束期和成熟中期表现出最强抗氧化能力（分别为0.09、0.017 8 mmol/L）;羟自由基清除率总体呈下降趋势,在风干期抗氧化能力达到最大。十二烷基硫酸钠-聚丙烯酰胺凝胶电泳结果表明,从原料腿到成熟结束期肌肉中蛋白质逐渐发生降解,产生小分子抗氧化肽。本实验为下一步研究羊肉火腿多肽的抗氧化机制提供参考。