African Journal of Food Agriculture Nutrition and Development, Vol. 8, No. 2, June, 2008, pp. 170-191
Solubility, Emulsion And Foaming Properties Of Coconut (Cocos Nucifera) Protein Concentrates
HD Mepba*1, SC Achinewhu1 and T Ademiluyi2
*Corresponding author: Email: firstname.lastname@example.org
Full-fat and defatted coconut protein concentrates containing 27.80% and 30.20% proteins (on a dry weight basis), respectively, were prepared from freeze-dried coconut meal samples. Selected functional properties such as: nitrogen solubility, emulsion and foaming properties were determined. Nitrogen solubility was measured in the range of PH 2.0-12.0 in three dispersion media including water, 0.1M NaCl (low salt) and 1.0M NaCl (high salt). The emulsion properties of the protein concentrates were measured at varying PH values (2.0-10.0) and sample concentrations.
Foaming properties were determined using same parameters including the use of additives (NaCl solutions and carbohydrates). Below and above the isoelectric PH (4.0-5.0) the nitrogen solubility increased. The coconut protein samples showed a fairly high solubility (more than 54.0% and 53.0%) for full-fat coconut protein concentrate (FFC-PC) and defatted coconut protein concentrate (DFC-PC), respectively at pH 2.0. FFC-PC sample had maximum solubility (more than 73%) at 10.5 and minimum solubility (13.2%) at PH 4.0. On the alkaline PH scale, FFC-PC sample had maximum solubility of more than 43% at PH 10.5 in high salt (1.0M NaCl) solution while DFC-PC sample had maximum solubility of 50.2% in 1.0M NaCl solution and more than 80% in low salt (0.1M NaCl) dispersion medium.
The emulsions prepared had good stability. A maximum of 92.5ml/g protein of emulsification capacity, at PH 10.0 and minimum of 45.3ml/g protein at PH 4.0 were obtained for FFC-PC while DFC-PC had a maximum EC of 86.4ml/g protein. FFC-PC samples produced significantly (P≤0.05) higher emulsion capacities than DFC-PC samples at all the dispersion PH and sample concentrations investigated. The FFC-PC samples had a significantly low foaming capacity than the DFC-PC samples at all the tested PH values. Similarly, the foaming capacities of FFC-PC and DFC-PC decreased with increasing PH of the sample medium. Both FFC-PC and DFC-PC foams collapsed completely, after 3hr standing at ambient temperature. Foaming was concentration dependent. The FFC-PC and DFC-PC foams increased sharply at 0.2% NaCl content of the sample slurries and progressed steadily to a peak of 92.0% and 113% in FFC-PC and DFC-PC samples respectively in 0.8% NaCl solution and then declined gradually with increased salt concentration. Addition of polymeric carbohydrates (sucrose, corn starch, gum acacia and pectins) significantly improved foaming properties of the protein concentrates but the foam stabilities were not significantly affected.
Key words: Coconut, Protein, Solubility, Foaming, Emulsion
The advent of modern technology in processing of oil-bearing materials for extraction of oil has brought in a variety of oil-free residues, which are rich in protein. Coconuts, though abundant in the Niger Delta region and hinterland of southern Nigeria, are rarely processed but eaten raw as fresh fruits. Coconuts contain in good measure, excellent proteins, carbohydrates and minerals beside a good supply of edible oil .
After oil extraction the residual cake is generally unfit for human consumption and is used for animal feed or fertilizer . Special attention is now being devoted to concentration and isolation of proteins from oil seeds. Such protein concentrates/isolates offer greater opportunity for incorporation of and /or supplementation of a broad variety of food and may probably fit the needs of infants whose needs are critical.
To be useful and successful in food applications, proteins should have satisfactory intrinsic properties such as: nutritional properties and organoleptic attributes. Besides, the proteins should have several desirable characteristics referred to as functional properties, such as solubility under varying conditions, emulsifying and foaming capacity. Emulsifying capacity (EC) is an important protein functionality trait related to their ability to lower the tension at the interface of water and oil. Emulsifying properties of protein are affected by source of protein, protein concentrations, medium PH incubation temperature and mechanical force .
Use of plant proteins in emulsified foods systems could promote fat binding to reduce cooking losses, improve EC and maintain stability of the emulsion system . Similarly, various proteins could be used as foaming agents because their surface properties are mainly responsible for foam development . Protein foams are important in many processes in the beverage and food industries and this has stimulated interest in their formation and stability. Foams are used to improve texture, consistency and appearance of food and are commonly encountered in baked, confectionery and other goods.
Protein functionality is governed by their structural characteristics, which are themselves influenced by many factors of their environment. Although the behaviour of a protein is influenced by its solubility characteristics, the correlations between the two have been crude . The present investigation were undertaken to evaluate the functional properties of coconut protein concentrate with the objective to compare full fat and defatted coconut protein concentrates for nitrogen solubility, emulsification and foaming properties in various dispersion media.
MATERIALS AND METHODS
Sample source and preparation
Matured ripe coconuts (the West African Tall variety) were collected from the Integrated Agriculture Project Farms, Kaani, Khalga, and Rivers State. Preparation of protein concentrate was as described in a previous study .
Test of functional properties
Nitrogen solubility of both full-fat coconut protein concentrate (FFC -PC) and defatted coconut protein concentrate (DFC-PC) samples in various dispersion media were determined by a modified method of Betschart .
Two hundred mg samples each were dispersed with distilled water, 0.1M NaCl (low salt) and 1.0M NaCl (high salt) solution. The media were adjusted to different PH levels (2.0 -12.0) with 1M NaOH or 1M HCl. The dispersed samples were shaken for 30min and centrifuged at 1000 x G for 20 min.
Duplicate aliquots of supernatant were analyzed for nitrogen contents by micro-Kjeldahl. The percentage nitrogen solubility was calculated based on total nitrogen in the supernatant to total nitrogen in the sample. AOAC  methods were used to determine the moisture (7.003), protein (7.080) fat (14.019), crude fiber (7.061), and ash (dry ashing method) content of FFC-PC and DFC-PC. Carbohydrate was determined by difference of the constituents from 100.
Emulsions were prepared according to the method of Beuchat . The sample (2g) was blended in a Moulinex electric blender with 5.0ml of oil in 100ml distilled water for 30seconds at high speed. Oil was added in 5 ml portions continuing blending. The drop in consistency (from a maximum), judged by a decrease in resistance to blending (subjectively) was considered to be the point of discontinuation of oil addition. The amount of oil added up to this point was interpreted as the emulsifying capacity of the sample. The emulsion so
prepared was then allowed to stand in a graduated cylinder and volume of water separated and time noted in each case to study the emulsion stability. All experiments were conducted in duplicate and at room temperature (30 ± 1C). Effects of PH on emulsion capacity were evaluated using 2% (w/v) slurries and adjusting the PH to a desired value (PH 2.0- 10.0) with IN HCL or IN NaOH.
Effect of sample concentration
Effects of concentration on emulsion capacity were evaluated at concentrations of 2, 4, 6, 8 and 10 (w/v).
Foaming capacity and stability were studied according to the method of Coffmann and Garcia . A 2g sample was whipped with 100ml distilled water for 5mins in a Moulinex electrical blender at high speed and was poured into a 250ml-graduated cylinder. The total volume at time intervals: 0.0, 0.5, 1.0, 1.5, 2.0, 3.0 and 4.0 hrs were noted. Volume increase (%) was calculated according to the following equation.
Effects of PH on foaming properties Effects of PH on foaming capacity were studied on 2% (w/v) slurries by adjusting the PH to the desired value (PH 2.0 -10.0) before whipping.
Effects of concentration on foaming capacity
Effects of concentration on foaming capacity were evaluated by whipping 2,4,6,8, and 10% (w/v) slurries as described above. Salt (NaCl) concentrations of .0.2, 0.4, 0.6, 0.8, 1.0, 1.5, 2.0% (w/v) slurries were employed to study the effects of salt on foaming capacity.
Effect of certain carbohydrates on foaming properties
The effects of certain carbohydrates on foaming properties of coconut protein concentrates were evaluated. Galactose, sucrose (all from BDH chemicals, England), Gum acacia, pectins and corn starch (from Hopkins and Williams Laboratory Reagents, Essex, England) were used. Samples (0.25g of each carbohydrate per gram of protein concentrate) were blended in 100 ml of distilled water for 5mins. Samples were then transferred into 250ml graduated cylinders and examined for volume increase as described above. All experiments were conducted at room temperature, in duplicates and means reported.
All data were examined statistically by analysis of variance  and adopting the least significance difference (LSD) as the test for significance
Mean values for the proximate composition of full fat (FFC-PC) and defatted (DFC-PC) coconut protein concentrates are presented in Table 1.The FFC-PC and DFC-PC contained 27.80% and 30.20% proteins, respectively. The lipid, carbohydrate, crude fiber and ash contents differed significantly (P≤ 0.05).
Nitrogen solubility profiles of FFC- PC and DFC-PC as a function of PH and ionic/salt concentrations are shown in Fig 1 and Fig 2. Generally, the nitrogen solubility response of the samples to changing levels of PH in water and low salt dispersion differed from that of the high salt suspension. The Full-fat coconut protein concentrate and DFC-PC samples showed minimum solubility between PH 4.0 -5.0 and maximum solubility in the range of PH 9.0 - 10.5.
The FFC-PC sample had maximum solubility (more than 73.0%) at PH 10.5 and minimum solubility (13.2%) at PH 4.0-5.0 (Fig 1). The coconut protein showed fairly high solubility at PH 2.0 (about 54.6% and 53.2%) in FFC-PC and DFC-PC, respectively, Fig 1. However, at PH 4.0 -5.0, protein extraction increased with increasing concentration of salt. DFC-PC samples were less soluble than their unextracted counterpart in this PH range. Whereas the reverse was the case beyond the isoelectric PH, on the alkaline side there was a steep increase in solubility.
At high (1.0M) NaCl concentration, the solubility decreased drastically, while at the lower (0.1M) NaCl concentration, solubility increased at all the PH levels. FFC-PC samples had a maximum solubility of more than 43% at PH 10.5 in 1.0M NaCl solution.
Similarly, the DFC-PC samples had maximum solubility of about 50.2% in 1.0M NaCl solution and more than 80% in 0.1M NaCl dispersion medium. At the lower salt (0.1M NaCl) concentration, the solubility of both FFC-PC and DFC -PC samples increased slightly in the PH range 8.0- 10.5 indicating the “salting-in” effect while at higher salt concentration (1.0M NaCl), the solubility decreased which revealed the “salting-out” effect on the complex protein system.
Emulsion properties: Effect of PH The emulsion capacities of FFC-PC and DFC-PC over a range of PH (2.0 -10) are shown in Table 2. The Emulsion capacity (EC) of coconut protein concentrate suspensions was markedly improved by adjusting the PH from 2.0 to 10.0. Alkaline PH improved the EC more than the acid PH A maximum value of 92.5ml/g protein of emulsification capacity at PH 10.0 and a minimum value of 45.3ml/g protein at PH 4.0 were observed for FFC-PC, while DFC-PC had a maximum EC of 86.4ml/g protein at PH 10.0. Both samples showed positive correlation with PH and nitrogen solubility. Emulsion capacity of the protein suspensions was greatest at the two extreme PH levels (2.0 and 10.0) and least at PH 4.0. Removal of lipids with n-hexane significantly (P≤0.05) reduced the EC of DFC-PC when compared with FFC - PC.
Table 3 shows the emulsion stabilities of FFC-PC and DFC-FFC samples held for various times at ambient temperature. The emulsions prepared from FFC-PC and DFC-PC had good emulsion stabilities. First phase separation occurred after 36hrs and 48hrs for DFC-PC and FFC-PC, respectively when held at ambient temperature.
Effect of sample concentration
The effects of concentration on emulsion capacity of the protein concentrate are summarized in Table 4. In comparing the emulsion capacities of FFC-PC and DFC-PC samples, quantities of oil emulsified increased as sample concentration increased, though not proportionately. However, when expressed on a unit protein basis, emulsion capacity actually decreased with increased sample concentration.
Data in Table 2 show the effect of PH on the foaming capacities of FFC-PC and DFC-PC samples. FFC-PC samples had a significantly (P<0.05) lower foaming capacity than the DFC-PC samples at all the tested PH values. Similarly, the foaming capacities of FFC-PC and DFC-PC decreased with increasing PH of the dispersion medium. A maximum volume increase of 26.4% for (FFC-PC) and 41.2% (DFC-PC) at PH 2.0 and a minimum volume increase of 16.4% and 28.4% at PH 10.0 were obtained for FFC-PC and DFC-PC samples, respectively.
Data in Table 3 show the foam stabilities of FFC-PC and DFC-PC samples. Both FFC-PC and DFC-PC foams had low stability. In both samples, foam volume reduced drastically after 30mins then stabilized for about 1hr before a gradual decline for 1.5hrs and collapsed after 4hrs. Full-fat coconut protein concentrate maintained a significantly higher foam volume than DFC -FC for up to 2.5hrs.
Effect of sample concentration
Data in Table 4 show the effects of concentration on foaming capacities of FFC-PC and DFC-PC. Foaming was concentration dependent and increased with increasing concentrations of protein concentrate. There was a rapid increase in foam volume up to 6(w/v)% solids concentration with a maximum at 10(w/v)% solids concentration. The Defatted coconut protein concentrate had a significantly higher foaming capacity than FFC-PC samples.
Effect of salt concentration on formability
The effect of salt additions on the foaming capacities of FFC-PC and DFC-PC are presented in Table 5. Addition of salt (NaCl) significantly (p≤0.05) improved the foamability of FFC-PC and DFC-PC. In both samples foam volume increased sharply at 0.2% NaCl content of the slurries then improved steadily to a peak of 92.0 % and 113% in FFC-PC and DFC-PC samples respectively at 0.8(w/v)% salt solution then declined gradually with increased salt concentration.
Table 6a and 6b present the effect of carbohydrates on foaming properties of FFC-PC and DFC-PC respectively. Addition of carbohydrates other than galactose at levels of 0.25g/g protein concentrate significantly (P≤0.05) increased the foaming capacity over that of the control. Higher foam volumes were obtained in samples treated with pectins, gum acacia and corn starch.
Similar observations were made with DFC-PC (Table 6b). However foam volume was higher in DFC-PC than in FFC-PC for all treatments with carbohydrates.
Defatted coconut protein concentrate had a significantly higher protein but lower lipid content than full fat coconut protein concentrate. These differences in the proximate composition of these proteins could reflect differences in their conformation, molecular flexibility and physiochemical properties [5, 13]
Test of functional properties: Nitrogen Solubility
As depicted in Fig 1 and Fig 2, nitrogen solubility was PH dependent. The nature of the nitrogen solubility curve of coconut protein concentrates resembles the nitrogen solubility curve of many oilseeds (14-16). Similar to other oilseeds, the minimum solubility for coconut proteins was at PH 4.0-5.0. Gbenle and Onyekachi  observed that the minimum solubilities for groundnut, soya bean and melon seed were at PH 4.0-6.0. Similarly, they reported that groundnut and et al. proteins showed fairly high solubilities of about 56% and 44% respectively at PH 2.0.
The high solubilities of coconut proteins at acid PH suggest that they may be used in acid foods and beverage . Kwon et al.  also reported increased protein extraction at the isoelectric PH with increasing concentration of salts. At this PH (isoelectric) protein extraction is least extractable in water . Similar to other oilseed proteins, coconut protein demonstrated higher solubilities at alkaline PH. Available results [14-16] indicate that the nitrogen solubilities of oilseed protein isolates at their maximal PH of solubility were 88% for groundnut, 82% for the C. citrullus and 77% soyabean. The change of solubility profile of DFC-PC resulting from lipid removal may indicate the modification (alteration of ionic charges) on the protein structure as suggested by Wang and Kinsella .
Data in Table 2 show that the emulsion capacities of FFC-PC and DFC-PC were dependent on PH Similar PH dependence of emulsifying capacities of proteins has been reported . The emulsifying capacities of soluble proteins depend upon the hydrophilic-lipid balance, which is affected by PH. The net charge at the interface may impede or facilitate emulsifying activity of proteins.
Our observations on increased E C. in the alkaline PH are in agreement with the findings of Bera and Mukherjee . The DFC-PC samples contained more carbohydrate (40.68%) than FFC-PC samples (36.21%). Poor emulsion properties of DFC-PC samples were due to presence of carbohydrates. The lipid content of FFC-PC might have influenced the hydrophilic-lipid balance of proteins to more favourable level and hence had higher EC. Similarly, the emulsions prepared from FFC-PC were more stable than those from DFC-PC samples (Table 3). This was expected because the lipid content of FFC-PC influenced EC and ES .
The emulsion capacity of FFC-PC and DFC-PC samples decreased with increasing sample concentration (Table 4). Lin et al. similarly reported decreased EC of sunflower and soybean flours and protein concentrates/isolates as sample concentration was increased. Phillips  explained the dependence of EC on protein concentration on the basis of absorption kinetics at a low protein concentration, the rate of adsorption is diffusion controlled but at high protein concentration, there is an activation barrier to absorption. Under the latter conditions the ability of the protein molecules to create space in the existing film and to penetrate and rearrange on the surface is rate determining.
The FFC-PC samples had significantly poor foaming capacities than the DFC-PC samples at all the tested PH values (Table 2). The poor foaming capacity of FFC-PC may be due to presence of lipids. Wang and Kinsella  observed that bound lipids in alfalfa leaf protein preparation depressed foaming. The FFC-PC sample contained more than 12.0% fat that may be responsible for poor foaming in comparison to the DFC-PC samples.
The stability of FFC-PC and DFC-FC foams was low (Table 3). DFC-PC foams collapsed completely after 48hrs at 30C. Foamability is related to the rate of decrease of surface tension of the air /water interface caused by adsorption of protein molecules. According to Grahams and Phillips  flexible protein molecules like (β-casein which can rapidly reduce surface tension give good foamability, whereas a highly ordered globular molecule such as lysozyme, which is relatively difficult to denature give lower foaming properties.
Foaming properties of all samples increased with an increase in protein concentration (Table 4). The effect may be explained to be similar to that of PH as more proteins become available to foam and to stabilize the foam., Canella.  reported maximum foam expansion of acidbutanol treated sunflower seed flour and sunflower seed protein isolate and soya protein isolate at 1% level. On the other hand, Huffman et al.  observed an increase in foam expansion up to 8%, concentration of sunflower seed flour at PH 9.0. From the above observation it appears that foaming properties may increase with increase in protein concentration up to a certain level, presumably, due to saturation point of protein solubility under the experimental conditions.
The effect of additives on the foaming properties was variable. NaCl addition (Table 5) increased foaming properties perhaps by increasing the protein solubility through partial denaturation . Kinsella  reported formation of high capacity low stability foams when salt was added to protein suspensions. Similar variable results were obtained with the addition of carbohydrates (Table 6a and 6b). Our results with the incorporation of carbohydrates on foam volume and stabilities are in accord with Sathe et al.  ; Gujska and Khan .
Full fat coconut protein concentrate and Defatted coconut protein concentrate containing 27.80% and 30.20% protein respectively (on a dry weight basis) were prepared from the West African tall coconuts. Both FFC-PC and DFC-PC samples showed minimum nitrogen solubility between PH 4.0-5.0 and maximum solubility in the range of PH 9 -10.5.
In low salt solutions (0.1 M NaCl), the solubility of both FFC-PC and DFC-PC samples increased slightly in the range of PH 8.0-12.0, while at higher salt concentration solubility decreased indicating the “salting out” effect on the complex protein system. Emulsion properties of the protein concentrate were PH dependent. The protein concentrate emulsion had good stability (36-48 hrs). Foaming properties were PH dependent with a maximum within the acid range. Coconut protein had poor foaming capacity and could be improved by increasing sample concentration as well as adding NaCl. Although carbohydrates other than galactose improved foam volume increase, foam stabilities were not significantly (p> 0.05) affected.
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