Enzymic and in sacco methods to estimate rumen degradation of food protein in cattle

BACKGROUND: Two factorial studies compared enzymic and in sacco methods to estimate degradation of ruminant foods. Enzyme degradation (in vitro = enzyme) was determined from the release of leucine‐equivalent amino acid (LA) crude protein (CP) from sunflower meal (SF), maize gluten meal (MG), distillers' dark grain (DG) and field beans (FB) after their separate incubations with Streptomyces griseus enzyme for 0–24 h. In sacco crude protein (CP) degradation of these foods was estimated during washing (0 h) and rumen incubations in fistulated cows for 2–24 h. The LA data were expressed as g LA per either kg of CP (LACP) or acid‐hydrolysable LA (HLA) of each food and compared with in sacco data. RESULTS: These methods showed comparable degradation with time (P < 0.01). The in sacco and HLA were greater than LACP for all foods except MG where in sacco value was either lower or equal to LACP depending upon the incubation time (P > 0.05 or P < 0.05). Conversely, HLA was significantly (P < 0.01) greater than LACP from 2 h onwards. At 0 h, in sacco values were significantly greater than those of enzyme for SF, DG and FB (P < 0.05) but not for MG. The foods differed significantly for degradation constants (a, b, c) in each method (P < 0.05). CONCLUSIONS: Despite variations between in sacco and enzyme estimates for different foods, the relationships between these estimates suggest that the HLA enzyme method has the potential to estimate food degradation. Copyright © 2007 Society of Chemical Industry     

In Sacco法评定玉米的不同加工处理方式对泌乳奶牛瘤胃干物质和淀粉降解规律的影响

以3头安装有永久性瘤胃瘘管的泌乳前期荷斯坦奶牛为试验动物,在精粗比为55：45的日粮条件下,采用In sacco法测定了不同加工处理的玉米在奶牛瘤胃内干物质和淀粉的降解规律.试验结果表明,不同加工处理方法可以程度不同的影响干物质和淀粉在瘤胃中的降解,与未处理玉米相比,挤压膨化和制粒加工处理能明显的提高玉米中淀粉在瘤胃中的降解率（P＜0.05）,压扁处理对干物质降解率无明显影响（P＞0.05）,但使淀粉瘤胃降解率明显提高（P＜0.05）,烘烤与焙炒加工处理能明显的降低玉米中干物质和淀粉在瘤胃中的降解率（P＜0.01）.淀粉瘤胃降解率由高到低顺序依次为膨化玉米＞颗粒玉米＞压扁玉米＞未处理玉米＞焙炒玉米＞140℃/35min烘烤玉米.淀粉在瘤胃中的降解的影响因素最主要是淀粉的快速可溶部分和降解速率.     

Nutrient composition and the use of solubility to estimate rumen degradability of food proteins in cattle

The use of protein solubility to estimate protein degradability in cattle was tested using 12 by‐products representing agricultural and distillers' food raw materials (RM). Agricultural RM included cereals (rice bran, maize gluten feed), legumes (field beans, soybean hulls) and oil‐seeds (rape meal, sunflower meal). Distillers' RM involved dark grains (DG1, DG2, DG3) and malts (MT1, MT2, MT3). Soluble proteins, as proportions of total soluble N (TSN), were extracted from RM and their undegraded residues (RU) after 18 h of in sacco rumen incubation in cattle (dg₁₈) by using water (albumin), salt (globulin), acid (glutalin 1) and alkali (glutalin 2) in succession. RM and RU differed significantly for all soluble proteins (P < 0.001). While albumin was the major protein in legume RM (0.74TSN), glutalin 1 was the major protein in legume RU (0.54TSN). Cereals were the most variable group, where maize gluten contained five times more albumin and two times more TSN in RM and three times more glutalin 1 + 2 in RU than those in rice bran. Distillers' RM contained slightly less albumin (0.53 vs 0.56TSN) but over twice as much glutalin 1 (0.29 vs 0.12TSN) as agricultural RM. Albumins were the most (0.55TSN, RM; 0.16TSN, RU) and glutalin 1 the least (0.21TSN, RM; 0.48TSN, RU) degradable proteins. There were moderate to strong correlations between protein solubility, dg₁₈ and acid detergent‐insoluble N (ADIN). ADIN correlated satisfactorily with glutalin 1 + 2 (r = ?0.55, RM and ?0.70, RU), TSN (r = ?0.75, RM and RU) and slowly degradable N, (SDN; r = ?0.70, all). TSN in all RM related well with SDN (r = 0.65) and dUN (r = 0.72). TSN in distillers' RM correlated closely with SDN (r = 0.91) and dUN (r = 0.96). Glutalin 1 in distillers' foods correlated extremely well (r = 0.94, RM and 0.93, RU) with dUN. Globulins correlated most consistently with SDN (r = 0.82, all; r = 0.77, agricultural; r = 0.92, distillers' RM). It is possible to predict protein degradability from the presence or absence of soluble proteins in various foods. While ADIN may be more suitable to predict DUN, globulins showed more potential to predict SDN in various foods. However, it may be necessary to be selective in choosing solvents for various foods. Further studies must validate this method by using a greater range of foods before suggesting its routine use for food evaluation. © 2001 Society of Chemical Industry     

Technical note: Methodological and feed factors affecting measurement of protein A,B, and C fractions,degradation rate,and intestinal digestibility of rumen-undegraded protein

When formulating dairy cow rations, characterization of protein in feeds requires estimation of protein degradation in the rumen and digestion in the intestine. The objective of this work was to evaluate experimental and feed-related factors that affect characterization using in situ, in vitro, or mobile bag techniques, of 0-h washout (A), potentially degradable (B), and undegradable (C) protein fractions, protein degradation rate (K_d), and digestibility of rumen undegradable protein (dRUP). Data sets of 136 studies on A, B, C, and K_d and 113 studies on dRUP were amassed from the literature. Mixed-effect linear models were used to relate these variables to methodological and feed factors while accounting for random differences among studies. Predictions of A, B, and C protein fractions were significantly influenced by crude protein and neutral detergent fiber interactions with bag pore size, incubation time, bag area, and sample-to-bag area ratio. For example, a 20.0% decrease in crude protein of a theoretical legume silage sample would increase A fraction prediction by 20.1%, but 34.7% with bag incubation time ?1 standard deviation below the mean. Similarly, reported K_d values were significantly influenced by crude protein interactions with bag area and sample-to-bag area ratio and by neutral detergent fiber interaction with pore size. Feed variables and measurement variables influencing protein digestibility measures suggest that these analytical factors are likely associated with variance among differing methodologies and within unique samples of the same feed. When predicting dRUP, the use of mobile bag method produced significantly different estimates compared with the in vitro 3-step method. The use of mobile bag resulted in an 8.9% (±3.8%) higher estimate of dRUP compared with the in situ technique. In 618 and 977 samples, sample variation to sample mean ratio for acid detergent fiber and pepsin-acid incubation time was 63.0 and 58.0%, respectively. Variation in feedstuff content and lack of standardization of methods used to measure protein disappearance led to a lack of robustness in the measurements commonly employed.     

Increased efficiency of wool growth and live weight gain in Merino sheep fed transgenic lupin seed containing sunflower albumin

The aim of this experiment was to assess, using sheep, the nutritive value of lupin seed transgenically modified to contain sunflower seed albumin. Eighty Merino wether sheep of mean live weight 32.3 kg were divided into two groups and fed 796 g dry matter (DM) day⁻¹ of a cereal hay‐based diet containing 350 g kg⁻¹ of either the transgenic or parent (unmodified) lupin seed for 6 weeks. Measurements were made of wool growth and live weight gain. After 6 weeks, half the sheep in each group were selected for a urine and faeces balance study in which organic matter (OM), nitrogen (N) and urinary purine metabolites were measured. Blood samples were taken from all sheep at the beginning and end of treatment and analysed for amino acids and plasma metabolites. A comprehensive chemical analysis of the grains showed that there was little difference between them in terms of most nutritional components, but the transgenic lupin seed contained a 2.3‐fold higher methionine concentration and 1.3‐fold higher cysteine than did the parent. There were no significant differences between grains in OM digestibility, rumen microbial protein synthesis or in sacco degradability of dry matter. Sheep fed the transgenic lupin grain had an 8% higher rate of wool growth (P < 0.01) and 7% higher live weight gain (P < 0.05) than sheep fed the parent grain. The sulphur (S) concentration of wool and the cysteine concentration of plasma were also higher in the sheep fed the transgenic lupin by 2.7% and 11.5% respectively (P < 0.05). Plasma methionine was increased by 10%, but the differences were not statistically significant (P > 0.1). Plasma urea N was lower in the sheep fed the transgenic grain than those fed the parent grain (6.5 vs 6.8 mmol l⁻¹, P < 0.05). The results show that genetic modification of a feed grain can improve its nutritive value for ruminants. The size and nature of the responses were consistent with the transgenic lupins providing more methionine to the tissues, a first‐limiting amino acid for sheep. © 2000 Society of Chemical Industry     

Technical note: Evaluation of a novel enzymatic method to predict in situ undigested neutral detergent fiber of forages and nonforage fibrous feeds

The purpose of this study was to optimize the conditions of a previously proposed enzymatic method used to estimate in situ undigested neutral detergent fiber (uNDF). We used a multi-step enzymatic approach, in which samples were first solubilized in NaOH solutions as a preincubation (PreInc) phase. After rinsing, samples were incubated (24 h at 39°C) in a buffered solution (pH 6) containing hemicellulase, cellulase, and Viscozyme L enzymes (Sigma-Aldrich s.r.l., Milan, Italy), followed by incubation (24 h at 39°C) in a buffered solution (pH 5) containing xylanase. Two sets of experiments were performed: a calibration trial (that tested different PreInc conditions on 9 selected forages) and a validation trial (that verified the results by testing multiple samples of 6 different forage types and a group of fibrous by-products). In the calibration trial, samples (300 mg in Ankom F57 filter bags; Ankom Technology Corp., Fairport, NY) were preincubated at 39°C in a 0.1 M NaOH solution for 90, 180, or 240 min, or in 0.2, 0.5, 1.0, or 2.0 M NaOH solution for 90 min. The results indicated that the best PreInc method, in terms of intra-laboratory repeatability and estimation of reference in situ values, was 90 min in a 0.2 M NaOH solution. Thus, we used this PreInc condition to determine enzymatic uNDF of 257 samples in the validation trial. Although the selected method generally had good accuracy in predicting in situ uNDF, inconsistencies were noted for certain forage types. Overall, when enzymatic uNDF was used to predict the in situ uNDF of all samples, the regression was satisfactory (intercept = 7.098, slope = 0.920, R² = 0.73). The regression models developed for alfalfa hays, corn silages, and small grain silages had also acceptable regression performances and mean square error of prediction (MSEP) values, and the main sources of MSEP variation were error due to incomplete (co)variation and random error. Even when R² values were >0.70, the MSEP value of the regression model for grass hays was 149.55, and that for nonforage fibrous feeds was 155.16. Although enzymatic uNDF partially overestimated the in situ uNDF, particularly in grass silages, the proposed procedure seems to be promising for accurately predicting in situ uNDF, because it generally had good repeatability and provided satisfactory estimates of in situ uNDF.     

Technical note: A simple back-mounted harness for grazing dairy cows to facilitate the sulfur hexafluoride tracer gas technique

We describe here a cattle harness to attach a gas collection vessel to facilitate the sulfur hexafluoride (SF₆) tracer gas technique. The harness consists of 2 major components: (1) a lightweight, robust body fabricated from an equine surcingle or lunge roller with padded thoracic trapezius pressure points, a bespoke shaping shaft for spine support, and adjustable buckles on both sides; and (2) an elastic flank-strap to prevent the harness from dislodging. The spine support consists of stainless steel laminated with carbon fiber. This support minimizes the contact area with the animal's skin, relieves the spine area of pressure, and creates free flow of ambient air below the platform, reducing sweat accumulation and hence preventing skin lesions. The harness weighs approximately 1.2 kg, allows for attachment of 2 gas collection vessels (animal and background sample), and is cost effective.     

非织造布购物袋热黏合接缝强力的测试方法探讨

超声波热黏合的非织造布接缝不同于普通机织物和针织物等的车缝接缝。运用正交试验设计方法,对纺粘法PP非织造布购物袋热黏合接缝断裂强力测试中的试样宽度、拉伸速率和隔距三个因素进行优化。试验结果表明,试样宽度对纺粘法PP非织造布热黏合接缝断裂强力的测试结果影响最大,并且接缝的断裂强力随着试样宽度的增加而增加。选用试样宽度为200mm、拉伸速率为50mm/min、隔距为100mm的组合所得出的测试结果能比较真实地反映实际的接缝断裂强力。     

Technical note: Effects of filter bags on neutral detergent fiber recovery and fiber digestion in vitro

Filter bags facilitate the measurement of amylase-treated neutral detergent fiber (aNDF) and in vitro (IV) undigested aNDF (uNDF) by eliminating the transfer of residues from beakers into filtration crucibles. The objectives of this study were to (1) determine effects of filter bags on recovery of aNDF and (2) evaluate effects of filter bags on IV uNDF. For study 1, 6 samples each of grass hay (GR), alfalfa (AL), and corn silage (CS) were selected. Large standard deviations (SD) of ash-free aNDF (aNDFom) for samples in each forage type indicated compositional diversity (15.1, 7.45, and 12.9% of DM for GR, AL, and CS, respectively), and starch SD for CS was 16.4% of DM. Samples were weighed into Berzelius beakers or filter bags [25-µm pores (F57) or 6-µm pores (F58); Ankom Technology, Macedon, NY] for measurement of aNDF and aNDFom. All samples were extracted with neutral detergent, thermostable α-amylase, and sodium sulfite, and then soaked in boiling water and then acetone. Residues from beakers were filtered through a sea sand–covered GF/D filter (Whatman, Marlborough, MA) in Gooch crucibles (CR). Filter bags were extracted in a pressurized chamber at 100°C. The aNDF values did not differ between F57 and CR, but F58 was greater than CR for CS and AL. For GR, F58 was greater than CR for aNDFom. For study 2, diverse samples with large SD of aNDFom (20.7, 7.45, and 12.9% of DM for GR, AL, and CS, respectively) were weighed as loose powder into medium bottles (LS) or F57 bags, which were weighted to prevent floating. Blended ruminal fluid from 3 steers fed a 30% aNDFom diet was used as inoculum. Three samples of 1 forage type were randomly assigned to 1 of 6 IV runs using both treatments (LS and F57), and 3 bottles of each sample–treatment combination were removed after 12 h and 2 were removed after 120 h to measure uNDF. For LS, residues were extracted as in study 1 for CR. For F57, bags were rinsed in cold water and extracted as described in study 1. After 12 h, uNDF of F57 was greater than LS in CS, AL, and overall types. Ash-free uNDF (uNDFom) after 12 h of F57 was greater than LS in CS and overall types. After 120 h, F57 was greater than LS for uNDF of CS, but no differences were detected for uNDFom. The SD of uNDFom, but not uNDF, was higher after 12 and 120 h for F57 compared with LS. From 6 to 96 h, overall gas production of F57 was less than LS, and F57 was less than LS for CS from 3 to 96 h. Overall, LS gave greater maximum and faster rates of gas production than F57, as did AL and CS, but lag did not vary. Results indicate that filter bags affected aNDF and aNDFom measurement and inhibited fermentation for some materials.     

Technical note: Methodological and feed factors affecting prediction of ruminal degradability and intestinal digestibility of essential amino acids

We hypothesized that ruminal degradability of essential AA (EAA) and the intestinal digestibility of the ruminally undegraded EAA residue in feeds could be evaluated in a meta-analysis. The objective was to characterize methodological factors for ruminal incubation (time of incubation of feed in situ) and method of simulating digestion of the ruminally undegraded AA (incubation of residue in digestive enzymes in vitro or in mobile bags inserted into the duodenum). To increase numbers of observations, feeds were categorized before ANOVA. An approach is described to predict differential ruminal degradability (or undegradability) of individual EAA by normalizing them as a proportion of total AA (TAA) degradability (undegradability) and similarly to normalize the intestinal digestibility of EAA using TAA. Interaction of feed category with individual EAA justifies future studies with a broader range of feeds and more replication within feed to bolster this approach. With broader data, the approach to normalize EAA as a proportion of TAA should allow a better defined EAA library to be integrated with more robust CP databases (that can be updated with specific feed information from more routine laboratory analyses) in dairy supply-requirement models.     

In situ degradability of conventional and unconventional starch sources for ruminants,and factors determining their washable fraction: methodological implications

BACKGROUND: The in situ technique (IS) is used for characterising, screening and evaluating feedstuffs in ruminants. However, it is often not adapted to the particular characteristics of feeds (i.e. kinetics of starchy feeds with a standard framework used in forage). This may lead to potential biases in the final conclusions. In two successive experiments, we evaluated the degradative characteristics of conventional (CON) and unconventional (UNC) starchy feedstuffs (ING) and factors affecting their washable fractions (WF). The suitability of IS was then assessed. RESULTS: Two well‐defined ruminal fermentation patterns (CON and UNC) were observed. The WF and insoluble washable (ISWF) fractions were affected by ING, state of presentation [WAY, fresh (F) or pre‐dried (D)], particle size (PSI) and their interactions. The UNC and F feeds had greater WF and ISWF than CON and D, respectively. Increasing PSI linearly reduced WF and its proportion of ISWF. CONCLUSION: The PSI and WAY are critical factors to consider when designing experiments for the evaluation of starchy feedstuffs for ruminants using IS. It is still very risky to propose ‘standard’ parameters as this will always depend on the particular ING evaluated. Conducting pre‐evaluation tests before implementing each research protocol could help to refine the procedure. Copyright © 2009 Society of Chemical Industry     

Technical note: a rapid lipid separation method for determining fatty acid composition of milk

A rapid method for milk lipid separation followed by transmethylation to produce fatty acid methyl esters from bovine milk samples is presented. Fat is separated by a nonsolvent method using centrifugation. The method was compared with the popular hexane:isopropanol solvent extraction method, and fatty acid proportions were statistically identical for both methods. In 108 replicates, variance accounted for by using the 2 methods was of a similar magnitude to variance due to repeat separations or repeat injections onto the gas chromatography column. It is concluded that the proposed method is accurate, simple, rapid, safe, economical, and especially suitable for large numbers of samples.     

Technical note: A comparison of methods to determine pH in silages

The techniques used to assess pH in silages vary greatly. The aim of this study was to evaluate the effects of water-to-sample ratio, extraction procedure, and standing time on pH determination. Silage samples (n = 20 for each silage) were chosen to represent diverse crops (corn, elephant grass, sugarcane, and forage peanut) to have a varied ensilability index and thus a wide range in final pH. Three water-to-sample ratios and 2 extraction procedures were used to measure pH at 0, 5, 10, 15, 30, 60, and 120 min of standing time. The ratios (undried silage to water) were 9:60, 25:100, and 30:270. The samples with the first 2 ratios were manually extracted, using a glass beaker and a glass stirring rod. The samples with the 30:270 ratio were extracted by using a stomacher blender for 4 min at 200 rpm. An electrode was used to perform pH readings. Dry matter (DM), water-soluble carbohydrates, and lactic acid concentrations were determined. The experimental design was completely randomized using a mixed repeated-measures model. Mean separation was performed using the Tukey test at P < 0.10 using the MIXED procedure of SAS (SAS Institute Inc., Cary, NC). The DM concentrations ranged from 24.5 to 40.2, 15.8 to 25.9, 26.9 to 30.6, and 17.8 to 21.4% for corn, elephant grass, sugarcane, and forage peanut silages, respectively. The lactic acid concentrations ranged from 2.9 to 10.1, 1.8 to 4.4, 0.7 to 11.4, and 0.3 to 1.4% of DM for corn, elephant grass, sugarcane, and forage peanut silages, respectively. The pH values measured by the 9:60 method were greater than other techniques at any standing times. The pH values from the 25:100 and 30:270 methods did not differ for elephant grass and forage peanut silages at any standing times. However, the 30:270 method had greater pH values for corn and sugarcane silages than the 25:100 technique at any standing times. The pH values measured by the 30:270 method did not vary for any of the silages according to standing time. Nevertheless, the pH values of the 25:100 method were greater from 0 to 10 min than for other standing times for all silages. Thus, for this method, there was pH stabilization from 15 min of standing time on. Overall, the 25:100 and 30:270 methods are recommended for assessment of silage pH. Fifteen minutes of standing time should be used for the 25:100 method but the pH reading can be performed immediately after mixing for the 30:270 technique.     

Technical note: estimating statistical power of mixed models used in dairy nutrition experiments

Statistical power is defined as the probability of correctly rejecting the null hypothesis. Power calculations may be useful in planning experiments. The objective of this technical note is to outline an applied method that estimates statistical power of a dairy nutrition experiment that employs a Latin square as the experimental design. Because the SAS MIXED procedure (PROC MIXED) is commonly used to analyze data sets, this note outlines basic programming procedures that may be used to estimate statistical power of a mixed model using this procedure.     

Technical note: assessment of recovery site of mobile nylon bags for measuring ileal digestibility of starch in dairy cows

The objective of this study was to evaluate recovery site of mobile nylon bags for measuring ileal digestibility of ruminally undegraded starch in dairy cows. Eight feed samples of untreated and treated concentrates were examined. Three lactating cows equipped with rumen fistula and duodenal and ileal cannulas were used in the experiment. The mobile nylon bags containing intact feeds or residues after a 12-h ruminal incubation were pretreated using a 2-step procedure to simulate abomasal digestion before insertion through the duodenal cannula. To assess the effect of hindgut fermentation on starch digestibility, approximately half of the bags were collected from the ileum and half from the feces. The results indicate that feed samples should be preincubated in rumen before insertion into duodenum, and that samples with relatively high fractions of rumen-undigestible starch should be collected from the ileum instead of from feces.     

Technical note: An in vivo method to determine kinetics of unsaturated fatty acid biohydrogenation in the rumen

Rumen microbial biohydrogenation (BH) of unsaturated fatty acids (UFA) has been extensively studied in vitro; however, in vitro BH pathways, rates, and extents may not parallel those in vivo. The objective was to develop an assay to assess in vivo rates, pathways, and extent of BH of oleic (OA), linoleic (LA), and α-linolenic (ALA) acids. Each UFA was characterized in a separate experiment, each using 4 ruminally cannulated lactating Holstein cows. A single bolus consisting of 200 g of a UFA-oil [experiment 1 (EXP1): 87% OA sunflower, experiment 2 (EXP2): 70% LA safflower, and experiment 3 (EXP3): 54% ALA flaxseed] and 12 g of heptadecanoic acid (C17:0) was mixed into the rumen through the fistula. Rumen digesta was collected at ?1, ?0.25, 0.1, 0.25, 0.5, 0.75, 1, 1.5, 2, 3, 4, and 6 h relative to the bolus. Overall, the triglyceride boluses increased total fatty acids (FA) in the rumen from 3.9 (standard deviation = ±1.4) to 7.3% (±1.4) of rumen dry matter and enriched C17:0 from 0.4 (±0.1) to 2.5% (±0.5) of FA. The bolus enriched OA from 8.9 (±1.0) to 30.1% (±4.6) of FA in EXP1, LA from 11.1 (±1.8) to 35.9% (±5.0) of FA in EXP2, and ALA from 2.1 (±0.1) to 19.8% (±4.3) of FA in EXP3. The disappearances of C17:0, OA, LA, and ALA were fit to a single exponential decay model. The first-order rate of C17:0 rumen disappearance (turnover) was 9.1, 6.9, and 5.2%/h in EXP1, EXP2, and EXP3, respectively, and was used as a marker of FA passage. The rate of total rumen turnover of OA was 54.1%/h, LA was 60.5%/h, and ALA was 93.0%/h in EXP1, EXP2, and EXP3, respectively. Rumen concentration of all 3 UFA reached prebolus concentrations within 4 h. The calculated extent of lipolysis and initial isomerization was 85.6% for OA, 89.8% for LA, and 94.7% for ALA in EXP1, EXP2, and EXP3, respectively. Assuming that BH equals total disappearance minus passage, the rates of lipolysis and initial isomerization were 45.0, 53.6, and 87.8%/h for OA, LA, and ALA in EXP1, EXP2, and EXP3, respectively. Analysis of the data using compartmental modeling showed that the normal BH pathways proposed in the literature explained 46.0, 37.3, and 49.8% of the BH of OA, LA, and ALA in EXP1, EXP2, and EXP3, respectively. Based on the model, BH of trans C18:1 FA was the rate-limiting step to complete BH. Importantly, oils were provided as triglycerides and the reported rates represent the rate of lipolysis and BH. In conclusion, the rate of ruminal BH of OA, LA, and ALA was higher than that commonly observed in vitro, but the extent of BH was near expected values. The method developed provides a potential in vivo assay of ruminal BH for use in future experiments and modeling efforts.     

Technical note: High-throughput method for antifungal activity screening in a cheese-mimicking model

In this study, we developed a high-throughput antifungal activity screening method using a cheese-mimicking matrix distributed in 24-well plates. This method allowed rapid screening of a large variety of antifungal agent candidates: bacterial fermented ingredients, bacterial isolates, and preservatives. Using the proposed method, we characterized the antifungal activity of 44 lactic acid bacteria (LAB) fermented milk-based ingredients and 23 LAB isolates used as protective cultures against 4 fungal targets (Mucor racemosus, Penicillium commune, Galactomyces geotrichum, and Yarrowia lipolytica). We also used this method to determine the minimum inhibitory concentration of a preservative, natamycin, against 9 fungal targets. The results underlined the strain-dependency of LAB antifungal activity, the strong effect of fermentation substrate on this activity, and the effect of the screening medium on natamycin minimum inhibitory concentration. Our method could achieved a screening rate of 1,600 assays per week and can be implemented to evaluate antifungal activity of microorganisms, fermentation products, or purified compounds compatible with dairy technology.     

Prediction of apparent heal protein digestibility in pigs with a two-step in-vitro method

The use of a two-step in-vitro method to predict the in-vivo ileal digestibility of proteins in pigs was investigated. It proved not possible to predict accurately the ileal protein digestibility with the in-vitro method. By dividing the samples into groups of closely related products, a good relationship (r² = 0.93) between in-vivo and in-vitro data was only obtained for wheat products, where only five samples were analysed. For beans, peas, rapeseed products and soya bean products it was still not possible to predict the in-vivo protein digestibility (r² = 0.03-0.60). The in vivo-in vitro relationship was mainly determined by the properties of the proteins and the presence of antinutritional factors, such as lectins and trypsin inhibitors. The first influences both the in-vitro and in-vivo protein degradability and the latter only reduces the in-vivo degradability by stimulating the secretion of endogenous protein. It is suggested that, with the in-vitro method, real ileal digestibility of proteins is predicted. The apparent ileal protein digestibility can only be predicted with the in-vitro method after making corrections for the influence of these antinutritional factors on the secretion of endogenous protein. Possibly corrections are also needed for microbial protein, and protein which is solubilised in the small intestine but not absorbed because of the physical state of the chyme.