RESUMEN
Our objectives were to determine if milk casein as a percentage of true protein (CN%TP) estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) is equivalent to CN%TP estimated by Kjeldahl, and to determine the proportion of casein (CN), casein proteolysis products (CNPP), and serum protein (SP) from milk true protein (TP) that goes into the Kjeldahl noncasein nitrogen (NCN) filtrate and the proportion that stays in the NCN precipitate using SDS-PAGE. Raw milk samples were collected from 16 mid-lactation Holstein cows twice a week for 2 wk. These milks were analyzed for Kjeldahl total nitrogen, nonprotein nitrogen, and NCN content in duplicate, and by SDS-PAGE. The CN%TP determined by Kjeldahl was compared with the CN%TP estimated by SDS-PAGE calculated in 2 ways: as a percentage of only intact caseins divided by TP and as a percentage of both intact caseins and CNPP divided by TP. Three milks varying in fat, lactose, TP, CN, and SP content were formulated. These milks were analyzed in duplicate for Kjeldahl total nitrogen, nonprotein nitrogen, and NCN content, and each of the NCN filtrate and NCN precipitate were analyzed in duplicate by SDS-PAGE for relative quantity (%) of CN, CNPP, and SP. We found that the estimate of CN%TP by Kjeldahl was higher than the estimate of CN%TP by SDS-PAGE that was calculated as only intact CN divided by the total of all protein bands. However, no difference was detected in the estimate of CN%TP by Kjeldahl compared with CN%TP by SDS-PAGE when CNPP were included as CN in the calculation of SDS-PAGE results. Based on SDS-PAGE results, we found that a majority (89%) of the CNPP from the milk (approximately 10.13 out of 11.41% TP) were retained in the Kjeldahl NCN precipitate. Thus, CN%TP measured by Kjeldahl underestimates the amount of proteolytic damage that has been done to CN in milk. It is important for the dairy industry to correctly and rapidly measure the extent of proteolytic damage to milk protein to correctly value milk from a product quality and yield point of view. A rapid and quantitative measure of proteolytic damage to milk protein is needed.
Asunto(s)
Caseínas , Leche , Animales , Bovinos , Electroforesis en Gel de Poliacrilamida/veterinaria , Femenino , Leche/química , Proteínas de la Leche/análisis , Dodecil Sulfato de SodioRESUMEN
Our objective was to determine the effect of mid-infrared (MIR) homogenizer efficiency on accuracy and repeatability of Fourier transform MIR predicted fat, true protein, and anhydrous lactose determination given by traditional filter and partial least squares (PLS) prediction models. Five homogenizers with different homogenization performance based on laser light-scattering particle size analysis were used. Repeatability and accuracy were determined by conducting 17 sequential readings on milk homogenized externally to the instrument (i.e., control) and unhomogenized milk. Milk component predictions on externally homogenized milks were affected by variation in homogenizer performance, but the magnitude of effect were small (i.e., <0.025%) when milks were pumped through both efficient and inefficient homogenizers within a MIR milk analyzer. Variation in the in-line MIR homogenizer performance on unhomogenized milks had a much larger effect on accuracy of component testing than on repeatability. The increase of particle size distribution [d(0.9)] from 1.35 to 3.03µm (i.e., fat globule diameter above which 10% of the volume of fat is contained) due to poor homogenization affected fat tests the most; traditional filter based fat B (carbon hydrogen stretch; -0.165%), traditional filter-based fat A (carbonyl stretch; -0.074%), and fat PLS (-0.078%) at a d(0.9) of 3.03µm. Variation in homogenization efficiency also affected traditional filter-based true protein test (+0.012%), true protein PLS prediction (-0.107%), and traditional filter-based anhydrous lactose test (+0.027%) at a d(0.9) of 3.03µm. Effects of variation in homogenization on anhydrous lactose PLS predictions were small. The accuracy of both traditional filter models and PLS models were influenced by poor homogenization. The value of 1.7µm for a d(0.9) used by the USDA Federal Milk Market laboratories as a criterion to make the decision to replace the homogenizer in a MIR milk analyzer appears to be a reasonable limit, given the magnitude of effect on the accuracy of fat tests. In the future, as new PLS models are developed to measure other components in milk, the sensitivity of the accuracy of the predictions of these models to factors such as variation of homogenizer performance should be determined as part of the ruggedness testing during PLS model development.
Asunto(s)
Lactosa , Leche/química , Animales , Calibración , Análisis de los Mínimos Cuadrados , Proteínas de la Leche , ProteínasRESUMEN
Our objective was to develop partial least square models using data from Fourier transform mid-infrared (MIR) spectra to predict the particle size distributions d(0.5) and d(0.9), surface volume mean diameter D[3,2], and volume moment mean diameter D[4,3] of milk fat globules and validate the models. The goal of the study was to produce a method built into the MIR milk analyzer that could be used to warn the instrument operator that the homogenizer is near failure and needs to be replaced to ensure quality of results. Five homogenizers with different homogenization efficiency were used to homogenize pasteurized modified unhomogenized milks and farm raw bulk milks. Homogenized milks were collected from the homogenizer outlet and then run through an MIR milk analyzer without an in-line homogenizer to collect a MIR spectrum. A separate portion of each homogenized milk was analyzed with a laser light-scattering particle size analyzer to obtain reference values. The study was replicated 3 times with 3 independent sets of modified milks and bulk tank farm milks. Validation of the models was done with a set of 34 milks that were not used in the model development. Partial least square regression models were developed and validated for predicting the following milk fat globule particle size distribution parameters from MIR spectra: d(0.5) and d(0.9), surface volume mean diameter D[3,2], and volume moment mean diameter D[4,3]. The basis for the ability to model particle size distribution of milk fat emulsions was hypothesized to be the result of the partial least square modeling detecting absorbance shifts in MIR spectra of milk fat due to the Christiansen effect. The independent sample validation of particle size prediction methods found more variation in d(0.9) and D[4,3] predictions than the d(0.5) and D[3,2] predictions relative to laser light-scattering reference values, and this may be due to variation in particle size among different pump strokes. The accuracy of the d(0.9) prediction for routine quality assurance, to determine if a homogenizer within an MIR milk analyzer was near the failure level [i.e., d(0.9) >1.7µm] and needed to be replaced, is fit-for-purpose. The daily average particle size performance [i.e., d(0.9)] of a homogenizer based on the mean for the day could be used for monitoring homogenizer performance.
Asunto(s)
Análisis de Fourier , Leche , Animales , Tamaño de la Partícula , Valores de ReferenciaRESUMEN
Our objectives were to develop a method to produce milk somatic cell count (SCC) reference materials for calibration of electronic somatic cell count (ESCC) using gravity separation and to determine the effect of refrigerated storage (4°C) and freeze-thaw stability of the skim and whole milk SCC reference materials. Whole raw milk was high-temperature short-time pasteurized and split into 2 portions. One portion was gravity separated at 4°C for 22 h and the second portion was centrifugally separated to produce skim milk that was also gravity separated with somatic cells rising to the surface. After 22 h, stock solutions (low SCC skim milk, high SCC skim milk, high SCC whole milk) were prepared and preserved (bronopol). Two experiments were conducted, one to compare the shelf-life of skim and whole milk SCC standards at 4°C and one to determine the effect of freezing and thawing on SCC standards. Both experiments were replicated 3 times. Gravity separation was an effective approach to isolate and concentrate somatic cells from bovine milk and redistribute them in a skim or whole milk matrix to create a set of reference materials with a wider and more uniformly distributed range of SCC than current calibration sets. The liquid SCC reference materials stored using the common industry practice at 4°C were stable (i.e., fit for purpose, no large decrease in SCC) for a 2-wk period, whereas frozen and thawed reference materials may have a much longer useful life. A gradual decrease occurred in residual difference in ESCC (SCC × 1,000/mL) versus original assigned reference SCC over duration of refrigerated storage for both skim and whole milk SCC samples, indicating that milk ESCC of the preserved milks was gradually decreasing during 28 d of storage at 4°C by about 15,000 SCC/mL. No difference in the ESCC for skim milk was detected between refrigerated and frozen storage, whereas for whole milk the ESCC for frozen was lower than refrigerated samples. Future work is needed to determine the time and temperature of longer term frozen storage over which the SCC results are stable.