Influence of Plantain and Sprouted Soybean Pastes on the Nutrient and Proximate Composition of Two Species of Cocoyam Puddings as a Complementary Food

Nutrients and proximate composition of the puddings prepared from ede-ocha (Xanthosoma sagittifolium) and ede-cocoindia (Colocasia esculenta) pastes were evaluated. Paste from each cocoyam variety was separately blended with firm ripe plantain, sprouted soybean pastes in the ratio of 100% cocoyam, 90% cocoyam:10% soybean, 25% cocoyam:65% plantain:10%soybean, and 45% cocoyam:45% plantain:10% soybean and mixed with equal quantities of seasonings. The formulated blends were wrapped in plantain leaves and boiled for 30 minutes. Values obtained showed that pudding from ede-ocha had a higher moisture content (56.25%), protein (4.47%), fat (1.84%), ash (2.57%), crude fiber (2.46%), iron (0.58mg), and calcium (5.12mg) than ede-cocoondia, which had 5.52%, 4.29%, 1.49%, 2.45%, 2.11%, 0.36% and 5.01% respectively. Also, puddings from ede-cocoindia had higher carbohydrate (39.28%), vitamin A precursor (62.90μg), vitamin C (33.05mg) and zinc (0.35mg) than that from ede-ocha which had 35.46%, 60.80 μg, 20.50 mg, and 0.32mg respectively. The ede-ocha, therefore, made a better nutritious pudding than ede-cocoindia.


Analysis
All the pudding samples were subjected to nutrient and proximate analyses separately in triplicates as described below. The mean of the triplicate values were used for statistical analyses. Carbohydrate was calculated as difference (100% -values of all components).

Moisture content determination
Gravimetric Oven Drying Method protocol described by Onwuka [31] was used. Ten grams (10g) of the sample was put into a previously cleaned and weighed moisture can, dried in the oven at 105°C for 3 hours, cooled in a desiccator and weighed after. The process was repeated at an hour interval until a constant weight was obtained. The final dry weight was recorded and used to calculate the percentage moisture content of the sample as shown below: where W1 = initial weight of empty can, W2 = weight of can + sample before drying, W3 = weight of can + sample after drying.

Crude protein determination
Kjeldahl Method of Onwuka [31] was used. One gram (1.0g) of the sample was mixed with 10mls of concentrated H2SO4 in a digestion flask. A tablet of selenium catalyst was added before heating in a fume cupboard until a clear solution was obtained (i.e. the digest) which was diluted to 100mls in a volumetric flask.
10mls of the digest was mixed with equal volume of 45% NaOH solution in a kjeldahl distillation apparatus. The mixture was diluted into 10mls of 4% buric acid containing 3 drops of mixed indicator (bromoscresssol green/methyl red). A total of 50mls of distillates was collected and titrated against 0.02N EDTA from green to deep red endpoint. The N2 content and hence the protein content was calculated using the formula below: % Protein = % N2 x 6.25 where w = weight of sample, N = normality of titrant (0.02 H2SO4), Vt = total digest volume (100m/s), Va = volume of digest analyzed (10ml). T = titre value of sample and B = titre value of blank.

Ash content determination
Muffle furnace ignition method described by Onwuka [31] was used. Three grams (3g) of the sample was measured into washed, dried and weighed porcelain crucible and ignited in the muffle furnace at 550°C. The sample was allowed to ash to a grayish white ash, brought out from the furnace using a forcep and left in a desiccator to cool. The cool porcelain was weighed and ash content calculated as shown below: where W1 = weight of empty crucible, W2 = weight of crucible + food before drying and W3= weight of crucible + ash.

Fat content determination
Soxhlet ether extraction protocol of Onwuka [31] was employed. Three grams (3g) of the sample was weighed into a thimble and placed into a reflux flask fitted to a weighed 300ml round bottom flask. A 300ml round bottom flask was filled with 250ml of petroleum ether (Bp 40 to60 0 C) and placed on a heating mantle preset at 60°C to reflux for about 6h during which the vapour rises and leaches all the oil from the sample in the thimble into the round flask. Thereafter, the thimble containing the sample was removed from the reflux flask and the excess was either recovered by

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heating leaving only the oil in the round bottom flask. The flask was detached from the set up and placed on the oven set at 105°C to dry off excess ether, allowed to cool in a desiccator and then reweighed and the oil was calculated as shown below: % Fat = Weight of fat Weight of sample × 100 1 %

Crude fiber determination
The method of Onwuka [31] was used. Two (2g) grams of each sample were digested with 200 ml of 1.25% H2SO4 solution under reflux for 30 min boiling. The digest was allowed to cool and then filtered with Buckner funnel equipped with muslin cloth. The residue was washed thrice with hot water, scooped into a conical flask and digested with 200 ml of 1.25% NaOH solution under reflux for 30 min boiling. The digest was cooled, filtered and washed thrice with distilled water. The residue was drained and scooped into a previously dried and weighed crucible and then put into the oven to dry at 105°C to a constant mass. The dish with its content was reweighed after drying and then placed in the muffle furnace to ash at temperature of 550°C for 3 h. The ash was withdrawn at the end and put in a bell jar and reweighed. The weight of fiber was calculated as a percentage of weight of sample analyzed as below: where W2= weight of crucible + sample after boiling, washing and drying and W3= weight of crucible + sample as ash.

Calcium determination
The EDTA complexometric titration method described by James [32] was used. 10g of the sample was dispensed into separate conical flasks, pinches of the masking agents (potassium cyanide, potassium ferrocyanide and hydroxyl hydrochloride) were measured into each flask and 20ml of ammonia buffer was added to raise the pH. The flask containing sample at10.0 pH a pinch of Erichrome dark black indicator was added and titrated against 0.02N EDTA solution while Solochrome dark blue indicator was added and titrated against 0.02N EDTA solution at pH of 12.0. A reagent blank was titrated as a control. The calcium content of the samples was calculated using the standard that 1ml of 1N EDTA has an equivalence of 20.04mg calcium.
where W= Weight of sample analyzed, Vf = Volume of extract, Va = Volume of extract used and N = Normality.

Zinc determination
Zinc was determined according to AOAC [33] method. One gram (1g) of the sample was first digested with 20ml of acid mixture (650ml concentrated HNO3, 80ml perchloric acid (PCA). About 5ml of the digest was diluted to100ml with distilled water and subjected to AAS reading. Also a standard solution of various zinc concentrations of 0.0, 0.2 and 1.0 was prepared and subjected to AAS to generate the standard curve. The concentration was calculated by extrapolation on the standard curve.

Determination of iron
The iron content was determined by spectrophotometric method of James [32]. One gram (1g) of the sample was first digested with 20ml of acids mixture (650ml concentrated HNO3, 80ml perchloric acid and 20ml concentrated H2SO4). The digest was diluted by making up to 100ml with distilled water. 2ml of the sample solution was pipette inside a flask before 3ml buffer solution, 2ml hydroquine solution and 2ml bipyridyl solution were added. The absorbance reading was taken at wavelength of 520nm and the blank was used to zero the instrument. Also, a standard solution of iron Sustainable Food Production Vol. 5 was prepared by dissolving 3.512g of Fe (NH4)2.(SO4). 6H2O in water and two drop of 0.5N HCL was added and diluted to 500ml with distilled water. The iron standard was further prepared at different concentration at 2ppm to 10ppm by diluting with distilled water. 3ml buffer solution, 2ml hydroquinone solution and 2ml bipyridtyl solution were added. Absorbance reading was taken at 520nm. The readings were used to plot a standard iron curve for extrapolation.

Vitamin A determination
Spectrophotometric method of Okwu and Josiah [34] was employed. Five gram (5g) of sample was dissolved in 30ml of absolute alcohol (ethanol) and 3ml of 5% potassium hydroxide was added. The mixture was boiled under reflux for 30 minutes, cooled rapidly with running water, filtered; 30ml of distilled water was added and transferred into a separating funnel. The lower layer was discarded and the upper layer was washed with 50mls of distilled water. The extract was evaporated to dryness and dissolved in 10mls of isoprophyl alcohol and its absorbance was measured at 325nm and vitamin A was calculated as.
where au = absorbance of test sample, as = absorbance of standard solution, c = concentration of the test sample and w = weight of sample.

Determination of vitamin C
The method used was as described by Ukwu and Josiah [34]. 10g of the sample was extracted with 50ml EDTA/TCA extracting solution for 1 hour and filtered through a Whatman filter paper into a 250 ml conical flask, 10ml of 30% KI and 50mls of distilled water was added. Starch indicator (2ml of 1%) was added and titrated against 0.01ml CuSO4 solution to a dark endpoint.
where Vf = volume of the extract, T = Sample titre -blank titer.

Statistical analyses
Mean data obtained from triplicate analyses were subjected to analysis of variance (ANOVA). A completely randomized design using SPSS version 22 was used to analyze the data. Means were separated using Duncan multiple range test at 95% confidence level (p<0.05).

Results and Discussion
Proximate results of the puddings from both cocoyam varieties were presented in Table 2. Moisture content (56.25%) of ede-ocha pudding in sample 101 (100% cocoyam) was significantly (p<0.05) higher than 52.55% from ede-cocoindia pudding in sample 105 (100% cocoyam). The difference could be due to variety. Same reason may explain higher MC in sample 104 (45% cocoyam, 45% plantain, 10%) than 108 (45% cocoyam, 45% plantain, 10% soybean) which were their least MC values. The puddings MC from 100% cocoyam of both varieties were significantly higher than their counterparts with plantain and soybean pastes in their formulations. Protein must have bound with some water [30,35] resulting lower MC. The more the moisture, the softer the pudding texture. Water also helps to maintain smooth paste consistency [5], lubricates and adds juiciness during eating and swallowing. Therefore, puddings from 100% ede-ocha will be easier to swallow than that from ede-cocoindia and those with plantain and soybean in their formulation.

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SFP Volume 5 significantly their protein contents, but the increase in ede-ocha puddings were more than that in edecocoindia. Despite these, consumption of 300 to 400g and 310 to 420g of puddings from ede-ocha and ede-cocoyam respectively will meet the recommended protein daily intake of 9.1g/d [36] which is possible considering infants' stomach capacity of 200ml [37]. Protein is an essential nutrient for proper growth and development of the body of infants and young children and a major structural component of muscle tissues which helps to repair increase and maintain children muscle mass. Besides, protein is also a component of child's blood, organs, skin and glands [38]. There is varietal influence on fat content of the puddings of both cocoyam varieties. This was reflected on significantly (p<0.05) higher fat content of ede-ocha pudding (1.84%) in sample 104 (45% cocoyam, 45% plantain, 10% soybean) than 1.49% from ede-cocoyam pudding in sample 108 (45% cocoyam, 45% plantain, 10% soybean). The variation may also be attributed to higher (39.28%) carbohydrate content of ede-cocoindia than 35.46% from ede-ocha puddings which may have proportionally reduced the total percent fat content despite higher fat content (1.21%) of edecocoindia than ede-ocha (0.95%). Besides, all the pudding samples from both varieties with same formulations were significantly difference (p<0.05) from each other. Fat contribution from the puddings of both cocoyam varieties agrees with recommendations of vegetable oil inclusion in infant and children foods [39] to increase the energy density and transport vehicle for fat soluble vitamins. Fat will aid infant in swallowing the puddings while enhancing the flavor for increase acceptability.
Ash, an index of mineral content of the puddings exhibited significant (p<0.05) variations with cocoyam variety. Maximum ash content (2.57%) was from ede-ocha pudding in sample 104 (45% cocoyam, 45% plantain, 10% soybean) while the lease value (2.31%) was from ede-cocoindia in sample 104 (45% cocoyam, 45% plantain, 10%). Also, significant higher ash content (2.14%) of sample 105 containing 100% ede-cocoindia than 2.01% in sample 101 with 100% ede-ocha follow suit and also validated the report of [10] that ede-cocoindia contains more mineral than ede-ocha. It is interesting to note that despite same formulations and higher ash content of ede-cocoindia, ash content of sample 104 (45% cocoyam, 45% plantain, 10% soybean) was higher than sample 108 (45% cocoyam, 45% plantain, 10% soybean). This may mean that vitamin -mineral interactions [29] which are nutrient loss [40], and leaching of mineral [16] may have been higher in ede-cocoindia than in ede-ocha. Ash is an indication of the amount of minerals (trace elements) with a well defined biochemical functions in the human body [41]. Iron works in synergy with protein and copper to produce red blood cells that transport oxygen from lungs to all the tissues for maintaining all body's life functions like fuelling the cell division and growth of a developing body [42]. Iron deficiency leads to anemia which is a preventable disease through food fortification or formulations for infants and young children [43]. Calcium is mainly used for bones, teeth blood clotting, nerve, muscle health, and others [42,44].
Maximum crude fiber content of ede-ocha pudding (2.46%) in sample 103 (25% cocoyam, 65% plantain, 10% soybean) which was significantly (p<0.05) higher than 2.10% from ede-cocoindia pudding in sample 107 (25% cocoyam, 65% plantain, 10% soybean) confirmed the differential varietal effects. Both samples have the same formulations. The value (2.46%) from ede-ocha pudding compared better to 2.54% reported by Olayiwola et al [45] from pudding prepared from cocoyam flour than 2.10% from ede-cocoindia pudding. The difference could be as a result of lower MC of cocoyam flour which may have increased the ash content by proportion. Consumption of 2 to 3g/d of puddings from both cocoyam varieties will meet fiber RDI of 5g.d [46] for infants aged between 6months to one year which made them a good fiber source [8]. Crude fiber is a carbohydrate subtype consisting of soluble and insoluble portions responsible for normalizing infant's bowel movement prevents constipation and helps control blood pressure [46].
Significant higher carbohydrate content of ede-cocoindia pudding (39.28%) in sample 105 (100% cocoyam) than ede-ocha (35.46%) in sample 104 (100% cocoyam) could be traced to varietal difference. Same reason holds for the significant (p<0.05) higher carbohydrate content from all edecocoindia puddings than their counterpart from ede-ocha with same formulation. This validated the superiority of ede-cocoindia over ede-ocha reported by Akpan and Umoh [10]. However, consumption of 158 to 330 g/d and 175 to 370g/d respectively of puddings from ede-cocoindia and Sustainable Food Production Vol. 5 ede-ocha will meet the infant's total carbohydrate RDI of 60 to 95g/d per serving for infants aged between 6 to 12 months [36]. These will also meet 130g/d of total carbohydrate daily intake for children beyond 12 months [47]. Lower quantity of ede-cocoindia is required to meet the RDI than ede-ocha. Carbohydrates provide energy needed to fuel children's metabolism, supports growth, keeps their brain and nervous systems working and maintains overall health [47].

Vitamin composition of the puddings
Vitamin content results of puddings from both cocoyam varieties were presented in Table  3.Varietal effects increased significantly (p<0.05) the vitamin A content of the puddings from 60.80µg (ede-ocha puddings) in sample 103 (25% cocoyam, 65% plantain, 10% soybean) to 62.90µg (ede-cocoindia) pudding in sample 107 (25% cocoyam, 65% plantain, 10% soybean).The difference was validated by higher vitamin A content (13.55µg) of 100% ede-cocoindia than 12.95µg from 100% ede-ocha puddings despite same formulations. Vitamin A is a vital micronutrient in complementary foods which deficiency is of public health concern worldwide. However, puddings from ede-cocoindia require ingestion of 794g which is lower than 822g from ede-ocha to meet vitamin A RDI of 500 µg/d [48] for infant and young children aged between 6 months to 2 years.
Varietal difference also manifested in the maximum vitamin C content of ede-cocoindia (33.05mg) in sample 107 (25% cocoyam, 65% plantain, 10% soybean) which was significantly (p<0.05) higher than 20.50mg from ede-ocha pudding in sample 103 (25% cocoyam, 65% plantain, 10% soybean). Also, vitamin C content (6.32mg) of 100% ede-cocoindia was significantly (p<0.05) higher than 4.81mg from100% ede-ocha puddings. Lower vitamin C values obtained in 100% cocoyam of both varieties could be attributed to non incorporation of plantain and soybean which may mean that their vitamin C content is low. Plantain is a rich source of vitamin C [18,19]. Though puddings from both cocoyam varieties were between 'good' and 'very good' sources of vitamin C [49], but more (195 to 832g) quantities of pudding from ede-ocha is needed to meet vitamin C RDI of 40mg for infants aged between 1 to 3yrs [50] than 121 to 633g from ede-cocoindia puddings. This implies that ede-cocoindia will meet vitamin C RDI easier than ede-ocha puddings and therefore a better source. Besides, as vitamin C is needed in trace amount for biological functions [32], it implies that cocoyam of both varieties in this study are adequate to meet infants' vitamin C RDI of 40mg. Vitamin C helps in maintaining healthy immune systems, fight infections, synthesis of collagens which gives structure and maintains healthy muscle, vascular tissue, tendons, ligaments, teeth, bones, gum, cartilage, joints, lining, skin and blood vessels [50]. Severe vitamin C deficiency leads to scurvy, a fatal disease [51] which causes malformation of infants' bone [49].

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SFP Volume 5  Table 4 presents the results of the mineral content of the puddings from both cocoyam varieties. The ede-ocha puddings which recorded maximum iron content of 0.58mg/100g in sample 103 (25% cocoyam, 65% plantain, 10% soybean) was significantly (p<0.05) higher than 0.56mg/100g from ede-cocoindia pudding in sample 107 (25% cocoyam, 65% plantain, 10% soybean). Varietal difference can explain this as 100% ede-ocha pudding in sample 101 had more iron (0.45mg/100g) than 0.40mg/g from ede-cocoindia pudding in sample 105. Besides, both puddings had same formulation. Though puddings from both cocoyam varieties responded linearly to iron increase due to plantain inclusion in their formulations [18], but those from ede-ocha still maintained their significant superiority when compared to their corresponding counterparts. General lower iron content from the puddings of both cocoyam varieties apart from variety may be attributed to leaching during boiling [16] and interaction with calcium and vitamin C in the puddings [29]. Meeting iron RDI of 11mg/d [52] for infants aged between 6 months to 2 years requires consumption of higher quantities of the puddings (>1.896kg) from both cocoyam varieties which makes the puddings poor sources of iron unless other iron rich food nutrients are used in the formulations. Therefore, the puddings should be taken along with iron rich foods. Iron works in synergy with protein and copper to produce red blood cells that transport oxygen from lungs to all the tissues where they are needed for maintaining all body's life functions like fuelling the cell division and growth of a developing body [42]. Advanced stage of iron depletion leads to anemia which is characterized with fatigue and shortage of blood [53].

Mineral composition of puddings
Maximum zinc content (0.35mg/100g) obtained from ede-cocoindia pudding in sample 107 (25% cocoyam, 65% plantain, 10% soybean) was significantly (p<0.05) higher than 0.32mg/100g from ede-ocha pudding in sample 103 (25% cocoyam, 65% plantain, 10% soybean) which may be traced to varietal differences as both puddings have same formulations. This was substantiated by significant (0<0.05) higher zinc content of 100% ede-cocoindia pudding in sample 105 (0.29mg/100g) than 0.28mg/100g from ede-ocha pudding. Zinc values obtained from puddings of both cocoyam varieties were lower than 0.44mg obtained by Olayiwola et al [45] from pudding prepared from cocoyam flour which may be due to drying and cocoyam variety used. Zinc content of all the puddings of both varieties will meet the RDI of 2mg to 3mg/day for infants aged between 6 months to 2 years [48] by consuming 625 to 1079g for ede-ocha puddings and 571 to 857g for edecocoindia puddings. Puddings from ede-cocoindia are therefore better zinc source and can meet the zinc RDI easier than that from ede-ocha. Despite this, both puddings are good zinc source for older children aged (>1y) than young children as consumption of these quantities per day is feasible. Adequate zinc intake supports protein metabolism, wound healing, growth, immune function and others. In young children zinc deficiency results in retarded growth and learning ability [54].
Varietal influence was also noticed in the puddings calcium content. Maximum calcium value (5.12 mg/100g) from 100% ede-ocha puddings in sample 101 was significantly (p<0.05) higher than 5.01mg/100g obtained from 100% ede-cocoindia in sample 105. This justified the earlier report that plantain [18] and cocoyam [55] are good sources of minerals with ede-ocha being superior [10]. It is worthy to note that despite the difference, puddings from both cocoyam varieties exhibited lower calcium values in those puddings with plantain and soybean pastes inclusions than in their 100% counterparts. This could be attributed to interactions between protein and calcium content of the formulations [4]. Calcium is of tremendous importance to infants and young children especially in healthy development of their bones and teeth [56]. Due to general low calcium content of the puddings from both cocoyam varieties, consumption of more than 2700g will be required to meet calcium RDI of 270 to 600 mg/day [56] for infants aged between 7 to 12 months. This makes the puddings poor calcium sources for young infants, but their 100% cocoyam puddings were better.