Effect of the addition of pancreatic lipase on the ripening of dry-fermented sausages - Part 1. Microbial, physico-chemical and lipolytic changes

The effect of the addition of nine different amounts (3-500 units) of pancreatic lipase on the microbial and physico-chemical parameters and lipid fractions during the ripening of dry fermented sausages has been studied. No differences between conventional and lipase-added sausages were found for pH, dry matter and water activity. The addition of lipase caused a greater accumulation of products resulting from the triglyceride breakdown, mainly diglycerides and free fatty acids (FFA). The maximum rate of lipolysis was observed during the first week of the ripening process, specially in the fermentation phase. The greater the pancreatic lipase added, the higher lipolysis observed. At the end of the ripening, the levels of total FFA were clearly higher (1·5 to 5-fold) in all lipase-added batches than in the controls. This fact gave rise to the accumulation of a great amount of FFA, which can contribute either by themselves to the flavour of the sausage or can be available as substrates for further transformations which may generate other flavour compounds.     

Sensory evaluation of beef patties and sausages containing lipids with various component fatty acids

Sensory scores of beef patties and emulsion-type sausages containing varying levels of lipids with various component fatty acids were investigated. Sensory scores of beef patties containing solid fat were relatively constant at every fat level examined. However, scores of beef patties containing liquid oil decreased with increasing level of added oil. In the case of sausages, on the other hand, sensory evaluation scores markedly decreased with increasing the level of solid fat, while no significant change was observed in panel scores when liquid oil was mixed into sausages. Even 10% level of added lipids satisfactorily gave preferable acceptability to beef patties and sausages.     

Influence of starter cultures on the free fatty acids during ripening in Tea sausages

The influence of starter cultures on the free fatty acids content during ripening of Tea sausages, typical dry sausages produced in the south of Croatia, was studied. Three batches of Tea sausages were produced using different starter mixtures (Staphylococcus xylosus S81 and Lactobacillus sakei G20; Staphylococcus xylosus S142 and Lactobacillus sakei G20; Staphylococcus xylosus S206 and Lactobacillus sakei G20), while the control batch was produced without a starter. The amounts of free fatty acids present in the samples at the end of the ripening period were not significantly different, suggesting that the lipolytic enzymes naturally occurring in meat could play a predominant role in the free fatty acids release. Oleic and linoleic acids were present in the highest concentrations, while only small quantities of short-chain fatty acids were detected, with acetic acid being the most representative one.     

Preliminary analysis of the lipase gene (gehM) expression of Staphylococcus xylosus in vitro and during fermentation of naturally fermented sausages (in situ

Coagulase-negative catalase-positive cocci (CNCPC) play a very important role during the fermentation of sausages. In particular, they are involved in the aroma formation of the final product, because they release lipases that are able to free short-chain fatty acids that are contributing to the sensory characteristics of the fermented sausage. Few studies have been undertaken to elucidate the regulation of lipase gene expression in Staphylococcus xylosus by substrate molecules or products of lipolysis. The aim of this study was to analyze the gehM gene expression of S. xylosus DSMZ 6179 in vitro with growth media containing different concentrations of lipids and in situ during the maturation of fermented sausages. The results obtained suggest that a concentration that increases in triglycerides in the growth medium suppresses the expression of the lipase gene.     

微生物酶与肉组织酶对干发酵香肠中游离脂肪酸的影响

以戊糖乳杆菌31-1和木糖葡萄球菌为发酵剂生产干发酵香肠,就发酵剂对干发酵香肠成熟过程中游离脂肪酸的影响进行了研究。研究结果表明,在灌肠后的1周内,脂肪的水解主要是由组织酶引起的,微生物对脂肪微弱的水解能力主要表现在干发酵香肠成熟的后期,并且发酵剂不能改变发酵香肠中游离脂肪酸的变化模式。     

Differential modulation of enterocyte-like Caco-2 cells after exposure to short-chain fatty acids

The response of intestinal epithelial cells to short-chain fatty acids, which are increasingly used as food additives, was investigated. Human small intestinal epithelial cell model Caco-2 cells were exposed to formate, propionate and butyrate to assess their effect on cellular growth, metabolism, differentiation and protection against bacteria. The Caco-2 cells were entirely grown in the different short-chain fatty acids and respective growth patterns were determined. Differentiated cells were exposed to 0-20 mM short-chain fatty acids for 48 h and changes in DNA, RNA, (glyco)protein syntheses, sucrase isomaltase activity, transepithelial electrical resistance and protection against Salmonella enteritidis were measured. The short-chain fatty acids, altered linearly and differentially the growth pattern ranging from stimulation by formate to inhibition by butyrate. Formate inhibited cellular metabolism. Low concentrations of up to 5 mM propionate and 2 mM butyrate stimulated metabolism, while higher doses were inhibitory. Formate had no effect on sucrase isomaltase enzyme activity and transepithelial electrical resistance, whereas propionate and butyrate increased these markers of differentiation. Infection with S. enteritidis did not benefit from the short-chain fatty acid-induced transepithelial electrical resistance. It is concluded that formate, propionate and butyrate selectively and differentially modulate growth characteristics, cellular metabolism, sucrase isomaltase activity and transepithelial electrical resistance in a concentration- and carbon atom-related fashion. The short-chain fatty acid-induced transepithelial electrical resistance does not confer protection against S. enteritidis.     

Chemoenzymatic synthesis of structured triacylglycerols with conjugated linoleic acids (CLA) in central position

A chemoenzymatic process for the production of structured triacylglycerols (TAG) containing CLA at sn2 position and lauric acid at external ones is proposed. First, castor bean oil was chemically dehydrated and isomerised to obtain a new modified oil with very high proportion of CLA (>95%). Then, this new oil was used for enzymatic transesterification allowing the grafting of lauric acid at external positions of the TAG backbone by using 1,3 regioselective enzymes. Among these, Aspergillus niger lipase was not satisfactory giving very low lauroyl incorporation (<5%) On the contrary, lipases from Thermomyces lanuginosa (Lipozyme TL IM) and from Carica papaya latex allowed good reaction yields. The effect of the type of acyl donor was studied. With alkyl esters T. lanuginosa lipase provided a final incorporation of 58.9% after 72 h corresponding to 88.4% transesterification yield. Concerning C. papaya lipase, incorporation of lauroyl residues was lower than Lipozyme TL IM. This lipase exhibited higher performance with lauric acid accounting for 44.7% lauroyl incorporation at the end of reaction for a 67.1% transesterification yield. The effect of the substrates mole ratio was also evaluated. It was observed that a 1:3 TAG/acyl donor mole ratio was the most efficient for both lipases. Finally, fatty acids regiodistribution of the newly formed structured TAG was determined. With Lipozyme TL IM, the proportion of lauric acid incorporated at the sn2 position did not exceed 5.4% after 72 h while with C. papaya lipase a more pronounced incorporation of lauroyl residues at the central position (8.8%) was observed.     

Comparison of the compositional, microbiological, biochemical and volatile profile characteristics of three Italian PDO fermented sausages

Three Italian PDO fermented sausages, Varzi, Brianza and Piacentino, were compared for compositional, microbiological, biochemical and volatile profile characteristics. Mean values for the gross composition varied especially due to moisture, fat, total protein and nitrate concentration which reflected differences in the ingredients and some technological parameters. Cell numbers of the major microbial groups were almost similar among sausages. The major differences were found for Brochothrix thermosphacta, enterococci and moulds. Apart from their use as starters, Lactobacillus sakei and Lactobacillus curvatus were the dominant lactic acid bacteria and, as well as Staphylococcus xylosus, dominated the population of coagulase-negative staphylococci. Sausages differed for the hydrolysis of myofibrillar proteins and secondary proteolysis. Varzi, the sausage subjected to prolonged fermentation at 23–25 °C for 10 days before ripening, showed the highest degree of secondary proteolysis. Varzi and Brianza, the two fermented sausages manufactured by using microbial starters, showed the highest concentration and similar profiles of free amino acids. The peptidase activities contained in the aqueous extracts agreed with the above findings. A total of 52 volatile components, mainly alcohols, aldehydes and terpenes, were identified by solid-phase micro-extraction coupled with gas chromatography–mass spectrometry analysis. The volatile profiles of the three Italian PDO fermented sausages differed in part and, except for terpenes, the highest levels of the other chemical classes were found in Varzi and Brianza sausages. The composition of free fatty acids of the three Italian PDO sausages was rather similar. Monounsaturated fatty acids (MUFA) were found at the highest relative percentage followed by saturated (SFA) and polyunsaturated (PUFA) fatty acids. Oleic, palmitic, linoleic and stearic were the main free fatty acids found in all fermented sausages.     

浓缩型冻干发酵剂在鸭肉发酵香肠中的应用

将浓缩型冻干发酵剂应用于鸭肉发酵香肠中,对比液体发酵剂和自然发酵对香肠发酵过程中的影响。结果表明：接种冻干发酵剂的香肠48 h内pH值降到5.3以下,能够快速发酵产酸达到发酵香肠安全控制范围,且水分含量呈下降趋势,发酵末期降到30%左右;对各处理组鸭肉香肠的质构进行分析比较,接种冻干发酵剂处理组（S2处理组）以及液体发酵剂（L2处理组）的香肠硬度、弹性和咀嚼性均显著（P＜0.05）高于自然发酵（对照组）;此外,香肠发酵过程中酸价呈上升趋势,过氧化值先上升后下降,各处理组差异不显著（P＞0.05）,推断内源性脂酶比微生物发酵剂产生的脂酶发挥更大作用;各类型脂肪酸占总游离脂肪酸的比例大小顺序为：饱和脂肪酸＞单不饱和脂肪酸＞多不饱和脂肪酸,接种发酵剂的处理组游离脂肪酸总量显著高于对照组（P＜0.05）,表明发酵剂的添加对游离脂肪酸的释放起到一定促进作用,在一定程度上丰富了香肠风味的前体物质。     

Study of the shelf life of a dry fermented sausage "salchichon" made from raw material enriched in monounsaturated and polyunsaturated fatty acids and stored under modified atmospheres

The microbiological, physico-chemical and sensory properties of salchichon with high unsaturated fat content, packed under vacuum and 20/80% CO(2)/N(2) modified atmosphere, were evaluated to determine its quality changes during storage under refrigeration. These sausages were manufactured with pork meat and pork backfat obtained from pigs fed with three different diets (control diet-CO, high oleic diet-HO and high linoleic diet-HL). In general, few significant differences were found in counts of different groups of microorganism between the three types of sausages and no difference between the packaging methods. A reduction in pH values was observed during storage and no great differences were determined by storage period on water activity (a(w)). Both parameters (pH and a(w)) presented similar results to those found in different Spanish sausages and other European dry fermented products. The sensory results denoted that sensory quality gradually decreased during storage under both packaging conditions (vacuum and 20/80% CO(2)/N(2)), so it is not advisable to store longer than 150days. On the other hand, fermented sausages with high content of unsaturated fatty acids had similar sensory properties to those of conventional sausages, and even a comparable sensory stability. In conclusion, the results showed healthier salchichons (HO and HL) similar to the traditional (CO) one could be manufactured and stored under refrigeration after slicing for a reasonable period, but the advantage of the gas mixture packaging (20/80% CO(2)/N(2)) versus vacuum packaging was not clear.     

Free fatty acids and oxidative changes of a raw goat milk cheese through maturation

Free fatty acids (FFA) and lipid and protein oxidation changes were studied throughout maturation process of a raw goat milk cheese with protected designation of origin. Cheeses were analyzed at 4 different times of maturation, at 1, 30, 60, and 90 d. All FFA significantly increased during maturation and the relative increase was higher for long-chain than medium- or short-chain FFA. At the end of maturation, oleic (C18:1 n9), butyric (C4:0), and palmitic (C16:0) acids were the most abundant. The higher levels of short-chain fatty acids (SCFA) regarding total FFA obtained at the end of Ibores cheese ripening compared with other raw goat milk cheeses, highlight the notable role of SCFA on the flavor of this cheese owing to their low-odor thresholds. Lipid oxidation values significantly increased during maturation process but low levels of malondialdehyde were reported; however, protein oxidation did not significantly change during ripening.     

桑椹多酚对广式腊肠风味的影响

植物多酚因能有效抑制肉制品的过氧化近年来备受关注,然而由此造成肉制品风味的变化还鲜有报道。鉴于此,将不同剂量的桑椹多酚提取物（0.5 g/kg和1.0 g/kg）加入到肉糜中,制成广式腊肠,分析多酚添加前后广式腊肠的挥发性风味、游离脂肪酸、游离氨基酸的组成和含量变化,以及桑椹多酚对广式腊肠感官评价的影响。研究发现,添加桑椹多酚后,腊肠中挥发性风味物质由空白组的49种减少到32种,腊肠游离脂肪酸的含量明显下降。值得注意的是,由于蛋白质水解增强,添加桑椹多酚后,腊肠中呈味氨基酸（Glu和Arg）的释放得到促进。储藏末期（10 d）,添加1.0 g/kg桑椹多酚的腊肠游离氨基酸的含量达到5904.91 mg/100 g,较对照组提高了10.82%。总体而言,桑椹多酚的添加会抑制腊肠中某些挥发性风味物质的形成,抑制腊肠中的肉腥味,促进呈味氨基酸的释放,但对腊肠游离脂肪酸组成的影响并不显著。     

Influence of lamb rennet paste on the lipolytic and sensory profile of Murcia al Vino cheese

The influence of lamb rennet paste (71.1% chymosin, 177 international milk-clotting units/mL, 4.57U/g of lipase activity) during the ripening of Murcia al Vino goat cheese was studied. The aim of this study was to improve the knowledge of the effect of lamb rennet paste on the lipolytic patterns in this type of cheese by reference to the evolution of total and free fatty acids. A sensory analysis was carried out to compare cheeses made with commercial and paste rennet. The rennet paste showed higher lipolytic activity, enhancing the production of short-chain free fatty acids. In addition, the cheese produced with lamb rennet paste had a slightly more bitter and piquant taste, making it an attractive commercial alternative that can be used to develop new varieties of goat cheese.     

Nutritional and sensory properties of dry fermented sausages enriched with n-3 PUFAs

Enrichment of dry fermented sausages with n − 3 fatty acids through a partial substitution of pork backfat by deodorised fish oil resulted in improved nutritional properties with regard to conventional sausages, without affecting sensory properties and oxidation status. The developed products supplied 0.64 g EPA/100 g and 0.46 g DHA/100 g product, and showed PUFA + MUFA/SFA ratio of 1.76 and n − 6/n − 3 ratio of 2.97. No signs of oxidation were found in any type of sausages, control and modified. None of the dienals and trienals reported as secondary lipid oxidation products and typical for fish oil were detected in the modified sausages. Instrumental colour differences were detected, however, without relevance in the sensory analysis. The sensory evaluation panel did not find differences in general acceptability. The modified dry fermented sausages can be considered a technologically viable functional food.     

Effects of dietary clenbuterol and salbutamol on meat quality in veal calves

Sixty male calves were assigned to one of four treatments, including (1) an untreated control, (2) clenbuterol; 1·6 mg/(calf.day), (3) and (4) salbutamol: 60 and 100 mg/(calf.day). Animals were milk-fed during 24 weeks, and β₂-agonists (BA) treatment occurred during the last 4 weeks including a 3-day withdrawal period before slaughter. Under identical feed intake amongst groups, treatments with BA had significantly positive effects on carcass weights and dressing percentages. BA induced muscle hypertrophy as evidenced by an improved EUROP carcass classification, 1 19–24% higher m. longissimus loin eye area and higher muscle protein/DNA and protein/RNA ratios. Through a dilution effect, this might explain the lower haem content of the m. semimembranosus and m. longissimus lumborum (LL) and diminished Hunter a^* value (redness) of the latter muscle. A higher ultimate pH was found in the m. semitendinosus and m. psoas major of BA-treated calves. The m. longissimus thoracis glycogen content appeared to be lowered by salbutamol. No effects of BA were encountered for water-holding properties of this muscle. Significantly higher shear force values (31–45%) for treated animals were measured for the LL which were parallelled by higher initial values for the calpain-inhibiting activity of calpastatin and a decrease of the μ-calpain/calpastatin ratio. EUROP fatness of carcasses and LL intramuscular fat content tended to be lowered by both BA. Effects of clenbuterol and of the higher salbutamol dose, which on a molecular base was administered at a 70 times higher level, were roughly of a comparable order. The lower dose mostly had intermediate effects. This means that in veal calves, clenbuterol is biologically more potent than salbutamol.     

Screening of microbes for lipases specific for saturated medium and long-chain fatty acids of milk fat

Over 250 microbial strains were screened for lipase activity specific towards saturated medium and long-chain fatty acids. Strains showing trimyristin (C14:0) hydrolysis on agar medium were further streaked on tricaprylin (C8:0) and on olive oil (mostly C18:1) agar to identify strains hydrolysing C12–C15 fatty acids with modest or no activity towards short-chain and unsaturated fatty acids. The twenty strains showing the desired specificity were grown as liquid cultures and analysed for p-nitrophenyl (p-NP) myristate hydrolysis in the presence of Triton X-100. Cells from the cultures showing the highest activity towards p-NP myristate were analysed for specificity towards p-NP esters of different fatty acid chain lengths. On the basis of the p-NP ester profiling, four lipase samples were selected for hydrolysis experiments on butter oil. Of these, the sample obtained from the cultivation of Didpodascus capitatus met the screening criteria: C12:0–C16:0 specificity and no preference towards C18:0 and C18:1 fatty acids.     

中性大蒜果聚糖体外发酵产短链脂肪酸

大蒜果聚糖是大蒜深加工后废弃蒜渣中的主要成分,不能被人体消化利用,可选择性增殖肠道益生菌,具有开发成新型益生元的潜力。体外发酵产短链脂肪酸的分析对进一步评价大蒜果聚糖对肠道微生态的影响有重要意义。方法:本研究通过体外模拟人体肠道系统的pH、温度和营养条件,接种人体新鲜粪便悬浮液进行间歇式静态厌氧培养,采用高效液相色谱法(HPLC)分析不同聚合度(平均DP为11、16、21)的中性大蒜果聚糖为唯一碳源,培养0、4、8、12、24 h产短链脂肪酸的情况。结果:发酵12 h时短链脂肪酸浓度达到最高,之后便开始下降。其中样a(平均DP为11)的发酵液中总SCFA浓度最高,达86.38μmol/mL。总短链脂肪酸产量为样a(DP 11)>样b(DP16)>样c(DP 21)。24 h时,各发酵液的短链脂肪酸中丁酸和乳酸的比例较乙酸和丙酸高。结论:大蒜果聚糖能被人体肠道微生物发酵利用在体外产短链脂肪酸。其中丁酸和乳酸的摩尔浓度较乙酸和丙酸高。聚合度低的大蒜果聚糖产酸高于聚合度高的果聚糖。     

The antioxidant properties of baechu-kimchi and freeze-dried kimchi-powder in fermented sausages

The antioxidant properties of baechu-kimchi in fermented sausages were evaluated by investigating its effect on the changes in total free fatty acid content, peroxide value, and TBARS during ripening. The conditions for its antioxidant effectiveness were optimized in a model system (beaker sausages). For this, the conditions of added kimchi – form (kimchi and freeze-dried kimchi-powder), fermentation temperature (7 and 20 °C) and concentrations (5%, 10% and 15%) – were varied. The results were compared with that of the control without kimchi. According to the evolution of peroxide value and TBARS, the addition of 5% and 10% kimchi clearly indicated retardation of the development of lipid oxidation. However, an increase in the amount of kimchi added caused a prooxidant effect. The antioxidant effect of kimchi has been found to be dependant on its fermentation temperature as the batches treated with kimchi fermented at 20 °C displayed better protection against the formation of oxidative products than those fermented at 7 °C. Kimchi-powder did not show any antioxidant properties.     

Fat effect on physico-chemical, microbial and textural changes through the manufactured of dry-cured foal sausage Lipolysis, proteolysis and sensory properties

The effect of fat content on chemical traits related to dry-curing process (pH, moisture and water activity), color and textural properties and changes of free fatty acids and amino acids compositions during the processing of foal dry-cured sausages were studied. For this purpose, three batches (20 units per batch) of dry fermented sausages with different pork back fat content (5%, 10% and 20%) were manufactured; low fat (LF), medium fat (MF) and high fat (HF), respectively. Samples at 0days (mix before stuffing), and after 7, 14, 28, 42 and 49days of ripening were taken. The fat level affected color and textural parameters at the end of the process, showing dry-cured foal sausage with the higher level of fat, the highest values of luminosity and the least hardness. No significant differences (P>0.05) among batches were detected on total viable counts, lactic acid bacteria and Microccaceae during the process. Regarding lipolysis and lipid oxidation it can be deduced that the increase in the fat level encouraged the production of free fatty acids and 2-thiobarbituric acid reactive substances. At the end of the ripening individual free fatty acids followed this order: oleic, palmitic, linoleic and stearic acid, representing 82-95% of the total free fatty acids. Final level of TBARS index was in the worst case of 1.23mg MDA/kg of sausage. On the contrary, the batch with lesser fat content showed the highest levels of free amino acids at the beginning and at the end of the process, showing final values of 1.6%.