Formulation of Senescent Plantain Dish with Various Local Cereal and Leguminous Flours for Feeding Rats : Growth Performance Evaluation

The aim of this study was to valorize senescent plantain. Therefore, a traditional dish named Dockounou was prepared with a mixture of senescent plantain and various millet, soybean, sorghum, cassava, maize or rice flours. The growth performance of several Wistar rats feed by Dockounou was followed. Thus, batches of rats were fed for 15 days with three formulations (F1, F2, F3) in proportion of 90:10, 80:20 and 75:25 (senescent plantain dough/flours) obtained after two cooking modes (dry cooking: baked ; wet cooking: boiled). The effects of these formulations were compared to control diet (C. diet). Beyond the control diet, rats fed with the soybean baked Dockounou presented, the best following growth parameters: weight gain (2.82 to 4.19 g/d), food intake (8.92 to 9.72 g/d), feed efficiency (0.10 to 0.42), proteins intake (8.28 to 19.67), proteins efficiency (0.13 to 3.15). The physicochemical and nutritive characteristics of soybean baked Dockounou were as follow: ash (2.93 ±0.15 %), proteins (10.62±0.59 %), carbohydrates (15.46±1.53 %), calcium (232.04 – 558.20 mg/100g), potassium (313.97 – 385.11 mg/100g), magnesium (42.40 – 72.22 mg/100g), sodium (211.24 – 303.85 mg/100g) and phosphorus (330.70 – 433.71 mg/100g). Also, the study showed that, two formulations, 80:20 and 75:25, have really impact on rats growth. These results suggest that soybean baked Dockounou with important proportions, 80 % and 75 %, of senescent plantain dough can be effectively used in the diet of laboratory Wistar rats regarding the good zoological performances there are obtained.


Introduction
Plantain provides more than 25 % of the carbohydrate requirements for over 70 million people in Africa (IITA, 1998).In Ivory Coast, plantain is the third crop (1 624 354 tons/year) after yam (5 731 719 tons/year) and cassava (2 436 495 tons/year) (FAOSTAT, 2013).In spite of this importance, plantain is a highly perishable foodstuff due to ethylene production associated with rudimentary conditions of harvesting, transport and storage (Rodrí guez et al., 1999;Emaga, Wathelet, & Paquot, 2008).This perishability leads to post-harvest losses estimated at 35 to 60 % of the annual production (Atanda et al., 2011).Indeed, the production of plantain depends on season marked by a period of high abundance which goes from September to April (Kuperminc, 1998;Sery, 1988).During this period, the difficulties of plantains conservation make the fruits, while ripening at the ambient temperature undergo depreciation, and also qualitative and quantitative degradations throughout the distribution chain (Chia & Huggins, 2003).In order to reduce the post-harvest losses, the ripe fruits of plantain are often processed by Ivorian farmer women into traditional dish called Dockounou (N'guessan, Yao, & Kehe, 1993;Lassois, Jean-Pierre, & Haï ssam, 2009;Dzomeku, Dankyi, & Darkey, 2011;Honfo, Tenkouano, & Coulibaly, 2011).Indeed, the Dockounou is a baked or boiled mixture of over ripe banana paste and cereals flours (Akoa et al., 2012(Akoa et al., & 2013)).Several studies have been performed on this food primarily designed for human consumption.In this way, Kra et al. (2013) and Kouadio et al. (2014) have showed that Dockounou is an energetic dish for human.Moreover, the reports of Akoa et al. (2014) have revealed the optimized parameters for this food preparation with best hygienic and nutritive qualities.As far as animal feeding is concerned, preliminary studies of Kouadio et al. (2015) highlighted the Wistar rats interest for this dish.Indeed, rats are experimental mammals, mostly used in biomedical, behavioral, toxicological and nutritional research projects (Baker, Lindsey, & Weisbroth, 1979).Therefore, using rats in experimental design could give an accurate response to some issues related to nutritional aspects of Dockounou dish.Furthermore, the use of senescent plantain in laboratory rats diet can contribute to reduce plantain post-harvest losses.Thus, several formulations using feed ingredients such as cereal or leguminous flours in mixture with various proportions of senescent plantain dough were investigated in order to evaluate their effect on the Wistar rats growth performance and contribute therefore to valorize the fruits at the senescent stage.
As for the animal material, it consisted in a total of one hundred and eighty five (185) young Wistar rats (60 to 75 g) with 50±3 days age.They were kindly provided by the animals' barn of the UFR Biosciences of Felix Houphouë t-Boigny University (Abidjan, Côte d'Ivoire).

Sorghum, Millet, Maize and Soybean Flours
Sorghum, millet, maize and soybean flours were prepared using the method described by Lombor, Umoh, & Olakumi (2009).The grains were cleaned and soaked in clean tap water and covered container.The soaked grains were allowed to ferment at room temperature (27 °C) for 24 h.After fermentation, the water was removed and the grains rinsed with 500 mL of water and oven dried at 80 °C for three (3) hours.The soybean grains were sorted, cleaned and blanched at 100 °C for 10 min.The blanched grains were, dehulled and rinsed with 500 mL of water in order to remove the seed coat.The rinsed seed were then dried in oven at 80 °C for five (5) h.Each dried sample was separately milled and sieved with a 100 μm particle size sieve.

Cassava Flour
Method described by Younoussa et al. (2013) was used.The cassava roots were peeled, cut in small pieces and washed before being sun-dried for three to seven (3-7) days.After drying, the pieces were then ground to obtain the flour which was sifted with 100 μm size sieve.

Rice Flour
Rice flour was obtained using the method described by Akoa et al. (2014).The rice grains were soaked in water for two (2) hours.After drying, the grains were crushed in a home clean wooden mortar and the flour was sieved (100 μm).

Preparation of Dockounou
Methods described by Kra et al. (2013) and Akoa et al. (2014) were used in the processing step.Fruits of senescent plantain were washed, peeled and crushed in a traditional mortar to obtain homogenous dough.Three (3) types of formulations (plantain dough: flour) were obtained.Thus, the first formulation F1 (90:10) was made with 90 % of senescent plantain dough and 10 % of flour.The second formulation F2 (80:20) was achieved with 80 % of senescent plantain dough and 20 % of flour and the third formulation F3 (75:25) was achieved with 75 % of senescent plantain dough and 25 % of flour.Each sample obtained was fermented during four (4) hours before being wrapped (in portions of 100 g) in leaves of Thaumatococcus danieilii.One part of samples has been boiled (1 h) and the second part was baked in oven at 150 °C for one (1) h.

Animals Feeding
The experiments were conducted with young Wistar rats from the animals' house of the UFR Biosciences of the Felix Houphouë t-Boigny University (Abidjan, Côte d'Ivoire).The average temperature of the room was 26 °C, and the humidity was 70 %, with 12 hours of daylight.Thirty seven (37) groups of five (5) Wistar rats each were used.They were divided as follow: one (1) group of five (5) young rats was fed with control diet (C), and for each formulation (F1, F2, F3), twelve (12) groups of five (5) young rats were fed with baked and boiled Dockounou (six groups for each cooking mode).The control diet (see Appendix A) used in this study was a granular provided by SFACI Company.Rats were acclimatized for three (3) days during which, they were fed with control diet and thereafter, fed with different experimental diets.Rats were disposed in individual screened bottomed cages designed separately to feed "ad libitum" for fifteen (15) days.

Experimental Design for Data Statement
Method described by Adrian, Rabache, & Fragne (1991) was used to determine the growth parameters.During the experimental period, the feed intake was measured daily while the body weight, length of body, tail and shin were measured each 3 days between 7h30 and 8h30 am.Each food was weighed before being given to rats and the following day, the leftover food was also weighed, in order to determine the food intake.The dry matter of each feed was determined according to AOAC (1990).The table in Appendix B shows the formulas used to determinate growth parameters.
During the experimental period, faeces of each rat were collected and the nitrogen (N) balance was evaluated by using the Kjeldahl method (AOAC, 1990).

Physicochemical and Biochemical Analysis of Dockounou
Physicochemical and biochemical characteristics of Dockounou involving best zoological performances were determined.Moisture, dry matter and ash content were determined by using the AOAC (1990) methods while the pH was directly measured with a pH-meter (Roucaire, Metr Ohm 632, Germany).Total titrable acidity was evaluated on the puree of Dockounou obtained from 40 g of paste.Raw proteins contents were measured by the Kjeldahl method according to BIPEA (1976) using 6.25 as conversion factor.Lipid content was determined after extraction with hexane in a Soxtherm system during 6 h (BIPEA, 1976).Total sugars contents were evaluated with the method of Dubois et al. (1956).As for carbohydrate and starch contents, they were calculated by the expression described by Coulibaly (2008): Starch content = 0.9 (Carbohydrate (%) -total sugars (%)).
Energy value was also calculated using the relation described by Atwater and Rosa (1899).

Mineral Analysis of Dockounou
Minerals contents were determined by the AOAC (1990) method.Five (5) grams of samples were digested with a mixture of concentrated nitric acid, sulfuric acid and perchloric acid (10:0.5:2,v/v) and analyzed using an atomic absorption spectrophotometer (GBC 904AA; Germany).Total phosphorus was determined as orthophosphate by the colorimetric ascorbic acid method (APHA, 2001).After acid digestion and neutralization using phenolphthalein as indicator reagent, the absorbance was read at 880 nm in a spectrometer (Spectronic 21 D, Miltonroy, New York, USA).Blank analysis was performed before, in the same conditions using distilled water.

Statistical Analysis
Statistical analysis of data was done by the one way Analysis of Variance (ANOVA) using software IBM SPSS Statistics version 17.0.Differences between means were tested using the Duncan Multiple Range Test with 5 % level of significant difference and figures were drawn on EXCELL 2013 Software.Dry matter intake of rats is given in Figure 2.For all formulations, feed intake is higher for baked Dockounou, than boiled Dockounou.Except for the C diet where the highest intake noted for F2 and F3 formulations was recorded by the baked D soybean.The registered values were 8.92±0.28g/d and 9.72±1.54g/d for F2 and F3 formulations, respectively.However, C diet registered the most important intake, followed by D. soybean.Food efficiency ratios of all diets are reported in Table 1.The statistical analysis showed that there is no significant difference at 5 % level between food efficiency ratios of boiled and baked Dockounou for F1 formulation.Nevertheless, a significant difference (p<0.05 ) was observed between food efficiency ratios of boiled and baked Dockounou for F2 and F3 formulations.At the whole, food efficiency ratios of baked D soybean (0.31±0.07 to 0.42±0.06)are higher than those of boiled D soybean (0.25±0.10 to 0.29 ± 0.06) for F2 and F3 formulations.However, the food efficiency ratios of D soybean for both formulations were higher than those of the others Dockounou.Results showed (Table 2) that only the D soybean diets resulted in a visible variation of length of body, shin and tail of young rats followed by the rice Dockounou diet which had just given a relatively weak variation of these parameters.There are significantly different (p<0.05) between both cooking method and type of formulations, mainly as F1 and F2 are concerned.The higher length of body was obtained for rats fed with the C diet (2.28±0.07cm), followed by those of the D soybean (0.26±0.18 cm ˂ 0.92±0.56cm ˂ 2.76±0.30cm in F1, F2 and F3 formulations, respectively).The rats body length evolution was similar with both boiled and baked D soybean in the F3 formulation (2.70±0.48cm and 2.76±0.30cm for boiled and baked D soybean, respectively).Rats fed with the baked D rice, in 75:25 proportions (F3) sleazy, obtained an increase in length of body estimate to 0.54±0.18cm.About the other diets (D. millet, D. sorghum, D. cassava and D. maize),, there was no increase in length of body, tail and shin for rats whatever the cooking mode was.

Proteins Intake
Table 3.Proteins intake of rats fed with baked and boiled Dockounou diets Table 3 presents the amount of total proteins intake in rats.At the whole, it was observed that the contents of proteins ingested by rats fed with baked Dockounou were higher than those of rats fed with boiled Dockounou.The proteins intakes for the baked preparations were 14.22 ±0.44 g and 19.67 ±3.18 g for D soybean in F3 and F2 formulations, respectively.Whatever, the amount of proteins intake of rats increased with incorporation of flours.
The amount of proteins intake by rats fed with C diet (25.25±1.45g) was higher than those of baked D soybean (8.28±0.72 g ˂ 14.22±0.44g ˂ 19.67±3.18g for F1, F2 and F3 formulations, respectively).Proteins efficiency ratios of different formulations for baked and boiled Dockounou are shown in Table 4.The whole values in the table were negative, except for those obtained with C. diet and D. soybean which were positive.The statistical analysis revealed that there was no significant difference (p>0.05) between values of proteins efficiency ratios of baked and boiled D soybean for F1 and F2 formulations.These values in the both formulations were higher than those of the C diet.However, for the F3 formulation, proteins efficiency ratios of boiled Dockounou were higher than those of baked Dockounou (5.71±1.38 > 3.15±0.44).Proteins content of Dockounou for F1, F2 and F3 formulations are presented in Figure 3. Values were significantly different (p˂0.05) between cooking methods and between formulations.Proteins contents of types of Dockounou ranged from 1.23±0.00% to 14.44±0.00% for D cassava and D soybean in F1 and F3 formulations, respectively.Control diet recorded the highest protein content, followed by those of baked and boiled D soybean in F2 and F3 formulations.All the other contents were at far slighter and statistically similar.The nutritive parameters of soybean Dockounou are presented in Figure 4. Results revealed significant difference (p ˂ 0.05) between the parameters of the whole diets (C diet and the boiled and baked D soybean).Generally, the C diet recorded higher contents of dry matter, ash, proteins, total carbohydrate and starch than boiled and baked D soybean.However, both Dockounou registered the same amount of ash, proteins, fat, total carbohydrate and starch.Data in table 5 also revealed significant differences between the three types of Dockounou, as far as the pH, total sugar content and energy are concerned.If Control diet registered the most important pH (6.20±0.10)and energy (293.11±7.46), it also topped the slightest amount of total sugars (0.21±0.03).All the diets presented statistically, similar acidity, but the pH (5.70±0.10 and 5.60±0.10) of both boiled and baked D. soybean were lower than that of the Control diet.Acidity.Baked D. soybean is richer in total sugars (0.29±0.03 g) than boiled D. soybean (0.23±0.02 g) which is less energetic (165.67±10.60Kcal) than the first (171.35±9.98Kcal).The whole diets contained many various minerals such as calcium, sodium, phosphorus, magnesium, manganese, zinc, cobalt, iron, copper, but also lead (Pb).The mineral content of soybean Dockounou and C diet, showed no significant difference (p ˂ 0.05) between copper (Cu), iron (Fe), potassium (K) and lead (Pb) contents of soybean baked Dockounou and those boiled D soybean as reported in the Table 6.At the whole, the C diet had higher mineral contents than soybean Dockounou, but Ca, Co and Mg contents of boiled D soybean are higher than those of baked D soybean.

Discussion
The length and weight are parameters that assess the growth and well-being of individual for a particular time (Morgane et al., 1993;Turlejski, 1996), but more especially appreciate of the nutritional state (WHO, 1995).Rats fed with Dockounou which were being incorporated the soybean flour had given best results zoological parameters at the end of the experiment.According to Ijarotimi & Oluwalana (2013), the weight gain is influenced by the quality of food composition and the quantity of food intakes by rats.Soybean is rich in protein and fats (Bender & Bender, 1999) and this could explained the fact that Dockounou without soybean could not allow Wistar rats growth.This fact highlights the importance to add nutrient-rich ingredients in formulations for better growth.
The consumption increased with proportion of soybean flour added and the cooking method.Thereby, an increase in weight of rats was also a link to these conditions.Udensi et al. (2012) reported the same observation in their study.In fact, soybean is a good source of protein and is generally used to enrich the foods poor in nutrients.The high food intake for baked D soybean might suggest that rats found this food more acceptable than the others as reported by Onweluzo & Nwabugwu (2009).The food efficiency ratio means the efficiency of food digestion, in other words, the weight gain obtained per unit of food consumed.Results of this parameter confirmed easly the lost or the gain of weight of rats according the diet.
The same applies to proteins intake and proteins efficiency ratio for which the statistical analysis revealed that there was no significant difference between baked and boiled Dockounou but proteins efficiency ratio of boiled Dockounou were higher than that baked.According to Ahenkora et al. (1997), the nature of the processing medium influenced nutritional qualities in the products.Indeed, the cooking mode influences proteins content.
With the dry cooking, the food becomes dry and proteins which it contains are made difficult to digest by young rats.In contrast, with the wet cooking, the presence of water in the food makes easier the digestion of proteins by young rats.These proteins in a wet food are more accessible.Swaminathan & Gangwar (1961) estimated that the losses of nutrients are due to leaching into the cooking water.Generally, nutritional qualities of plantain fruits varied with the cooking processing method employed (Baiyeri et al., 2011).Thus, proteins content of the boiled D soybean were more ingested by young rats than proteins of baked D soybean.For the other types of formulated Dockounou, weak proteins intake could be explained by the fact that these cereal foods and roots of cassava are less rich in proteins and more important source of energy (Gomez, 1985;Tonukari, 2004).The high proteins intake, and protein efficiency ratio of baked D soybean could be due to his high food intake by the animals in the one hand and the other hand, by the content of soybean.Proteins content of the baked D soybean determined in our study highlight the wealth of soybean in protein.Indeed, this content of proteins is high than the others types of Dockounou.Moreover, the studies showed that a high food intake resulted in a sharp increase in proteins intake because proteins is required for proper growth, healthy living, maintenance and production tissues and cells of the young body (Messina, 1999).Also, the addition of legumes such as soybean to foods, increases the nutritional benefits by increasing of proteins content of food (Gomez, 1985;Nnam, 2001;Perez et al., 2008).
The quantity of proteins recommended in the diet of rats adults is least 12 % (Rogers, 1979).Consequently, the proteins content of soybean Dockounou in 80:20 formulation could be used for feeding adult rats.Concerning minerals, the recommended dietary allowance (RDA) (FAO, 2004) are: calcium (1000 mg/day) and phosphorus (800 mg/day).Thus, the soybean boiled Dockounou could cover 55 % of calcium RDA and 41 % of phosphorus RDA.While, the soybean baked Dockounou could cover 23 % of calcium RDA and 54 % of phosphorus RDA.Calcium and Phosphorus are associated for growth and maintenance of bones, teeth and muscles (Turan et al., 2003).

Conclusion
The present study showed that the cooking method could affect nutrients intake of Dockounou.The baking processing provides also,a better nutrients intake than the boiling one.However, soybean baked Dockounou formulated in 80:20 and 75:25 proportions used to feed the rats and which provide them a high physical and biological performance are the best diets.The proportions of senescent plantain (80 and 75 %) in this diet reveled that feeding rats with Dockounou could help to valorize the very ripe banana in order to reduce its post-harvest losses.

Figure 1 .
Figure 1.Weight gain per day of rats fed with various types of boiled and baked Dockounou for F1, F2 and F3 formulations and control diet Data are represented as Means ± SD (n=5).Means with no common letter (a, b, c, d) are significantly different (p˂0.05)

3. 2
Figure 2. Dry matter intake by rats with F1, F2 and F3 formulations of boiled and baked dockounou and control diet.DMI= dry matter intake Data are represented as Means ± SD (n=3).Means with no common letter (a, b, c, d) are significantly different (p˂0.05)

Figure 3 .
Figure 3. Proteins content of various Dockounou for F1, F2 and F3 formulations Data are represented as Means ± SD (n=3).Means with no common letter (a, b, c, d) are significantly different (p˂0.05)

Figure 4 .
Figure 4. Nutritive properties of baked and boiled D soybean and C diet Data are represented as Means ± SD (n=3).Means with no common letter (a, b, c, d) are significantly different (p˂0.05) for each parameter.

Table 1 .
Food efficiency ratios of rats fed with baked and boiled Dockounou diets Values are mean ± standard deviation of five determinations.Values with different letters in the same line of each parameter indicate statistical difference (p ˂ 0.05).D: Dockounou, F: Formulation

Table 2 .
Length of body, shin and tail of young rats fed with control diet and various types of baked and boiled Dockounou in the three formulations F1, F2, F3

Table 4 .
Proteins efficiency ratios of rats fed with the control, baked and boiled Dockounou diets Values are mean ± standard deviation of five determinations.Values with different letters in the same line indicate statistical difference (p ˂ 0.05).D: Dockounou, F: Formulation

Table 5 .
Physicochemical and biochemical properties of baked and boiled D soybean and C diet Values are mean ± standard deviation of five determinations.Values with different letters in the same line of each parameter indicate statistical difference (p ˂ 0.05).D: Dockounou, F: Formulation

Table 6 .
Minerals composition of soybean baked and boiled Dockounou and C diet