Major Whey Proteins Research Articles

Analysis of major proteins and fat fractions associated with mare's milk fat globules

Although several studies have aimed to identify mare's milk proteins, only the major whey proteins and some caseins have yet been characterized. Incomplete sequencing of the equine genome and the difficulty of recovering highly hydrophobic proteins mean that little is known to date about the proteins associated with milk fat globules, which have been shown to play an important role in newborns' defense mechanisms. The fat fraction, in particular the distribution of unsaturated fatty acids, has been more extensively studied, but complex lipids are only partially elucidated. This study reports a 2-DE approach combined with a powerful method for de novo protein sequencing, and quali-quantitative data on complex lipid composition determined by high performance TLC (HPTLC) and GC. The presence in mare's milk of long-chain highly unsaturated fatty acids, and the evidence of close similarity between equine and human milk fat globule membrane proteins, support the use of mare's milk for human nutrition.

The use of “lab-on-a-chip” microfluidic SDS electrophoresis technology for the separation and quantification of milk proteins

The use of a microfluidic “lab-on-a-chip” technique for the separation and quantification of milk proteins is described and compared with traditional sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). The microfluidic chip technique could separate all major milk proteins when standard protein solutions were used. In a milk system, α-lactalbumin, β-lactoglobulin, α s-casein, β-casein and κ-casein were readily separated; the resolution was comparable with SDS-PAGE. However, the immunoglobulins, lactoferrin and bovine serum albumin could not be resolved from the background in the microfluidic chip technique, but were easily resolved by SDS-PAGE. Standard curves for the major whey proteins were linear with both techniques and the calculated concentrations of the major proteins in a milk sample were comparable using both microfluidic chip and SDS-PAGE techniques. These results indicate that the microfluidic chip technology may be a rapid alternative for the separation and quantification of proteins in milk protein based systems.

Rational Design for Crystallization of β-Lactoglobulin and Vitamin D3 Complex: Revealing a Secondary Binding Site

β-Lactoglobulin (LG) is a major milk whey protein containing primarily a calyx for vitamin D3 binding, although the existence of another site beyond the calyx is controversial. Using fluorescence spectral analyses in the previous study, we showed the binding stoichiometry for vitamin D3 to LG to be 2:1 and a stoichiometry of 1:1 when the calyx was “disrupted” by manipulating the pH and temperature, suggesting that a secondary vitamin D binding site existed. To help localize this secondary site using X-ray crystallography in the present study, we used bioinformatic programs (Insight II, Q-SiteFinder, and GEMDOCK) to identify the potential location of this site. We then optimized the occupancy and enhanced the electron density of vitamin D3 in the complex by altering the pH and initial ratios of vitamin D3/LG in the cocrystal preparation. We conclude that GEMDOCK can aid in searching for an extra density map around potential vitamin D binding sites. Both pH (8) and initial ratio of vitamin D3/LG (3:1) are c...

Beta-lactoglobulin and its nanocomplexes with pectin as vehicles for ω-3 polyunsaturated fatty acids

The reduction of fat consumption calls for enrichment of non-fat foods and beverages with essential oil-soluble nutraceuticals, including ω-3 fatty acids. However, the low water-solubility and sensitivity to oxidation require new ways to solubilize and protect such sensitive compounds without compromising the desired sensory attributes of the enriched product. Beta-lactoglobulin (β-Lg), the major whey protein of cow milk, is a natural molecular nano-carrier for hydrophobic molecules. The present work provides apparently first evidence for the spontaneous binding ( K b = (6.75 ± 1.38) × 10 5 M −1) of docosahexaenoic acid (DHA) to β-Lg. Based on a recent study from our group [Ron, N. (2007). β-Lactoglobulin as a nano-capsular vehicle for hydrophobic nutraceuticals. M.Sc. thesis, Advisor: Dr. Yoav D. Livney, The Technion, Israel Institute of Technology, Haifa, Israel], we herein show the formation of colloidally stable nanocomplexes of DHA-loaded β-Lg and low methoxyl pectin below the isoelectric point of β-Lg (5.2), at pH = 4.5. By adding excess of pectin, negatively charged particles were formed containing ∼166 times higher DHA concentration than the surrounding serum. This enabled the formation of dilutable nanoparticle dispersions, which formed transparent solutions containing 0.05% β-Lg and DHA at a 1:2 (β-Lg:DHA) molar ratio, with a very good colloidal stability and average particle size of ∼100 nm. The entrapment by β-Lg, and moreover, the formation of nanocomplexes with the pectin provided good protection against degradation of DHA during an accelerated shelf-life stress test: only about 5–10% lost during 100 h at 40 °C, compared to about 80% lost when the unprotected DHA was monitored. This study presents a new way to nanoencapsulate long chain polyunsaturated fatty acids like DHA, useful for enrichment of clear acid drinks.

A longitudinal study of the protein components of marsupial milk from birth to weaning in the tammar wallaby ( Macropus eugenii)

The major milk whey proteins of the tammar wallaby ( Macropus eugenii) have been identified over the total period of lactation using proteomic analysis techniques comprising two-dimensional electrophoresis, comparative image analysis, matrix assisted laser desorption ionisation mass spectrometry (MALDI MS), de novo peptide sequencing and cross species protein matching. Samples were collected at the periods coinciding with major milestones of immunological development in the developing marsupial and in the four phases of milk production, specifically, Days 0, 5 (Phase 1); 27, 68 (Phase 2A); 137, 174 (Phase 2B) and 250 (Phase 3). Major changes in the protein content of marsupial milk whey correlated with the changing needs of the pouch young for stages in growth and development. We have shown that the levels of milk whey proteins vary with the developmental stage of the young animal, with a high number of proteins detected in early and late milk compared with the middle phases of lactation. Over 41 proteins were confidently identified, of which most had known roles in immunological protection. Proteins providing immunological protection across the lactation period included transferrin, β2 microglobulin, haptoglobulin and a 78 kDa glucose regulated protein. Immunoglobulin IgJ linker chain and a known antimicrobial cathelicidin, were only detected for the first 100–137 days, after which time Complement B factor was found to be present (Phase 2B). The changes which correlated with development and growth in the pouch young were reflected by the presence of proteins such as an α-fetoprotein like protein and clusterin found in early milk (Phase 1–2A) and two unknown proteins which were apparent in very early mammary gland secretions. This is the first comprehensive proteomic study of the major whey proteins of a marsupial across the entire period of lactation and provides fundamental data on proteins secreted by the mammary gland during key stages of immunological development of the young animal.

Effect of high-pressure treatment at various temperatures on activity of indigenous proteolytic enzymes and denaturation of whey proteins in ovine milk

The effect of high-pressure (HP) processing (200, 450 or 650 MPa) at various temperatures (20, 40 or 55 °C) on the total plasmin plus plasminogen-derived activity (PL), on plasminogen activators (PA), on protease activity (which seemed to be equivalent to cathepsin D), and on denaturation of major whey proteins in ovine milk was investigated. There were synergistic effects between pressure and temperature at 450 and 650 MPa for partial inactivation of PL, PA and cathepsin D. In contrast, processing at 200 MPa did not affect the activity of these enzymes. Treatment at 650 MPa was necessary for the denaturation of α-lactalbumin, and denaturation was enhanced with increasing temperature. In contrast, denaturation of β-lactoglobulin occurred at all combinations of HP and temperature studied. In conclusion, HP affected the activity of various indigenous proteolytic enzymes and the denaturation of major whey proteins of ovine milk, which may influence its behaviour during the manufacturing of dairy products.

Crystal Structure of A Chimeric b-Lactoglobulin, Gyuba

β-lactoglobulin is a major whey protein of 162 residues and is composed of nine β-strands (A-I strand) and one a -helix. Eight out of the nine strands form an upand-down b -barrel structure. Under physiological conditions, equine βlactoglobulin (ELG) is monomer, whereas bovine β-lactoglobulin (BLG) exists as a dimer in which I strand forms an intermolecular β-sheet. In this study, we report the 2.0 resolution crystal structure of Gyuba which was constructed from the amino acid sequences of secondary-structured regions of BLG and those of loop regions of ELG. This structure revealed that Gyuba formed dimer in which I strand forms an intermolecular β-sheet. This dimerization is similar to BLG.

Separation of minor protein components from whey protein isolates by heparin affinity chromatography

Heparin affinity chromatography has been used to extract minor protein components from two whey protein isolates (WPIs), one produced by ion-exchange chromatography (IEC-WPI) and the second by microfiltration/ultrafiltration (MF/UF-WPI). Only 1–13% of the major whey proteins bound to the heparin column. However, insulin-like growth factor-I (IGF-I), transforming growth factor-beta2 (TGF-β2) and lactoferrin were concentrated by factors of 24–38, 10–0 and 32–93, respectively, in the fraction eluted with 0.5 m NaCl from IEC-WPI and MF/UF-WPI. Two-dimensional gel electrophoresis of this fraction from IEC-WPI (but not from MF/UF-WPI) also revealed the presence of minor proteins, including heart fatty acid-binding protein, transthyretin, β2-microglobulin, and angiogenin. These observations suggest that heparin affinity chromatography can be used not only to separate the major proteins contained in some WPIs, but also to concentrate the minor cationic proteins and some milk growth factors.

Effect of high-pressure treatment at various temperatures on indigenous proteolytic enzymes and whey protein denaturation in bovine milk

The objective of the present study was to determine the effect of high pressure (HP) processing (200, 450 and 650 MPa) at various temperatures (20, 40 and 55 degrees C) on the total plasmin plus plasminogen-derived activity (PL), plasminogen activator(s) (PA) and cathepsin D activities and on denaturation of major whey proteins in bovine milk. Data indicated that transfer of both PL and PA from the casein micelles to milk serum occurred at all pressures utilized at room temperature (20 degrees C). In addition to the transfer of PL and PA from micelles, there were reductions in activities of PL (16-18%) and PA (38-62%) for the pressures 450 and 650 MPa, at room temperature. There were synergistic negative effects between pressure and temperature on residual PL activity at 450 and 650 MPa and on residual PA activity only at 450 MPa. Cathepsin D activity in the acid whey from HP-treated milk was in general baroresistant at room temperature. The residual activity of cathepsin D decreased significantly at 650 MPa and 40 degrees C and at the pressures 450 and 650 MPa at 55 degrees C. Synergistic negative effects on the amount of native beta-lactoglobulin were observed at 450 and 650 MPa and on the amount of native alpha-lactalbumin at 650 MPa. There were significant correlations between enzymatic activities (PL, PA and cathepsin D) and the residual native beta-lactoglobulin and alpha-lactalbumin in bovine milk. In conclusion, HP significantly affected the activity of indigenous proteolytic enzymes and whey protein denaturation in bovine milk. Reduction in activity of indigenous enzymes (PL, PA and cathepsin D) and transfer of PL and PA from the casein to milk serum induced by HP is expected to have a profound effect on cheese yield, proteolysis during cheese ripening and quality of UHT milk during storage.

Moisture-Induced Aggregation of Whey Proteins in a Protein/Buffer Model System

Moisture-induced protein aggregation in a dry or intermediate-moisture food matrix can contribute to the loss of product acceptability. The present study evaluated the molecular mechanisms and controlling factors for moisture-induced whey protein aggregation in a premixed protein/buffer model system. Insoluble aggregates rapidly formed during the first 3 days of storage at 35 degrees C with a slower rate afterward. Evaluation of the insoluble aggregates by solubility tests in solutions containing SDS/urea/guanidine HCl/dithiothreitol and gel electrophoresis showed that the formation of intermolecular disulfide bonds was the main mechanism for protein aggregation, and all major whey proteins were involved in the formation of insoluble aggregates. Effects of various factors on aggregation were also investigated, including moisture content, medium pH, and the addition of NaCl. The dependence of aggregation on moisture content was bell-shaped, and the maximal extent of aggregation was achieved at a moisture content of around 70-80% on a dry weight basis.

Whey fermentation by thermophilic lactic acid bacteria: Evolution of carbohydrates and protein content

Whey, a by-product of the cheese industry usually disposed as waste, is a source of biological and functional valuable proteins. The aim of this work was to evaluate the potentiality of three lactic acid bacteria strains to design a starter culture for developing functional whey-based drinks. Fermentations were performed at 37 and 42 °C for 24 h in reconstituted whey powder (RW). Carbohydrates, organic acids and amino acids concentrations during fermentation were evaluated by RP-HPLC. Proteolytic activity was measured by the o-phthaldialdehyde test and hydrolysis of whey proteins was analyzed by Tricine SDS–PAGE. The studied strains grew well (2–3 log cfu/ml) independently of the temperature used. Streptococcus thermophilus CRL 804 consumed 12% of the initial lactose concentration and produced the highest amount of lactic acid (45 mmol/l) at 24 h. Lactobacillus delbrueckii subsp. bulgaricus CRL 454 was the most proteolytic (91 μg Leu/ml) strain and released the branched chain amino acids Leu and Val. In contrast, Lactobacillus acidophilus CRL 636 and S. thermophilus CRL 804 consumed most of the amino acids present in whey. The studied strains were able to degrade the major whey proteins, α-lactalbumin being degraded in a greater extent (2.2–3.4-fold) than β-lactoglobulin. Two starter cultures were evaluated for their metabolic and proteolytic activities in RW. Both cultures acidified and reduced the lactose content in whey in a greater extent than the strains alone. The amino acid release was higher (86 μg/ml) for the starter SLb (strains CRL 804+CRL 454) than for SLa (strains CRL 804+CRL 636, 37 μg/ml). Regarding α-lactalbumin and β-lactoglobulin degradation, no differences were observed as compared to the values obtained with the single cultures. The starter culture SLb showed high potential to be used for developing fermented whey-based beverages.

Selective Separation of the Major Whey Proteins Using Ion Exchange Membranes

Synthetic microporous membranes with functional groups covalently attached were used to selectively separate β-lactoglobulin, BSA, and α-lactalbumin from rennet whey. The selectivity and membrane performance of strong (quaternary ammonium) and weak (diethylamine) ion-exchange membranes were studied using breakthrough curves, measurement of binding capacity, and protein composition of the elution fraction to determine the binding behavior of each membrane. When the weak and strong anion exchange membranes were saturated with whey, they were both selective primarily for β-lactoglobulin with less than 1% of the eluate consisting of α-lactalbumin or BSA. The binding capacity of a pure β-lactoglobulin solution was in excess of 1.5 mg/cm2 of membrane. This binding capacity was reduced to approximately 1.2 mg/cm2 when using a rennet whey solution (pH 6.4). This reduction in protein binding capacity can be explained by both the competitive effects of other whey proteins and the effect of ions present in whey. Using binary solution breakthrough curves and rennet whey breakthrough curves, it was shown that α-lactalbumin and BSA were displaced from the strong and weak anion exchange membranes by β-lactoglobulin. Finally, the effect of ionic strength on the binding capacity of individual proteins for each membrane was determined by comparing model protein solutions in milk permeate (pH 6.4) and a 10mM sodium phosphate buffer (pH 6.4). Binding capacities of β-lactoglobulin, α-lactalbumin, and BSA in milk permeate were reduced by as much as 50%. This reduction in capacity coupled with the low binding capacity of current ion exchange membranes are 2 serious considerations for selectively separating complex and concentrated protein solutions.

Interaction of β-Lactoglobulin with Resveratrol and its Biological Implications

Beta-lactoglobulin (beta-LG), the major whey protein in the milk of ruminants, has a high affinity for a wide range of compounds. Resveratrol (3,5,4'-trihydroxystilbene), a natural polyphenolic compound found in grapes and red wine, exhibits many physiological effects associated with health benefits. In this study, the interaction of resveratrol with beta-LG was investigated using circular dichroism, fluorescence and UV-vis absorbance. Self-association of resveratrol possibly occurs at high concentrations. Resveratrol interacts with beta-LG to form 1:1 complexes. Resveratrol is bound to the surface of the protein because beta-LG-bound polyphenol is in a weaker hydrophobic environment relative to 75% ethanol. The binding constant for the resveratrol-beta-LG interaction is between 10(4) and 10(6) M (-1), as determined by protein or polyphenol fluorescence. The beta-LG-resveratrol interaction may compete with self-association of both the polyphenol and the protein. It has no apparent influence on beta-LG secondary structure but partially disrupts tertiary structure. Complexing with beta-LG provides a slight increase in the photostability of resveratrol and a significant increase in its hydrosolubility.

Short Communication: Duplication in the 5′-Flanking Region of the β-Lactoglobulin Gene is Linked to the BLG A Allele

β-Lactoglobulin (β-LG) is the major whey protein in the milk of cows and other ruminants. It is well established that the predominant genetic variants β-LG A and B are differentially expressed. Extensive investigation of the genetic variation in the promoter region of the BLG gene revealed the existence of specific haplotypes associated with the A and B variants. However, the genetic basis for the differentially expressed BLG A and B alleles is still elusive. In this study additional genetic variation further upstream in the 5′-flanking region of the BLG gene was identified, including 6 single nucleotide substitutions, a single nucleotide deletion, and a 7-bp duplication. Comparison of DNA sequences showed that the investigated 5′-flanking region is highly conserved between ruminants, and the duplication g.–1885_–1879dupCTCTCGC and the substitution g.–1888A>G are only found in the BLG A and D alleles in cattle. The cytosine at position g.–1957 and the thymines at positions g.–2008 and g.–2049 are only found in BLG B alleles of cattle. It is suggested that the described genetic variability contributes to the differential allelic expression of the BLG gene.

Induction of cell death in RAW 264.7 cells by alpha-lactalbumin

Alpha-lactalbumin (α-LA), a major human milk whey protein, has been reported to exhibit bactericidal properties, immune suppressive effects, anti-proliferation and apoptosis in transformed cells; however, little is known about its anti-inflammation and related molecular mechanism. In this study we investigated the effects of α-LA on macrophages. We found that treatment with high concentration α-LA (≥100 μg/ml) could result in a time- and dose-dependent decrease in growth activity, morphological changes, increase in hypodiploid DNA population, and DNA fragmentation in RAW 264.7 cells. We also found that high dose α-LA could induce cellular apoptosis and necrosis, as determined by Annexin V binding assay. The α-LA could enhance the expression levels of cytochrome c, active caspase 3, active caspase 8, extracellular signal-regulated kinase (ERK1/2) and c-Jun N-terminal kinase (JNK) activation without changing the protein levels, but suppress the protein level of Bcl-2. The broad-spectrum caspase inhibitor, Boc-D-fmk, failed to block cell death, indicating that α-LA-induced cell death was modulated in a caspase-independent manner. In addition, the ERK1/2 inhibitor, PD98059, could partially rescue α-LA-induced cell death, while the JNK inhibitor, SP600125, could weakly protect cells from death. Our results suggested that activation of ERK1/2 might mediate α-LA-induced cell death in RAW 264.7 cells.

Modified Polyethersulfone (PES) Ultrafiltration Membranes for Enhanced Filtration of Whey Proteins

Application of membrane technology to whey protein separation is an interesting development that has seen growth in recent years. In particular, modification of existing membranes to impart charge properties on the membrane surface or in the pores has been shown to improve membrane selectivity, product purity, and throughput of protein solutions. This paper focuses on exploring the effects of membrane charge and solution pH on filtration of the major whey proteins α‐lactalbumin (14.1 kDa) and β‐lactoglobulin (18.4 kDa) using functionalized PES membranes. The membranes have an open pore structure containing charged sulfonated grafted polymer chains that allows for greater protein retention. The modified membranes were synthesized by polymerization of styrene in the membrane pores followed by sulfuric acid treatment of the resulting polystyrene grafts. The charged membrane gave a calculated selectivity of five times better than the raw membrane at pH 7.2 based on data from single protein transmission experiments. The enhanced selectivity of the tailor‐made membrane was due to increased retention of β‐lactoglobulin due to a reduction in molecular sieving combined with electrostatic repulsion between negatively charged β‐lactoglobulin and the negatively charged membrane.

Molecular evolution of monotreme and marsupial whey acidic protein genes

Whey acidic protein (WAP), a major whey protein present in milk of a number of mammalian species has characteristic cysteine-rich domains known as four-disulfide cores (4-DSC). Eutherian WAP, expressed in the mammary gland throughout lactation, has two 4-DSC domains, (DI-DII) whereas marsupial WAP, expressed only during mid-late lactation, contains an additional 4-DSC (DIII), and has a DIII-D1-DII configuration. We report the expression and evolution of echidna (Tachyglossus aculeatus) and platypus (Onithorhynchus anatinus) WAP cDNAs. Predicted translation of monotreme cDNAs showed echidna WAP contains two 4-DSC domains corresponding to DIII-DII, whereas platypus WAP contains an additional domain at the C-terminus with homology to DII and has the configuration DIII-DII-DII. Both monotreme WAPs represent new WAP protein configurations. We propose models for evolution of the WAP gene in the mammalian lineage either through exon loss from an ancient ancestor or by rapid evolution via the process of exon shuffling. This evolutionary outcome may reflect differences in lactation strategy between marsupials, monotremes, and eutherians, and give insight to biological function of the gene products. WAP four-disulfide core domain 2 (WFDC2) proteins were also identified in echidna, platypus and tammar wallaby (Macropus eugenii) lactating mammary cells. WFDC2 proteins are secreted proteins not previously associated with lactation. Mammary gland expression of tammar WFDC2 during the course of lactation showed WFDC2 was elevated during pregnancy, reduced in early lactation and absent in mid-late lactation.

Reaction kinetic pathway of reversible and irreversible thermal denaturation of β-lactoglobulin

A reaction kinetic approach for the description of reversible (unfolding) and irreversible (aggregation) denaturation of β-lactoglobulin (β-Lg) was developed in this study. This novel ap- proach allows the calculation of the content of native, partially unfolded and irreversibly denatured β-Lg as a function of the heat treatment time. Moreover, it is able to give a mathematical inter- pretation of the sharp bend on the Arrhenius plot of the formal denaturation rate constant which is characteristic of the denaturation behaviour of both major whey proteins, α-lactalbumin and β- lactoglobulin. The developed theoretical backgrounds were applied for the denaturation kinetics of β-Lg in a protein solution containing 50 g·L −1 of the isolated whey protein fraction and verified by means of differential scanning calorimetry. β-lactoglobulin / thermal denaturation / unfolding / aggregation / reaction kinetic

Whey acidic protein (WAP) regulates the proliferation of mammary epithelial cells by preventing serine protease from degrading laminin

Whey acidic protein (WAP) is a major whey protein in milk that has structural similarity to the family of serine protease inhibitors with WAP motif domains characterized by a four-disulfide core. We previously reported that enforced expression of the mouse WAP transgene in mammary epithelial cells inhibits their proliferation in vitro and in vivo by means of suppressing cyclin D1 expression (Nukumi et al., 2004, Dev Biol 274: 31-44). This study was conducted in order to clarify the molecular mechanism of the inhibitory function of WAP in HC11 cells, a mammary epithelial cell line. The assembly of laminin, a component in the extracellular matrix, was much more prominent around WAP-clonal HC11 cells that stably expressed the WAP transgene than around mock-clonal HC11 cells, and the proliferation of WAP-clonal HC11 cells was particularly inhibited in the presence of laminin. A laminin degradation assay demonstrated that WAP inhibited the activity of the pancreatic elastase-mediated cleavage of laminin B1 and the phosphorylation of ERK1/2. ERK1/2 phosphorylation was blocked by an inhibitor of the epidermal growth factor (EGF) receptor AG1478. Treatment with pancreatic elastase was found to enhance the proliferation of mock-clonal HC11 cells, but had no effect on that of WAP-clonal HC11 cells. The proliferation of WAP-clonal HC11 cells was recovered by the addition of exogenous EGF. We concluded that WAP plays some role in regulating the proliferation of mammary epithelial cells by preventing elastase-type serine protease from carrying out laminin degradation and thereby suppressing the MAP kinase signal pathway.

Separation of Selenium, Zinc, and Copper Compounds in Bovine Whey Using Size Exclusion Chromatography Linked to Inductively Coupled Plasma Mass Spectrometry

To study the role of trace elements for the quality and nutritional value of bovine milk, the distribution of selenium, zinc, and copper in whey was investigated using a method linking size exclusion chromatography to inductively coupled plasma mass spectrometry (SEC-ICP-MS). Three major peaks were detected for selenium, two peaks for zinc, and five peaks for copper. More than 65% of the selenium was found in protein fractions, mainly in fractions coinciding with the major whey proteins beta-lactoglobulin and alpha-lactalbumin. All zinc was associated with low molecular weight compounds (<5 kDa) and one of these compounds was probably citrate. More than 60% of the copper eluted in protein fractions and two of the five major peaks probably contained metallothionein and citrate. This method was used to compare milk and whey produced by organic and conventional feeding procedures. The selenium content in whey and desalted milk produced using organic regimens was significantly lower than that in conventional samples. Moreover, the proportion of selenium in protein fractions of organic whey was significantly smaller than that in conventional whey, but the distributions of zinc and copper did not differ. This study showed that with the SEC-ICP-MS technique the distribution profiles of several trace elements in whey could be studied in the same run and that the selenium profile differed in whey produced by organic and conventional procedures.