Plasmin persists in many dairy products, leading to undesirable quality defects. High pressure jet (HPJ) processing may be a potential method to reduce the plasmin (PL) activity due to its strong shear and high pressure. However, the impact of HPJ processing on the activity of PL-system remains unknown. In the study, the activities of PL and plasminogen (PG) in raw milk treated with HPJ were evaluated. PL activity increased from 127.24% to 180.21% in milk treated HPJ at 100-300MPa for single-cycle. Meanwhile, HPJ processing increased the PG activators (PA) activity and reduced the PA inhibitors (PAI) activity, suggesting that the increase of PL activity was caused by the conversion of PG to PL. Furthermore, milk treated with HPJ at 200 MPa or 300 MPa for three cycles can substantially decrease the activities of PL and PG. The initial TPC values in milk decreased from 1.37 × 104 CFU/mL to 8.00 × 101 and 2.00 × 101 CFU/mL. Overall, these results suggested that increasing the cycle number is an effective method to inhibit PL activity and achieve sterilization, which provided preliminary mechanistic insights into the effects of HPJ treatment on the PL-system in milk.
ABSTRACT: Studies with sheep and cattle have identified the macroalga Asparagopsis taxiformis (AT) as a potent methane-mitigating feed additive but have also raised concerns related to bromoform's (a halogenated compound responsible for the antimethanogenic effect of AT) stability over prolonged storage. Previously, our laboratory conducted a study in which the effect of AT on enteric CH4 diminished over time. This effect had not been previously reported and warranted further investigation. The study was conducted with 18 Holstein cows in a replicated 3 × 3 Latin square design with 28-d periods. Treatments were control (basal diet), basal diet + 0.50% AT, and basal diet + 0.75% AT (feed DM basis). Methane emission was measured using the GreenFeed system. At the beginning of the experiment, bromoform concentration in AT was 4.41 mg/g DM. Daily CH4 emission decreased up to 30% with increasing AT dose. Inclusion of AT also decreased DMI by up to 22%, resulting in no effect on CH4 yield (g CH4/kg DMI). Milk true protein and lactose concentrations decreased up to 4% and 2% by AT, respectively. Asparagopsis taxiformis decreased milk fat yield up to 13%. Milk true protein and lactose yields were decreased up to 15% and 14% by AT, respectively, resulting in a decrease in ECM of up to 14%. Methane intensity (g CH4/kg of ECM) decreased by AT up to 18%. Additionally, there was a decrease in BW in cows fed AT, compared with the control. In conclusion, AT stored over 20 mo and included at up to 0.75% (feed DM basis) in the diet of dairy cows decreased daily CH4 emission but did not affect CH4 yield and decreased DMI and milk yield in mid-lactation dairy cows. These data confirm previous observations of inconsistent effects of AT on CH4 emission metrics and negative effects of AT on feed intake and productivity.
ABSTRACT: Colostrum quality is influenced by multiple factors, including its microbial load, which is determined by the cleanliness of collection and storage conditions. Additionally, the dam's diet and immune status play a crucial role in shaping colostrum quality by affecting IgG concentrations. Whereas many factors contribute to colostrum quality, this study will specifically use IgG content as the primary measure of quality. It is well established that feeding low-quality colostrum negatively affects calf health and growth, whereas feeding good-quality colostrum leads to better outcomes. However, it remains unclear if feeding colostrum above the recommended quality threshold offers additional benefits for calf health and growth. This study compared the effects of adequate- versus high-quality colostrum on dairy heifer calf growth, health, and the development of the fecal microbiome during the first 15 wk of life. We also measured the levels of apparent efficacy of absorption of IgG in both groups. Colostrum quality was initially determined and measured before feeding using a Brix refractometer (only feeding a minimum of 21% Brix); 93 heifer calves were assigned to treatment based on this data. Subsequently, laboratory analyses were conducted using radial immunodiffusion assays to measure the exact IgG levels in the colostrum; 72 calves were selected for continuation in the experiment based on the highest (n = 36) and lowest (n = 36) IgG levels in the colostrum. For these 72 calves, laboratory analyses were conducted using radial immunodiffusion assays to measure the exact IgG levels in the colostrum at the point of feeding and in the calf serum at 24 h after feeding. To ensure a fair comparison, the groups were balanced for calf birthweight, breed, and dam parity. Serum IgG at 24 h, weight, and health data were analyzed in SAS using linear mixed models and logistic regression. Alpha and β diversity were analyzed using R with ANOVA, permutational multivariate analysis of variance (PERMANOVA), and Benjamini-Hochberg P-value adjustments. Calves fed colostrum with a high IgG content (123.0 mg/mL IgG) exhibited higher rates of passive transfer compared with those fed adequate-quality colostrum (85.2 mg/mL IgG). Both groups had passive transfer rates >23 mg/mL IgG. Health outcomes were similar between the 2 groups, and ADG during the preweaning period was comparable, with calves gaining an average of 0.62 kg/d. Measures of α and β diversity in the fecal microbiome showed similar development in both groups. Apparent absorption efficacy was lower in calves fed high-quality colostrum (24.9%) compared with those fed adequate-quality colostrum (29.3%). The findings of this study support current recommendations for colostrum quality and suggest that calves may have a limited capacity to absorb higher concentrations of IgG. Whereas feeding higher-quality colostrum did not lead to significant improvements in growth, health, or microbiome diversity, it demonstrated that adequate-quality colostrum can be equally effective when combined with best practice management. Further research is needed to better understand the relationships between immunoglobulin absorption efficiency, calf health, microbiome development, and growth performance.
ABSTRACT: There is an urgent need for a rapid and sensitive method for the detection of Escherichia coli O157:H7, a class of hazardous foodborne pathogens in food safety. The traditional ELISA, a dominant rapid detection technique, has the disadvantage of low test sensitivity due to the insufficient enzyme loading capacity. In this study, we successfully synthesized self-assembled Au/polydopamine (PDA)/horseradish peroxide (HRP) nanocomposites with high enzyme loading on the outer surface and in the inner space. The high catalytic activity of Au/PDA/HRP was maintained by virtue of its hyperbranched flexible structure. For E. coli O157:H7 detection in milk samples, the proposed immunoassay achieved a visual cut-off value of 103 cfu mL−1 and a low limit of detection of 2.8 × 102 cfu mL−1, which are 33 and 46 times more sensitive than the traditional ELISA, respectively. The tremendous advantages of high sensitivity, excellent specificity, and adequate recovery make it promising for monitoring various kinds of pathogenic bacteria in food safety with greater sensitivity.
Hongqing Hu, Sébastien Franceschini, Pauline Lemal
et al.
ABSTRACT: Negative energy balance (NEB) during early lactation is a critical physiological challenge in high-producing dairy cows, affecting both their health and production performance. The objectives of this study were: (1) to compare the genetic architecture of logit-transformed predicted NEB (LPNEB), a logit-transformed novel energy deficiency score (LEDS), 15 biomarkers, and 3 production traits using SNP-based genomic correlation analysis; (2) to extend this study to a chromosomal level to identify specific genomic regions involved in the regulation of energy metabolism; and (3) to compare the independent contributions of 8 traits to the underlying genetic architecture of LPNEB and LEDS. The SNP effects estimated from single-trait models can be used to quickly calculate genomic correlations for 20 traits. The results indicate strong genomic correlations between LPNEB and LEDS, as well as with key metabolic biomarkers, particularly blood nonesterified fatty acids (NEFA), highlighting their importance in energy metabolism. Furthermore, NEFA was a strong independent contributor to both LPNEB and LEDS. Chromosome regions located on BTA19 and BTA25 were identified as potentially associated with NEB. By combining genomic correlation and contribution analyses, this study provides valuable insights into the genetic basis of NEB and related traits in dairy cows.
ABSTRACT: The objective of this study was to evaluate the possible role of the peroxisome proliferator-activated receptors (PPAR: PPAR-α, PPAR-β/δ, and PPAR-γ) in diet and CLA-induced milk fat depression (MFD) in dairy cows. We hypothesized that the expression of PPAR, which regulate lipid metabolism and bind to PUFA, could be modulated by biohydrogenation intermediates that induce MFD, thereby interfering with milk fat synthesis. First, tissue profiling revealed that PPAR-α and PPAR-β/δ had low expression in mammary tissue compared with the liver. A comparison of lactating and nonlactating tissue from the same cows showed that expression of all 3 PPAR isoforms did increase during lactation. Mammary expression of the PPAR family during MFD was then observed in 9 mid-lactation cows in a 3 × 3 Latin square design with MFD induced by a 3-d intravenous infusion of trans-10,cis-12 CLA or feeding a high-oil and low-forage diet. The expression of all 3 PPAR isoforms remained largely unaltered during CLA and diet-induced MFD, except for an increase in PPAR-α target genes CPT1A and ACADVL that are involved in β-oxidation. The interaction of PPAR-γ chemical agonist troglitazone and antagonist T0070907 and CLA was then investigated in bovine mammary epithelial cells. The activation and inhibition of PPAR-γ did not overcome trans-10,cis-12 CLA inhibition of lipogenesis despite the agonist stimulating PPAR-γ expression. Furthermore, PPAR-γ activation did not modify the expression of lipogenic genes. Overall, the results fail to support a functional role of the PPAR family in the inhibition of lipogenesis during MFD in dairy cows.
The growing global demand for minimally processed fresh foods has driven the development of innovative food processing methods. High-pressure processing (HPP), a leading non-thermal technique, has proven to be environmentally friendly, cost-effective, and versatile across various food types. Its ability to extend shelf life and ensure microbial safety is well established, particularly in dairy products. However, further research is needed to fully understand HPP’s impact on improving the digestion of milk compounds and assessing potential safety risks. Recent studies, for example, demonstrate HPP’s efficacy in enhancing the microbial safety and nutritional value of products like cheese, yogurt, and whey protein isolate. This review explores these advancements and highlights HPP’s role in improving the physicochemical, organoleptic, and functional properties of milk and dairy products.
Ziad W. Jaradat, Sally Abulaila, Ekhlas Al-Rousan
et al.
Escherichia coli O157:H7 is a pathogen associated with severe diarrhoeal diseases that lead to hemorrhagic colitis, hemolytic uremic syndrome, and death, especially in immunocompromised patients. The source of this bacterium is mainly beef cattle in which the pathogen seeps into the food chain from improper hygienic practices either during the slaughtering of the animal or during the after-processing. In addition, this pathogen gets introduced into other parts of the food chain such as vegetables and fruits, as well as dairy products. This document presents the first review of articles published on the prevalence of E. coli O157:H7 in foods in the MENA region in the last 20 years. Indeed E. coli O157:H7 was isolated from many foods of plant and animal origin. The presence of E. coli O157:H7 in meat products necessities good cooking procedures for these products to eliminate the pathogen and prevent foodborne illnesses. In addition, the presence of the pathogen in fruits and vegetables which are consumed mainly without cooking indicates a high risk of E. coli O157:H7 foodborne outbreaks. Thus, estimating the prevalence of this pathogen in these types of food will certainly help health authorities impose good agricultural and manufacturing practices to minimize the risk of such outbreaks. In this review, Google scholar and PubMed were used as the main search engines for our literature search using the following key words: Prevalence—E. coli O157:H7—Food—MENA—Meat—Chicken—Dairy products—Beef etc. and the names of the countries included in the MENA region were used as keywords.
ABSTRACT: The objective of this cross-sectional study was to determine associations between calf management practices, the number of antimicrobial treatments, and antimicrobial resistance in preweaning heifers on Canadian dairy farms. A composite of 5 fecal samples from preweaning calves was collected from 142 dairy farms in 5 provinces and analyzed for phenotypic antimicrobial susceptibility with the microbroth dilution method. Questionnaires were used to capture herd characteristics and calf management practices used on the farm. Calf treatment records were collected during the farm visits. Escherichia coli was isolated from all 142 fecal samples with the highest resistance to tetracycline (41%), followed by sulfisoxazole (36%), streptomycin (32%), chloramphenicol (28%), ampicillin (16%), trimethoprim-sulfamethoxazole (15%), ceftriaxone (4.2%), cefoxitin (2.8%), amoxicillin-clavulanic acid (2.1%), ciprofloxacin (2.1%), nalidixic acid (2.1%), azithromycin (1.4%), and gentamicin (1.4%). Multidrug resistance was observed in 37% of E. coli isolates. Three-quarters of farms used fresh colostrum as the most common type of colostrum fed to calves. Colostrum quality was checked on 49% of farms, but the transfer of passive immunity was only checked on 32% of farms in the last 12 mo. Almost 70% of farms used straw or hay or a combination as the bedding material for calves. Among the 142 farms, a complete set of calf records were collected from 71 farms. In a multivariable logistic regression model, farms with ≥1.99 to 32.57 antimicrobial treatments/calf-year were 3.2 times more likely to have multidrug resistant E. coli in calf feces compared farms with <1.99 antimicrobial treatments/calf-year. Farms using hay or straw beddings were 5.1 times less likely to have multidrug resistant E. coli compared with those with other bedding materials including shavings or sawdust. Bedding management practices on farms may need to be investigated to reduce the potential effect on disseminating multidrug resistant bacteria.
Matthias Schilde, Dirk von Soosten, Jana Frahm
et al.
Methanogenesis in ruminants contributes to both greenhouse gas emissions and feed energy losses whereby the latter becomes specifically important in energy-deficient periparturient cows. It was hypothesized that increased concentrate feed proportions (CFP) and feeding with the methane inhibitor 3-nitrooxypropanol (3-NOP), as well as their potential synergism, improve the energy status of peripartal cows. Periparturient dairy cows were fed low or high dietary CFP either tested without or combined with 3-NOP. The GreenFeed system was used to calculate the metabolic respiration quotient (RQ<sub>metabolic</sub>) and tissue energy retention (ER<sub>tissue</sub>) by methods of indirect calorimetry. The calorimetrically estimated ER<sub>tissue</sub> coincided with a conventionally calculated energy balance except for the antepartal period. Neither CFP nor 3-NOP affected the ultrasonographically assessed lipomobilization in adipose depots. In the group fed 3-NOP and a high concentrate feed proportion, the RQ<sub>metabolic</sub> significantly rose over the course of the experiment and the ER<sub>tissue</sub> was also increased. Serum non-esterified fatty acid concentrations were lower in the 3-NOP groups albeit ß-hydroxybutyrate (BHB) remained unaffected. Higher CFP reduced BHB and increased blood glucose levels. In conclusion, 3-NOP and high CFP improved the energy budget of the cows in an interactive manner, which was, however, not apparent in all of the examined parameters. The application of the GreenFeed system for indirect calorimetry is a promising approach, which needs further validation in the future.
ABSTRACT: The objective of this study was to compare 3-breed rotational crossbred (CB) cows of the Montbéliarde, Viking Red, and Holstein (HO) breeds with HO cows fed 2 alternative diets for dry matter intake (DMI), fat plus protein production (CFP), body weight (BW), body condition score (BCS), feed efficiency, and residual feed intake (RFI) from 46 to 150 days in milk (DIM) during first lactation. The CB cows (n = 17) and HO cows (n = 19) calved from September 2019 to March 2020. Cows were fed either a traditional total mixed ration diet (TRAD) or a higher fiber, lower starch total mixed ration diet (HFLS). The HFLS had 21% more corn silage, 47% more alfalfa hay, 44% less corn grain, and 43% less corn gluten feed than the TRAD. The 2 diets were analyzed for dry matter content, crude protein, forage digestibility, starch, and net energy for lactation. The BW and BCS were recorded once weekly. Daily milk, fat, and protein production were estimated from twice monthly milk recording with random regression. Measures of efficiency were CFP per kilogram of DMI and DMI per kilogram of BW. The RFI from 46 to 150 DIM was the residual error from regression of DMI on milk energy, metabolic BW, and the energy required for change in BW. Statistical analysis of all variables included the fixed effects of diet, breed group, and the interaction of diet and breed group. The CB cows fed HFLS had less DMI (−12%) and lower DMI/BW (−14%) compared with the HO cows fed TRAD. For CFP, CB and HO cows were not different when fed TRAD or HFLS. Furthermore, the CB cows fed HFLS had higher BW (+50 kg) compared with HO cows fed HFLS. The CB cows fed TRAD had higher BCS than HO cows fed TRAD and HO cows fed HFLS (+0.46 and +0.62, respectively). The HO cows fed TRAD had more DMI (+14%) and lower CFP per kilogram of DMI (−12%) compared with the HO cows fed HFLS. In addition, mean RFI from 46 to 150 DIM was lower and more desirable for CB cows fed HFLS (−120.0 kg) compared with HO cows fed TRAD (85.3 kg). Dairy producers may feed either TRAD or HFLS to CB cows without loss of CFP.
If validated for use in dairy cattle, interstitial continuous glucose monitors (CGMs) could be easily implemented, informative tools for research, clinical, and perhaps even on-farm applications. To evaluate their efficacy, 2 experiments were conducted, during which lactating Holstein cows were fit with indwelling jugular catheters, as well as FreeStyle Libre (FSL; Abbott) and Dexcom G6 (DexCom Inc.) CGMs secured either behind their polls, lateral to their ears, or beneath their pin bones on their upper rear legs. During the first experiment, blood (measured with a handheld glucometer) and interstitial glucose measurements were collected from 13 cows every 4 h for 96 h. In the second experiment, the same measurements were collected from 8 cows every 15 min for 6 h. At the mid-point of the sampling period (3 h), cows received a bolus dose of dextrose to facilitate comparisons across a broad range of glucose concentrations. Results from both experiments determined that functional longevity of the sensors was greatest for those sensors secured near the ear. Likewise, interstitial measurements from the ear sensors were most closely correlated with blood glucose concentrations (r = 0.82 and r = 0.71 for FSL ear and Dexcom G6 ear, respectively). Unfortunately, accuracy calculated as absolute relative error was low, at 60.7% or less. As a result of the low accuracy, even though both ear sensors detected an increase in glucose concentrations following the bolus dose, neither produced results exactly matching blood glucose measurements. The results of this work indicate that the FSL and Dexcom G6 CGMs are not currently capable of replacing blood-based glucose measurements.
Aya A. Muhammed Saeed, Alan A. Othman , Kaihan H. S. H Karim
et al.
The study of heavy metals in the human diet is important due to their dual effects as either essential or toxic to the human body. In this study, we determined the concentration of selected heavy metals in common potato and corn chips brands in Iraq markets. Ten common brands were selected and analyzed for their concentrations of the studied heavy metals by ICP-MS. The results shown that the average heavy metals level in potato and corn chips in (mg kg-1±SD) were 2.68±0.67 and 2.96±3.00 for Mn, 4.64±1.99 and 5.82±5.45 for Fe, 0.29±0.33 and 1.36±0.95 for Cu and 2.84±1.26 and 6.26±1.67 for Zn respectively. Corn chips were found to contain higher heavy metals than potato chips. The daily consumption of heavy metals from potato and corn chips (20 g daily intake) is lower than the recommended level set by the World Health Organization and Food and Agriculture Organization. Also, Daily intake of these metals is lower than the oral suggested amount and the upper tolerable daily intake set by the US. The contamination risk for the Iraqi population especially children from heavy metals exposure by this study performs to be non-significant, emphasizing the need to conduct more studies and confirm examining of heavy metals in foodstuffs especially chips brands.
Bulei Sheng, Søren D. Nielsen, Maria Glantz
et al.
ABSTRACT: Milk with different κ-casein (CN) phenotypes has previously been found to influence its gastric digestion rate. Therefore, the aim of the present study is to disentangle contributions of genetic variation and its related sialylation on the in vitro digestion process of κ-CN. Accordingly, κ-CN was purified from milk representing homozygous cows with κ-CN phenotypes AA, BB, or EE and used as substrate molecules in model studies using the INFOGEST 2.0 in vitro static digestion model. Furthermore, the effect of removal of the terminal sialic acids present on the O-linked oligosaccharides of the purified κ-CN A, B, and E protein variants were studied by desialylation enzymatic assays. The κ-CN proteins were purified by reducing anion exchange chromatography with purities of variants A, B, and E of 93.0, 97.1, and 90.0%, respectively. Protein degradations of native and desialylated κ-CN isolates in gastric and intestinal phases were investigated by sodium dodecyl sulfate-PAGE, degree of hydrolysis (DH), and liquid chromatography electrospray ionization mass spectrometry. It was shown that after purification, the κ-CN molecules reassembled into multimer states, which then constituted the basis for the digestion studies. As assessed by DH, purified variants A and E were found to exhibit faster in vitro digestion rates in both gastric and intestinal phases compared with variant B. Desialylation increased both gastric and intestinal digestion rates for all variants, as measured by DH. In the gastric phase, desialylation promoted digestion of variant B at a rate comparable with native variants A and E, whereas in the intestinal phase, desialylation of variant B promoted better digestion than native A or E. Taken together, the results confirm that low glycosylation degree of purified κ-CN promotes faster in vitro digestion rates, and that desialylation of the O-linked oligosaccharides further promotes digestion. This finding could be applied to produce dairy products with enhanced digestibility.
ABSTRACT: The objective of this study was to determine the effects of milk fat depression induced by supplementing conjugated linoleic acid (CLA; trans-10,cis-12 and cis-9,trans-11 CLA) or feeding a higher starch and oil-containing diet (HSO) on metabolic changes in dairy cows after calving. The main hypothesis was that the 2 strategies to decrease milk fat yield could have different effects on performance, energy balance (EB), and inflammatory status in early lactation. Thirty-three Nordic Red dairy cows were used in a randomized block design from 1 to 112 d of lactation and fed one of the following treatments: control (CON), CLA-supplemented diet, or HSO diet. Dry matter intake and milk yield were measured daily whereas milk composition was measured weekly throughout the experiment. Nutrient digestibility, EB, and plasma hormones and metabolites were measured at 3, 7, 11, and 15 wk of lactation in respiration chambers. The HSO diet led to lower intakes of dry matter, neutral detergent fiber, and gross energy compared with CON and CLA diets. The CLA diet and especially the HSO diet resulted in lower energy-corrected milk yield during the first 7 wk of lactation than those fed CON. The EB was numerically higher for HSO and CLA diets compared with CON at wk 3 and 7. Plasma glucose concentration was higher by the CLA diet at wk 3 and by the HSO diet from wk 3 to 15 compared with CON. Plasma nonesterified fatty acids were higher at wk 3 in the CON group (indicating more lipid mobilization) but decreased thereafter to similar levels with the other groups. The HSO-fed cows had higher plasma ceruloplasmin, paraoxonase, and total bilirubin concentrations in the entire experiment and showed the highest levels of reactive oxygen metabolites. These results suggest an increased inflammatory and oxidative stress state in the HSO cows and probably different regulation of the innate immune system. This study provides evidence that milk fat depression induced by feeding HSO (as well as CLA) decreased milk fat secretion and improved EB compared with CON in early lactation. The increase in plasma glucose and paraoxonase levels with the HSO diet may imply a better ability of the liver to cope with the metabolic demand after parturition. However, the negative effect of HSO on feed intake, and the indication of increased inflammatory and oxidative stress warrant further studies before the HSO feeding strategy could be supported as an alternative to improve EB in early lactation.
Yu-Hang Jiang, Jian-Ping Ying, Wei-Gang Xin
et al.
ABSTRACT: Phenyllactic acid (PLA) has been demonstrated to possess antibacterial activity and capacity to prolong food shelf life. However, studies on the performance of PLA in inhibiting Staphylococcus aureus and its effectiveness when applied to dairy products are largely lacking. Here, antibacterial activity (planktonic and biofilm states) of PLA against S. aureus CICC10145 (S. aureus_45) were investigated. The results showed that PLA inhibited growth of S. aureus_45 and formation of S. aureus_45 biofilm. Next, the antibacterial action target of PLA was uncovered from both physiological and phenotypic perspectives. The results showed that PLA decreased cell metabolic activity and cell viability, damaged cell membrane integrity, triggered leakage of intracellular contents (DNA, proteins, and ATP), and caused oxidative stress damage and morphological deformation of S. aureus_45. In practical application, the antibacterial activity of PLA against S. aureus_45 cells was further confirmed in skim milk and cheese as dairy food models, and the antibacterial effects can be adequately maintained during storage for 21 d, at least at 4°C. These findings suggested that PLA could be a potential candidate for controlling S. aureus outgrowth in dairy foods.
ABSTRACT: Milk preservative and freezing are used as strategies to prevent microbial growth and milk degradation, especially when immediate analytical processing is not feasible. The effects of the addition of preservative and freezing procedures have been investigated mainly in relation to milk gross chemical composition predicted through mid-infrared spectroscopy. This study aimed to determine whether different preservatives (i.e., no preservative, hydrogen peroxide, Bronopol, and Azidiol), freezing times (i.e., 0, 7, and 30 d), and temperatures of analysis (i.e., 5 and 21°C) influence the composition of milk protein fractions determined through reversed-phase HPLC. Bulk milk samples for the analysis of protein profile were collected from 5 commercial dairy farms. Data were analyzed with a linear mixed model, which included type of preservative, time of storage, temperature of analysis, and the interaction between type of preservative and time of storage as fixed effects, with the farm and the residual as random effects. Samples with no preservative had the greatest amount of all protein fractions, whereas Bronopol-preserved milk had the lowest amount. Increasing storage time under freezing conditions had a nonlinear detrimental effect on milk protein fractions. The temperature of analysis significantly contributed to the variation of κ-casein, β-casein, αS1-casein, β-lactoglobulin, and α-lactalbumin fractions. The z-scores were calculated to evaluate the similarity between detailed protein profile of fresh milk without preservative analyzed at 5°C and detailed protein profile of milk treated according to the tested conditions. Overall results suggested a good agreement between different analytical conditions. Still, short storage time under freezing conditions is recommended to avoid degradation of milk protein fractions and consequent analytical underestimation.