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Cholesterol content and functional properties of products fractionated from egg yolk Omotade I. OLOYEDE1 and AdesolaO. IKUELOGBON Department of Biochemistry, University of Ado-Ekiti, Ado-Ekiti, Nigeria.1E-mail: pjmoloyede@yahoo.com, Tel: 08033972332 Received 19 November 2003 Code Number: bk04007 AbstractIn low-salt aqueous solution, egg yolk was fractionated into delipidated fraction, plasma and granule fractions were obtained using centrifugation. The following determinations were made on each fraction: Emulsifying activity (EA) and stability (ES), foam capacity and stability, protein solubility index, total cholesterol, lipid and protein. Fractionation of egg yolk into plasma resulted in five fold reduction in cholesterol. In decreasing order, EA of yolk products in distilled water was: whole egg yolk > delipidated egg yolk > plasma > granules. Emulsion and foam stability exhibited by plasma suggest that they could be used as a stabilizer of dispersion system Key words: Egg yolk, plasma, cholesterol content, functional properties. INTRODUCTIONEgg yolk is a complex mixture composed of granule and a water soluble fraction, plasma. Each fraction contains a lipoprotein as the main constituent. Granules contain mainly 70% high density lipoprotein (HDL), 16% phosvitin and 12% low-density lipoprotein (LDL). [1,2]. Plasma is composed of 85% LDL and 15% livetin [3]. Egg yolk supplies protein, vitamins, minerals, and essential fatty acids to human dietary requirements. It has a lipid-to-protein ratio of about 2:1[4]. The lipid components of egg yolk are triacylglycerol, phospholipid and cholesterol [5]. Increasing awareness of relationships between excess dietary fat and health problems i.e. coronary heart disease has resulted in lower consumption of high fat (i.e. cholesterol) products such as egg yolk. In response to consumer demand, development of low-fat cholesterol egg yolk has been introduced [12]. This was achieved by alteration in the hen’s diet to induce compositional changes in the egg yolk [6], organic solvent extraction [4,7],marketing of low cholesterol egg blends containing egg white and vegetable oil, [8,9] and supercritical extraction with carbon dioxide [7]. However, removal of fat from egg yolk is feasible only if functional properties are not reduced substantially. Egg yolk protein is an effective emulsifying agent for food products such as bakery products and mayonnaise [16]. The surface-active proteins form an interfacial film, by orienting with the polar portion of the molecule to the aqueous phase and non polar to the oil phase, thus helping to form a stabilized emulsion. The objective of this study is to fractionate egg yolk into plasma and granules by centrifugation and to determine the cholesterol content and its functional properties. MATERIALS AND METHODS Materials All reagents were of analytical grade. Fresh eggs of hen (Local breed) and Corn oil were purchased in a market at Ibadan, Nigeria. Plasma and granules fraction from whole egg yolkEgg yolk was fractioned into plasma and granules using a modified version of the procedure (15) as follows: 50 ml of pooled egg yolk after separation from the albumen was diluted with 400ml of distilled water. The mixture was thoroughly mixed for 10mins to get an homogenous suspension by a magnetic stirrer, and centrifuged at 1,600 x g for 10 min at 10oC. The plasma fraction (upper layer) was carefully decanted and the granule fraction was removed by spatula from the centrifuge tubes. Both were stored at 4oC prior to analysis. Delipidated yolk fraction Delipidated egg yolk fraction was prepared using an adaptation of the procedure described by Aulisio and Shelokov [18]. Equal volume of egg yolk without albumen and 0.16M sodium chloride solution was placed in a screw-cap tube. The mixture was mixed thoroughly by shaking and double volume of chloroform was added. The tube was inverted several times and left at room temperature for 30 mins. The inversion was repeated 4 times at 30 mins interval. The mixture was left overnight at 4oC. The mixture was centrifuged the following morning at 2,000rpm for 10 min. The upper clear saline layer was pipetted off. The middle layer of depigmented yolk debris and the lowest pigmented chloroform layer were discarded leaving the saline layer (delipidated fraction). This fraction was kept at 4oC prior to analysis. Emulsifying activity and Emulsion stabilityEmulsifying activity (EA) was determined using the turbidimetric method of Pearce and Kinsella[13]. A 1% protein dispersion of samples was made in distilled water, 0.1, 0.2 and 0.3M NaCl solutions. Samples were homogenized for 15 min using a rotor mixer at room temperature (27oC). The pH was measured and recorded. The emulsion was made by pipetting 16.0ml of protein dispersion into 250ml glass beaker, followed by addition of 24.0ml corn oil. The mixture was homogenized using a rotor mixer for 1 min at room temperature. A 30ml aliquot was removed and diluted with 20ml of 0.1% sodium dodecyl sulphate (SDS) solution. Absorbance was measured at 500nm using a 0.1%SDS solution blank, samples were analysed in duplicates. Emulsion stability (ES) was determined by the modified method of Chung and Ferrier [4]. Emulsions prepared as above were allowed to stand for 24 hr at room temperature (27oC); the total height, along with heights of the emulsion layer, and the bottom aqueous layer, were recorded to the nearest 1.0mm. The ES was calculated as the percentage (v:v) emulsion layer remaining (Table 1). Total cholesterol and protein in yolk fraction. The total cholesterol was determined using a modified method of Strove and Makarera [10]. Sample preparation was performed by dispersing 0.2g of granules, plasma, delipidated yolk and whole egg yolk in 2ml of 1M NaCl solution. Samples were analyzed for cholesterol content as follows: 0.1ml of the samples were separately pipetted into 3.0 ml of glacial acetic acid. 2.0ml of colour reagent was added along the side of the test tubes. The steps above were repeated using 0.1ml of standard cholesterol (1mg/ml) and distilled water respectively. Absorbances were read at 530nm using a colorimeter. Two replicates and 2 determinations were performed for each sample. Foam Capacity and Foam StabilityFoam capacity and foam stability were determined using a modified version of the procedure described by Bera and Mukherjee [14].Briefly 1% protein dispersion of the samples was prepared and 15ml aliquot of the protein dispersion was transferred to 250ml glass beaker and homogenized for 1min using a rotor mixer. The mixture was immediately poured into a 100ml glass graduated cylinder and the initial foam volume (foam capacity) was measured after 30 secs. Foam stability was determined by measuring the foam volume every 15 mins for 60min. The percent foam remaining after 60 mins was calculated using the formula [16]. % foam remaining (foam stability) = where Vt = foam volume at 60min V0 = initial foam volume (foam capacity). Protein Solubility index (PSI) The protein solubility index was determined using a modification of the procedure described by Causeret et al [15]. A 1% protein dispersion of samples was prepared as described. 10ml aliquot of homogenate was placed into 2 centrifuge tubes and centrifuged for 30 mins at 6,000 rpm (10oC). Following centrifugation, 5ml of supernatant was carefully removed by pipette for kjeldahl protein determinations. Statistical AnalysisAll data were expressed and the means ± SD. Differences between groups were considered to be significant at P<0.05, using student t-test. RESULT AND DISCUSSION Emulsifying activity and emulsion stability Table 1 shows the effect of different salt concentrations on the emulsifying activity and stability of yolk products. In distilled water, differences occurred (P<0.05) between EA values for each of the products. Whole egg yolk showed the highest EA (0.47) compared to Delipidated (0.46), plasma (0.39) and granules (0.37). Increasing salt concentration has little effect on the EA of plasma, but higher EA value is observed at 0.3M NaCl concentration. This observation confirms the report of [16]. The emulsifying activity values of granules, delipidated and whole egg yolk steadily increased with increasing salt concentration. Emulsion stability data (Table 2) indicated that ES value of whole egg yolk showed a noticeable increase reaching 91.2% TABLE 1: Emulsifying activity and Stability of products fractionated from hen’s egg yolk
ES is emulsion stability while EA is emulsion activity of yolk products
Table 2: Total protein and Protein Solubility Index (PSI) of protein fractionated from hen’s egg yolk.
Table 3: Foam capacity and foam stability of products fractionated from hen’s egg yolk
Table 4: Cholesterol content of products fractionated from egg yolk
followed by granules 87.5% at 0.3NaCl. However, other investigators [14,17, 19,20] have indicated that granule may have efficient-stabilizing effect. Protein Solubility Index (PSI) Protein solubility index (PSI) of fractionated products (Table 2) showed distinct similarities. PSI values steadily increased with increasing salt concentration, delipidated fraction showed the highest PSI values (83.4%). The PSI values of plasma that is higher than granules supports the observation of [16]. A portion of the LDL must have been deposited into the granule during fractionation. Plasma would be expected to have a higher ratio of livetins (water- soluble globular proteins) to LDL. Levitins may account for the higher PSI value exhibited by the plasma. The highest value of PSI for all the yolk products was revealed at 0.3M NaCl. Protein concentration of the yolk products increased with increasing salt concentration. In this study, as salt concentration increased, proteins were dispersed in the yolk products and contribute to the increase in PSI values. This observation supports the report of [15]. Foam capacity and Foam StabilityTable 3 shows the effect of different salt concentration on the foam capacity and stability of yolk products. In distilled water, whole egg yolk exhibited the highest foam capacity (18ml). Foam stability values of whole egg yolk and granule decreased steadily with increasing salt concentrations. The greater foam capacity values of plasma compared to granule suggest that granule may have lost some of its LDL to plasma as a result of centrifugation. It appears as suggested by [16] that LDL may be important in a products’ foam capacity. CONCLUSIONAnalysis of the yolk products indicated the use of plasma and delipidated egg yolk as a replacement for whole egg yolk, could result in reduction in cholesterol and increase in protein. Furthermore, emulsion and foam stability values exhibited by plasma suggest they could be used as a stabilizer of dispersion system. Granules could possibly be used in smaller quantities than whole egg yolk, while achieving comparable emulsifying activities with a reduction in cholesterol level. REFERENCES
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