Lactoperoxidase potential in diagnosing subclinical mastitis in cows via image processing.

This report describes how image processing harnessed to multivariate analysis techniques can be used as a bio-analytical tool for mastitis screening in cows using milk samples collected from 48 animals (32 from Jersey, 7 from Gir, and 9 from Guzerat cow breeds), totalizing a dataset of 144 sequential images was collected and analyzed. In this context, this methodology was developed based on the lactoperoxidase activity to assess mastitis using recorded images of a cuvette during a simple experiment and subsequent image treatments with an R statistics platform. The color of the sample changed from white to brown upon its exposure to reagents, which is a consequence of lactoperoxidase enzymatic reaction. Data analysis was performed to extract the channels from the RGB (Red-Green-Blue) color system, where the resulting dataset was evaluated with Principal Component Analysis (PCA), Multiple Linear Regression (MLR), and Second-Order Regression (SO). Interesting results in terms of enzymatic activity correlation (R2 = 0.96 and R2 = 0.98 by MLR and SO, respectively) and of somatic cell count (R2 = 0.97 and R2 = 0.99 by MLR and SO, respectively), important mastitis indicators, were obtained using this simple method. Additionally, potential advantages can be accessed such as quality control of the dairy chain, easier bovine mastitis prognosis, lower cost, analytical frequency, and could serve as an evaluative parameter to verify the health of the mammary gland.

Assessment of pasteurisation of edible insects using enzymatic tests (activity of alkaline phosphatase and lactoperoxidase) applied in dairy products.

Industrialising edible insects goes along with quality control and hazard analysis and critical control points. One of those steps is assessing heat treatment. For the present contribution, the potential of enzymatic heat assessment tests used in the dairy industry (alkaline phosphatase and lactoperoxidase) to detect heat treatment in several insect species ( Acheta domesticus, Gryllus assimilis, Gryllus bimaculatus, Locusta migratoria, Schistocerca gregaria, Chilecomadia moorei, Galleria mellonella, Bombyx mori, Pachnoda marginata, Tenebrio molitor, Zophobas atratus, Apis mellifera, and Hermetia illucens) was evaluated. Insect material was homogenised, diluted, and the enzymatic tests (Lactognost®, Peroxtesmo®) were carried with these liquids as if they were milk. All species but C. moorei, B. mori, P. marginata, and A. mellifera showed alkaline phosphatase activity in raw samples and none in heated (10 min at 100 ℃) ones, while only G. mellonella, T. molitor, and Z. atratus reacted accordingly with lactoperoxidase. In trial 2 focusing only on alkaline phosphatase activity, inactivation of the enzyme after 5, 10, and 15 min of heating occurred species specific within a range of 60-86 ℃, i.e. within ordinary pasteurisation schemes. Thus and for the time being, heat treatment in many edible insect species can be assessed using alkaline phosphatase activity test kits. In contrast to milk samples, positive results may display bluish or greenish colours, and the time until a reliable reading is possible is extended to 1-1.5 h (24 h in the case of Gryllidae).

A proteomics study of colostrum and milk from the two major small ruminant dairy breeds from the Canary Islands: a bovine milk comparison perspective.

Colostrum and milk feeding are key factors for the newborn ruminant survival, affecting the future performance of the animal. Nowadays, there is an increasing interest in the potential of feeding newborn ruminants (mainly goat kids and lambs) with colostrum and milk from other more productive ruminant species (mainly cows). Although some studies regarding differences between colostrum and milk from these three species have been performed, herein we conduct for the first time a comparison using a proteomics 2-Dimensional Electrophoresis gel-based approach between these three ruminant species. In this study colostrum and milk samples from six Holstein cows, six Canarian sheep and six Majorera goats were used to determine the chemical composition, immunoglobulin G (IgG) and M (IgM) concentrations and proteomics profiles. Results showed that in general sheep colostrum and milk contained higher fat, protein and lactose percentages compared to bovine and goat samples. Additionally, no differences in the IgG or IgM concentrations were found among any of the three studied species, with the exception of sheep colostrum that showed the highest IgM concentration. With reference to the proteomics-based approach, some high abundant proteins such as serum albumin precursor, beta-caseins or different immunoglobulins components were found in colostrum, milk or even both. Nevertheless, differences in other proteins with immune function such as serotransferrin or lactoperoxidase were detected. This study shows that despite the similar immunoglobulin concentrations in colostrum and milk from the three studied species, differences in several immune components can be detected when these samples are studied using a proteomics approach. Finally, this study also provides a base for future investigation in colostrum and milk proteomics and metabolomics.

Set-up and application of an analytical approach for the quality control of purified colostrum as food supplement.

A validated analytical procedure is here described for the quality control of the protein fraction of purified bovine colostrum used in food supplements. The proposed procedure starts with 1D and 2D-gel electrophoresis. The sample is then separated into two fractions by protein G affinity chromatography: the IgG enriched and the IgG depleted fraction (IgG-d). A size exclusion chromatography coupled to UV is then applied to the IgG and IgG-d fractions for the quantitative analysis of IgG and IgM, respectively. The IgG-d fraction is then analysed by HPLC-MS analysis for the quantitative analysis of ß-lactoglobulins and α-lactoalbumin. The next step is to quantitatively measure a set of bioactive proteins selected from the bovine colostrum data bank on the basis of their claimed health benefits. The enzymatic activities of lactoperoxidase and xanthine dehydrogenase/oxidase are then tested as an index of protein functionality.

Change in viable bacterial count during preservation of milk derived from dairy cows with subclinical mastitis and its relationship with antimicrobial components in milk.

The objectives of the present study were to investigate the change in the number of viable pathogens during preservation of milk obtained from cows with subclinical mastitis and the association between the decreasing ratio of viable bacteria during preservation and the somatic cell count (SCC) and the values of lingual antimicrobial peptide (LAP), lactoferrin (LF) and lactoperoxidase (LPO). After preservation of milk at room temperature for 0, 0.5, 1, 2, 3, 4 and 5 hr, the bacterial colonies in the milk were counted to determine the number of colony forming units (CFUs). Fresh skim milk was used to determine the values of LAP, LPO and LF. Bacteria were not detected in 19.4% of milk samples, and this percentage increased up to 30% after 5 hr of preservation. The number of Staphylococcus aureus and Streptococcus uberis in milk did not change significantly during the 5-hr incubation, whereas significant decreases were observed in the number of coliforms, coagulase-negative staphylococci, yeasts and Corynebacterium bovis. High SCC significantly decreased CFUs of S. aureus and yeast after preservation of milk for 4 to 5 hr. High LF concentration in milk was associated with decrease in CFU of S. aureus during 4-hr preservation. These results suggest that the viable counts of some pathogens in milk decreased during preservation at room temperature after collection, which may be attributed to the leukocytes and antimicrobial components present in milk.

Development of a test strip for rapid detection of lactoperoxidase in raw milk.

Traditional methods for detecting lactoperoxidase (LP) are complex and time-consuming, so a test strip was made based on the enzymatic reaction principle to enable quick and convenient detection of LP in raw milk. In this study 0.1 mol/L citric acid (CA)/0.2 mol/L disodium hydrogen phosphate (NaP) buffer solution (pH 5.0), 22 mmol/L 3,3',5,5'-tetramethylbenzidine (TMB), 0.6 mmol/L hydrogen peroxide (H2O2), and 0.5% Tween-20 or 0.3% cetyltrimethyl ammonium bromide (CTAB) were optimal for preparing a quick, sensitive, and accurate LP test strip. The coefficient of variation (CV) of the estimated LP concentrations ranged from 2.47% to 6.72% and the minimum LP concentration detected by the test strip was 1-2 mg/L. Estimates of active LP in sixteen raw milk samples obtained using the test strip or the TMB method showed a good correlation (r=0.9776). So the test strip provides a quick, convenient, and accurate method for detecting the LP concentration of raw milk.

Raw milk from vending machines: Effects of boiling, microwave treatment, and refrigeration on microbiological quality.

In Italy, the sale of raw milk from vending machines has been allowed since 2004. Boiling treatment before its use is mandatory for the consumer, because the raw milk could be an important source of foodborne pathogens. This study fits into this context with the aim to evaluate the microbiological quality of 30 raw milk samples periodically collected (March 2013 to July 2013) from 3 vending machines located in Molise, a region of southern Italy. Milk samples were stored for 72 h at 4 °C and then subjected to different treatments, such as boiling and microwaving, to simulate domestic handling. The results show that all the raw milk samples examined immediately after their collection were affected by high microbial loads, with values very close to or even greater than those acceptable by Italian law. The microbial populations increased during refrigeration, reaching after 72 h values of about 8.0 log cfu/mL for Pseudomonas spp., 6.5 log cfu/mL for yeasts, and up to 4.0 log cfu/mL for Enterobacteriaceae. Boiling treatment, applied after 72 h to refrigerated milk samples, caused complete decontamination, but negatively affected the nutritional quality of the milk, as demonstrated by a drastic reduction of whey proteins. The microwave treatment at 900 W for 75 s produced microbiological decontamination similar to that of boiling, preserving the content in whey proteins of milk. The microbiological characteristics of raw milk observed in this study fully justify the obligation to boil the raw milk from vending machines before consumption. However, this study also showed that domestic boiling causes a drastic reduction in the nutritional value of milk. Microwave treatment could represent a good alternative to boiling, on the condition that the process variables are standardized for safe domestic application.

Lingual antimicrobial peptide and lactoferrin concentrations and lactoperoxidase activity in bovine colostrum are associated with subsequent somatic cell count.

The present study was undertaken to examine whether potential levels of innate immune factors (lingual antimicrobial peptide (LAP), lactoferrin (LF) and lactoperoxidase (LPO)) in colostrum are associated with subsequent milk somatic cell count (SCC) in dairy cows. Quarter milk samples were collected daily for 1 week postpartum to measure LAP and LF concentrations and LPO activity. SCC in milk was determined weekly for 2 months postpartum and its correlations to concentrations of LAP and LF and LPO activity were examined. Only small variations of all immune factors were found among four udders in each individual cow, whereas there were great differences in these factors among cows. Negative correlation was detected only between LPO activity and mean and maximum SCC, whereas its relationship was not significant. LAP and LF concentrations were significantly correlated positively to mean, maximum and minimum SCC. These results suggest that the great difference in innate immune factors among animals and high LAP and LF concentrations in colostrum may be associated with subsequent high incidence of SCC increase.

The impact of iron on the bleaching efficacy of hydrogen peroxide in liquid whey systems.

UNLABELLED: Whey is a value-added product that is utilized in many food and beverage applications for its nutritional and functional properties. Whey and whey products are generally utilized in dried ingredient applications. One of the primary sources of whey is from colored Cheddar cheese manufacture that contains the pigment annatto resulting in a characteristic yellow colored Cheddar cheese. The colorant is also present in the liquid cheese whey and must be bleached so that it can be used in ingredient applications without imparting a color. Hydrogen peroxide and benzoyl peroxide are 2 commercially approved chemical bleaching agents for liquid whey. Concerns regarding bleaching efficacy, off-flavor development, and functionality changes have been previously reported for whey bleached with hydrogen peroxide and benzoyl peroxide. It is very important for the dairy industry to understand how bleaching can impact flavor and functionality of dried ingredients. Currently, the precise mechanisms of off-flavor development and functionality changes are not entirely understood. Iron reactions in a bleached liquid whey system may play a key role. Reactions between iron and hydrogen peroxide have been widely studied since the reaction between these 2 relatively stable species can cause great destruction in biological and chemical systems. The actual mechanism of the reaction of iron with hydrogen peroxide has been a controversy in the chemistry and biological community. The precise mechanism for a given reaction can vary greatly based upon the concentration of reactants, temperature, pH, and addition of biological material. In this review, some hypotheses for the mechanisms of iron reactions that may occur in fluid whey that may impact bleaching efficacy, off-flavor development, and changes in functionality are presented. PRACTICAL APPLICATION: Cheese whey is bleached to remove residual carotenoid cheese colorant. Concerns regarding bleaching efficacy, off-flavor development, and functionality changes have been reported for whey proteins bleached with hydrogen peroxide and benzoyl peroxide. It is very important for the dairy industry to understand how whey bleaching can impact flavor and functionality of dried ingredients. Proposed mechanisms of off-flavor development and functionality changes are discussed in this hypothesis paper.

Lactoperoxidase activity in milk is correlated with somatic cell count in dairy cows.

Lactoperoxidase (LPO) is a milk protein with antimicrobial function. The present study was undertaken to examine the correlation between LPO activity and somatic cell count (SCC) in milk to use LPO activity as an indicator of mastitis. Composite milk of 36 cows and quarter milk of 3 cows were collected once per week from 0 to 300 d postpartum and twice per day for 1 wk, respectively. For the measurement of LPO activity, milk was mixed with tetramethylbenzidine solution and incubated at 37°C for 30 min, followed by the measurement of optical density. When only milk with low SCC (132±12×10(3) cells/mL) was used, a significant decrease in LPO activity was detected in primiparous cows from 0 to 4 mo postpartum. Lactoperoxidase activities of primiparous cows in mo 1, 2, and 3 postpartum were significantly higher than those in multiparous cows. When composite milk was divided based on LPO activity, the SCC was significantly higher in the groups with LPO activity >5 and from 3 to 3.9 U/mL in the second- and fourth-parity cows, respectively, compared with the group with LPO activity <2U/mL. Extremely high SCC were found in the ≥fifth-parity cows, even in low-LPO activity groups. In the case of quarter milk, higher LPO activity was associated with increased SCC in all 3 cows. The percentage of quarter milk samples with high SCC (4,062±415×10(3) cells/mL) increased with an increase in the LPO activity. The percentage of quarter milk samples with high SCC was 50.0 to 100% in the milk with LPO activity ≥5 U/mL. These results indicate that the correlation of LPO activity to the SCC in bovine milk may point to the potential use of the former as an indicator of SCC.

Integrated downstream processing of lactoperoxidase from milk whey involving aqueous two-phase extraction and ultrasound-assisted ultrafiltration.

The present work involves the adoption of an integrated approach for the purification of lactoperoxidase from milk whey by coupling aqueous two-phase extraction (ATPE) with ultrasound-assisted ultrafiltration. The effect of system parameters of ATPE such as type of phase system, polyethylene glycol (PEG) molecular mass, system pH, tie line length and phase volume ratio was evaluated so as to obtain differential partitioning of contaminant proteins and lactoperoxidase in top and bottom phases, respectively. PEG 6000-potassium phosphate system was found to be suitable for the maximum activity recovery of lactoperoxidase 150.70% leading to 2.31-fold purity. Further, concentration and purification of enzyme was attempted using ultrafiltration. The activity recovery and purification factor achieved after ultrafiltration were 149.85% and 3.53-fold, respectively. To optimise productivity and cost-effectiveness of integrated process, influence of ultrasound for the enhancement of permeate flux during ultrafiltration was also investigated. Intermittent use of ultrasound along with stirring (2 min acoustic and 2 min stirring) resulted in increased permeate flux from 0.94 to 2.18 l/m(2) h in comparison to the ultrafiltration without ultrasound. The use of ultrasound during ultrafiltration resulted in increase in flux, but there was no significant change in activity recovery and purification factor. The integrated approach involving ATPE and ultrafiltration may prove to be a feasible method for the downstream processing of lactoperoxidase from milk whey.

Alterations in the host defense properties of human milk following prolonged storage or pasteurization.

OBJECTIVES: Preterm infants are often fed pasteurized donor milk or mother's milk that has been stored frozen for up to 4 weeks. Our objectives were to assess the impact of pasteurization or prolonged storage at -20 degrees C on the immunologic components of human milk and the capability of the different forms of human milk to support bacterial proliferation. MATERIALS AND METHODS: The concentrations and activities of major host defense proteins in the whey fractions of mother's milk stored for 4 weeks at -20 degrees C or pasteurized human donor milk were compared with freshly expressed human milk. Proliferation of bacteria incubated in the 3 forms of human milk was assessed. RESULTS: Relative to freshly expressed human milk, the concentrations of lysozyme, lactoferrin, lactoperoxidase, and secretory immunoglobulin A were reduced 50% to 82% in pasteurized donor milk and the activities of lysozyme and lactoperoxidase were 74% to 88% lower (P < 0.01). Proliferation of bacterial pathogens in pasteurized donor milk was enhanced 1.8- to 4.6-fold compared with fresh or frozen human milk (P < 0.01). CONCLUSIONS: The immunomodulatory proteins in human milk are reduced by pasteurization and, to a lesser extent, by frozen storage, resulting in decreased antibacterial capability. Stringent procedure to minimize bacterial contamination is essential during handling of pasteurized milk.

Efficacy of enzymatic toothpastes for immobilisation of protective enzymes in the in situ pellicle.

AIM: Different enzyme-containing toothpastes are available on the market. The aim of the present in situ study was to investigate their efficacy for immobilisation of protective enzymes in the pellicle layer. METHODS: Pellicle formation took place in situ on bovine enamel slabs fixed to individual upper jaw splints carried by 6 subjects. After pellicle formation for 1 min, brushing was performed for 3 min with the commercially available toothpastes Enzycal, biotène and BioXtra, respectively. Before as well as 0, 20 and 40 min after brushing, samples were removed from the splints and tested for lysozyme, peroxidase and glucoseoxidase activity. The assays for the respective enzyme activities were based on fluorogenic substrates. Separate experiments were conducted for the different enzymes and toothpastes. RESULTS: Brushing with the toothpastes caused an extensive increase of glucoseoxidase activity in the pellicle, but it was of low tenacity whereas peroxidase activity was enhanced considerably. However, targeted accumulation of lysozyme in the pellicle was not very pronounced. Brushing without toothpaste had no effect on enzyme activities in the acquired pellicle. CONCLUSION: Targeted immobilisation of enzymes in the in situ pellicle can be achieved with toothpastes.

Antioxidants in colostrum and milk of sows and cows.

On account of the oxidative stress conditions that may appear during parturition, colostrum should provide with not only nutritional and immunological components but also antioxidative protection of newborn. There is evidence that apart from well-known antioxidative enzymes like glutathione peroxidase, superoxide dismutase, catalase or low molecular antioxidants, proteins like lactoperoxidase (LPO), lactoferrin (LF) and ceruloplasmin (CP) may exert antioxidative properties in colostrum. The aim of present study was to determine and to evaluate LPO, LF and CP activities in colostrum and milk of sows and cows. Samples were collected from 16 healthy cows five times: immediately after parturition, 12, 24 and 48 h, and 7 days postpartum as well as from 14 healthy sows five times: immediately after parturition, 6, 12, 24 and 36 h later. Examined parameters were determined spectrophotometrically at 412, 560 and 540 nm respectively. LPO activity was higher in sows as in cows and increased significantly within examined time. LF ability to inhibit superoxide radical generation was higher in sows as in cows and increased significantly within examined time. CP oxidase activity was higher in cows as in sows and decreased significantly during experimental period. In conclusion, antioxidative defence system in colostrum shows dynamic changes that allow for providing with necessary protection from oxidative stress conditions, which may appear after parturition.

Probiotic bacteria affect the composition of salivary pellicle and streptococcal adhesion in vitro.

INTRODUCTION: The use of probiotic bacteria is increasing worldwide and at least some of them can transiently colonize the oral cavity. Several studies have shown that probiotic bacteria, which are often thought of in relation only to intestinal health, can also affect the oral ecology, but the mechanisms for this are largely unknown. The aim of this study was to investigate in vitro if the probiotic bacteria used in commercial products affect the protein composition of the salivary pellicle and the adherence of other oral bacteria. METHODS: Salivary pellicle on hydroxyapatite and the adhesion of two oral streptococci, Streptococcus mutans and Streptococcus gordonii, were used as a model. RESULTS: Probiotic bacteria that bound to saliva-coated hydroxyapatite reduced the adhesion of S. mutans but the inhibitory effect on the adherence of S. gordonii was weaker. Salivary pellicle protein composition was modified by all the strains tested. The modifications in the pellicle affected the adherence of S. mutans but not of S. gordonii. Two of the proteins missing from the pellicles made of saliva-treated with the probiotic bacteria were identified as salivary agglutinin gp340 and salivary peroxidase. All bacterial strains bound salivary agglutinin gp340. The ability of the probiotic bacteria to degrade peroxidase was demonstrated with purified bovine lactoperoxidase and two of the probiotic strains. CONCLUSION: This in vitro study showed that probiotic strains used in commercial products may affect the oral ecology by specifically preventing the adherence of other bacteria and by modifying the protein composition of the salivary pellicle.

Utilization of microfiltration or lactoperoxidase system or both for manufacture of Cheddar cheese from raw milk.

The objective of the present study was to determine if application of microfiltration (MF) or raw milk lactoperoxidase system (LP) could reduce the risk of foodborne illness from Escherichia coli in raw milk cheeses, without adversely affecting the overall sensory acceptability of the cheeses. Escherichia coli K12 was added to raw milk to study its survival as a non-pathogenic surrogate organism for pathogenic E. coli. Five replications of 6 treatments of Cheddar cheese were manufactured. The 6 treatments included cheeses made from pasteurized milk (PM), raw milk (RM), raw milk inoculated with E. coli K12 (RME), raw milk inoculated with E. coli K12 + LP activation (RMELP), raw milk inoculated with E. coli K12 + MF (MFE), and raw milk inoculated with E. coli K12 + MF + LP activation (MFELP). The population of E. coli K12 was enumerated in the cheese milks, in whey/curds during cheese manufacture, and in final Cheddar cheeses during ripening. Application of LP, MF, and a combination of MF and LP led to an average percentage reduction of E. coli K12 counts in cheese milk by 72, 88, and 96%, respectively. However, E. coli K12 populations significantly increased during the manufacture of Cheddar cheese for the reasons not related to contamination. The number of E. coli K12, however, decreased by 1.5 to 2 log cycles during 120 d of ripening, irrespective of the treatments. The results suggest that MF with or without LP significantly lowers E. coli count in raw milk. Hence, if reactivation of E. coli during cheese making could be prevented, MF with or without LP would be an effective technique for reducing the counts of E. coli in raw milk cheeses. The cheeses were also analyzed for proteolysis, starter and nonstarter lactic acid bacteria (NSLAB), and sensory characteristics during ripening. The concentration of pH 4.6 soluble nitrogen at 120 d was greater in PM cheese compared with the other treatments. The level of 12% trichloroacetic acid-soluble nitrogen at 120 d was greater in RM, RME, and RMELP cheeses compared with PM, MFE, and MFELP cheeses. This could be related to the fact that cheeses made from raw milk with or without LP (RM, RME, and RMELP) had greater levels of NSLAB compared with PM, MFE, and MFELP cheeses. Cheeses at 60 d, as evaluated by 8 trained panelists, did not differ in bitterness, pastiness, or curdiness attributes. Cheeses at 120 d showed no differences in acid-taste, bitterness, or curdiness attributes. Sensory analysis at 60 d showed that PM and MFELP cheeses had greater overall sensory acceptability than RM and RME cheeses. The overall sensory acceptability of the cheeses at 120 d showed that PM, MFE, and MFELP cheeses were more acceptable than RM and RME cheeses.

The lactoperoxidase system links anion transport to host defense in cystic fibrosis.

Chronic respiratory infections in cystic fibrosis result from CFTR channel mutations but how these impair antibacterial defense is less clear. Airway host defense depends on lactoperoxidase (LPO) that requires thiocyanate (SCN-) to function and epithelia use CFTR to concentrate SCN- at the apical surface. To test whether CFTR mutations result in impaired LPO-mediated host defense, CF epithelial SCN- transport was measured. CF epithelia had significantly lower transport rates and did not accumulate SCN- in the apical compartment. The lower CF [SCN-] did not support LPO antibacterial activity. Modeling of airway LPO activity suggested that reduced transport impairs LPO-mediated defense and cannot be compensated by LPO or H2O2 upregulation.

A novel host defense system of airways is defective in cystic fibrosis.

RATIONALE: The respiratory tract is constantly exposed to airborne microorganisms. Nevertheless, normal airways remain sterile without recruiting phagocytes. This innate immune activity has been attributed to mucociliary clearance and antimicrobial polypeptides of airway surface liquid. Defective airway immunity characterizes cystic fibrosis (CF), a disease caused by mutations in the CF transmembrane conductance regulator, a chloride channel. The pathophysiology of defective immunity in CF remains to be elucidated. OBJECTIVE: We investigated the ability of non-CF and CF airway epithelia to kill bacteria through the generation of reactive oxygen species (ROS). METHODS: ROS production and ROS-mediated bactericidal activity were determined on the apical surfaces of human and rat airway epithelia and on cow tracheal explants. MEASUREMENTS AND MAIN RESULTS: Dual oxidase enzyme of airway epithelial cells generated sufficient H(2)O(2) to support production of bactericidal hypothiocyanite (OSCN(-)) in the presence of airway surface liquid components lactoperoxidase and thiocyanate (SCN(-)). This OSCN(-) formation eliminated Staphylococcus aureus and Pseudomonas aeruginosa on airway mucosal surfaces, whereas it was nontoxic to the host. In contrast to normal epithelia, CF epithelia failed to secrete SCN(-), thereby rendering the oxidative antimicrobial system inactive. CONCLUSIONS: These data indicate a novel innate defense mechanism of airways that kills bacteria via ROS and suggest a new cellular and molecular basis for defective airway immunity in CF.

Determination of minor proteins of bovine milk and colostrum by optical biosensor analysis.

Automated, rapid, sensitive, and label-free biosensor-based immunoassays for immunoglobulin G (IgG), folate binding protein, lactoferrin, and lactoperoxidase in bovine milk using surface plasmon resonance optical detection with direct binding assay format are described. Samples are prepared for analysis by direct dilution into buffer. Analysis conditions, including ligand immobilization, flow rate, contact time, and regeneration are defined and nonspecific binding considerations evaluated. The technique has been applied to the measurement of these proteins in consumer milks, colostrum, milk products, and infant formulas, and their temporal change during early bovine lactation followed.

Significance of frictional heating for effects of high pressure homogenisation on milk.

High pressure homogenisation (HPH) is a novel dairy processing tool, which has many effects on enzymes, microbes, fat globules and proteins in milk. The effects of HPH on milk are due to a combination of shear forces and frictional heating of the milk during processing; the relative importance of these different factors is unclear, and was the focus of this study. The effect of milk inlet temperature (in the range 10-50 degrees C) on residual plasmin, alkaline phosphatase, lactoperoxidase and lipase activities in raw whole bovine milk homogenised at 200 MPa was investigated. HPH caused significant heating of the milk; outlet temperature increased in a linear fashion (0.5887 degrees C/ degrees C, R2=0.9994) with increasing inlet temperature. As milk was held for 20 s at the final temperature before cooling, samples of the same milk were heated isothermally in glass capillary tubes for the same time/temperature combinations. Inactivation profiles of alkaline phosphatase in milk were similar for isothermal heating or HPH, indicating that loss of enzyme activity was due to heating alone. Loss of plasmin and lactoperoxidase activity in HPH milk, however, was greater than that in heated milk. Large differences in residual lipase activities in milks subjected to heating or HPH were observed due to the significant increase in lipase activity in homogenised milk. Denaturation of beta-lactoglobulin was more extensive following HPH than the equivalent heat treatment. Inactivation of plasmin was correlated with increasing fat/serum interfacial area but was not correlated with denaturation of beta-lactoglobulin. Thus, while some effects of HPH on milk are due to thermal effects alone, many are induced by the combination of forces and heating to which the milk is exposed during HPH.