Three different tubers namely white yam (Dioscorea rotundata), sweet potato (Ipomoea batatas) and cocoyam (Colocasia esculenta) were processed (roasted and boiled), the raw served as the control and all samples were characterised with respect to their proximate composition, anti-nutrients and phytochemical contents, sugars and functional groups.
Thereafter, starches were extracted from them and analysed for sugars (after digestion with α amylase enzyme) also, the structural morphology of the extracted starch granules were examined using scanning electron microscope .Results on proximate composition were significantly different p≥0.05, protein content was decreased in all the boiled and roasted samples while the ash content increased in all roasted tubers. Data on antinutrients and phytochemical revealed that tannin, saponin, terpenoid, oxalate and phytate were in the ranges (mg/g) 0.52-1.70; 17.36-169.27; 1.31-5.00, 0.18-1.16 and 6.59-22.25 respectively. Flavonoid was absent in the cocoyam while phlobatannin and alkaloid were below detection limit in sweet potato. Results on functional groups before and after starch extraction showed the presence of OH, C2H2n, CnH2n-2, RX and ROR. Scanning electron microscopy on the starch granules showed an increase in the surface area in the boiled samples. Complex carbohydrate digestion before and after addition of the enzyme revealed digestion were highest in all roasted samples. Processing generally had effects on the nutritive characteristics of all the tubers analysed.

Introduction
Tuber crops are important in human diet; they are staple foods and the major source of carbohydrates. They are also good source of dietary fibre and provide essential vitamins and minerals [1].Cocoyam belongs to the Aracea family and Colocasia genus.
All the parts of cocoyam are edible especially, the starchy tuberous root [2]. Sweet potato (Ipomoea batatas) belongs to the Convolvulaceae family [2]. Raw sweet potatoes are rich in starch, complex carbohydrates, dietary fibre and beta carotene. White yams (Dioscorea rotundata) are staple foods produced by annual and perennial vines. They are cultivated for their starchy tubers [3].It has been reported that processing caused a significant alteration in nutrient and chemical composition of tubers and the extracted starches, [4]. Therefore, proper processing is crucial to the nutrients and anti-nutrient of the final products. In recent years, several research studies have been carried out to establish the best processing method for some common tubers [5].
Generally, tubers undergo various processing methods (boiling/cooking, roasting, frying etc.). Boiling and roasting are common and they are the most important processing methods for tubers. However, there is lack of scientific information on the effects of these processing methods on nutrients, anti-nutrient and structural characteristics of tubers and their starches, Therefore, this present study was designed at characterising the effect of roasting and boiling on the nutrient, Antinutrients, phytochemicals, functional groups as well as the structural morphology of their extracted starches. The findings will suggest the better method of processing.

the tuber samples; white yam, sweet potato and cocoyam were bought from the main market in Akure, Ondo State. For boiling and roasting, 500g of each tuber were washed after peeling with distilled water to remove the dirt and adhering soils. The tuber samples were divided into 2 portions. Each of these portions were sub-divided into 3 parts, a part of the sub division boiled for 30mins, the second part roasted in a preheated oven to 350oC for 30mins and the third part unprocessed served as control. The boiled sample was drained and oven-dried (Gallenkamp Hotbox Oven, size2, Gallenkamp UK) at 1050C until constant weight. Hundred grams of the other second part was also milled into powdered form, soaked in 300ml of distilled water for 12hr and starch was extracted. All the samples (processed and raw) were milled into fine powder using electric grinder (NIMA-830Burma,Germany) until to pass through 0.425nm sieve, and finally packed into airtight polythene plastic bag and stored in the desiccator until required for analysis.

Chemicals such as Maltose, D-glucose, Sucrose assay kit and pure alpha-amylase enzyme were products of megazyme international, Ireland, UK. Acetic acid, sodium carbonate,chloroform,ferric chloride, dodecyl sulphate, potassium permanganate, ammonium thiocyanide, tannic acid were procured from BDH Chemicals Ltd. (Poole,Dorse,UK).All other chemicals used in this study were of analytical grade. Fourier transform infrared spectroscopy (FTIR) was done in National Research institute for Chemical Technology (NARICT) Zaria, Nigeria, while the scanning electron microscopy was done in Ahmadu Bello University, Zaria, and Kaduna State

Proximate analysis: Proximate analysis of the samples was carried out in triplicate, using the method of [6].Nitrogen was determined by micro-Kjeldahl method, described by [6] and percentage nitrogen was converted to crude protein by multiplying with 6.25.

Tannin: Finely milled and sieved flour solution was prepared by dissolving 200mg in 10 ml of 70% aqueous acetone extracted for 2hrs at 300C in a water bath using Gallenkamp orbital shaker at 120 r.m.p and filtered. The total polyphenols (as tannic equivalent) was determined in 0.05cm3 aliquot in test tube by the addition of distilled water to make to make it to 1.0 ml. This was followed by the addition of 0.5 ml of the Folin Ciocalteaus reagent (sigma St. Louis MD.USA) and the 2.5cm3 sodium carbonate solution. The tubes were vortexes and the absorbance recorded at 725nm after 40mins as described by [7]. The amount of total phenol (as tannic equivalent) was calculated from the standard curve, calibrated earlier obtained with pure tannic acid. Saponin: 20g of the samples were put into a conical flask and 100cm3 of 20% aqueous ethanol was added, the samples were
heated over a hot water bath for 4hrs with continuous stirring at about 550C.The mixture was filtered and the residue re-extracted with another 200 ml of 20% ethanol. The combined extracts were reduced to 40 ml over water bath at 90OC.The concentrate was transferred into 250 ml separatory funnel and shaken vigorously. The aqueous layer was recovered while the other layer was discarded .The purification process was repeated and 60 ml of butanol was added, the combined n-butanol extracts were washed twice with 10 ml of 5% aqueous NaCl, and the remaining solution was heated in a water bath. After evaporation, the samples were dried to constant weight; the saponin content was calculated as described by [8].
Terpenoid:0.5g of the milled sample was weighed into conical flask, 20 ml of chloroform and 10 ml of methanol were added and shaken .The mixture was allowed to stand for 15 min at room temperature and centrifuged at  000rpm.The supernatant was discarded and rewashed with 20 ml chloroform and 10 ml methanol and re-centrifuged. The precipitate was dissolved in 40 ml of 10% sodium dodecyl sulphate (SDS), 1 ml of 0.01M ferric chloride was added and allowed to stand for 30 min and the absorbance was read at 510nm as described by [9].

Oxalate: 1 g of the powered sample was soaked in 75 ml of 1.5 N H2SO4 for 1hr and filtered with Whatman No1 filtered paper. 25 ml of the filtrate was put inside conical flask and titrated with hot (80-900C) 0.1 M KMnO4 until a pink colour persisted as described by [10].

Phytin: For the quantification of Phytin, 8g of each finely ground sample was soaked in 200 ml 0f 2% HCl and allowed to stand for 3 hrs. The extracts were thereafter filtered through two layers of hardened filter paper and 50cm3 aliquot of the filtrate was pipette into 400cm3 capacity beakers before addition of 10cm3 of 0.3%ammonium thiocyanide solution as an indicator, and 107cm3 of distilled water to obtain the proper acidity (pH 4.5). The solution was then titrated with a standard iron chloride (FeCl3) solution containing 0.00195g (1.95 mg) Fe/cm3 until a yellow colour persisted for 5 minutes as described by [11].
Flavonoid content was determined by extracting 10g of the sample repeatedly with 100 ml of 80% aqueous methanol at room temperature. The whole solution was filtered through whatman filter paper No 42(125mm). The filtrate was later transferred into a crucible and evaporated into dryness over a water bath and weighed into constant weight as described by [12].

Phlobatannin: One gram of each sample was weighed and 10 ml of 4% methanolic vanillin solution was added and was allowed to stand for 1hr and decanted. One ml of 50% HCL was added to each sample. Blank was prepared by adding 10 ml of 40%methanolic vanillin to 1 ml of HCL, the mixture was allowed to stand for 15 minutes and the absorbance was read at 500nm Alkaloid: Five grams of the samples were weighed into 250 ml beaker and 200 ml of 10% acetic acid in ethanol was added, covered and allowed to stand for 4hrs. This was filtered and the extract was concentrated on a water bath to about 1/4 of the original volume. Concentrated NH4OH was added drop wise until the precipitate was complete. The solution was allowed to settle and the precipitate was collected and washed with dilute NH4OH and then filtered. The residue was dried and weighed as described by [13].

Determination of sugars: The principle of the method is based on an enzymatic cleavage of the disaccharide and specific measurement of the resulting glucose using an enzymatic procedure described by [14].
Method Specificity: The α- glycosidase hydrolyses the terminal, non-reducing 1, 4-linked α- D-glucose residue with the release of α-D- glucose

Standard curve: 50μg of anhydrous D-glucose was dissolved in 1000 ml of ethanol (48%). Dilutions were prepared ranging from 0.01-0.05 mg/ml to obtain the standard curve. Four ml of glucose oxidase/peroxidase enzyme solution was added and incubated for 1hr at room temperature (200C). The wavelength was set at 450nm.
Sample preparation: 1g of the milled vacuum dried sample (particle size 0.8mm) was suspended in ethanol (46%) to make up to 100 ml of suspension; the aim of the alcohol treatment was to precipitate the starch and proteins. The system was extracted in the shaker at room temperature during 40min.Afterwards the suspension was filtered by paper media filter 0.22μm. Glucose analysis: 1 ml of the filtered solution was mixed with 1 ml of buffer (pH 6) and 4 ml of oxidase/peroxidase (5.6g/100 ml) as described by [14]. The mixture was incubated for 1hr at room temperature and the absorbance read at 450nm.

Maltose analysis: 1 ml of the filtered solution was mixed with 1ml of acetate buffer (pH6.6) and 0.2 ml of α-glucosidase (240μ/ml), the mixture was incubated for 1hr at room temp according to the procedure of [15] .Afterwards, 0.1 ml of the obtained solution was mixed with 1.9 ml of water and 4 ml of glucose oxidase/ peroxidise (5.6g/100 ml) as described by [14]. The incubation process was 1hr at room temperature, and the absorbance was read at 450nm.
Sucrose analysis: 0.3 ml of the filtered solution was mixed with 1 ml of acetate buffer pH 4.6 and 0.640 ml of invertase (150 μ/ml), the mixture was incubated for 1hr at room temp and 0.5 ml 0f 0.5M tris buffer was added as described by [16]. Afterwards, 0.6 ml of the obtained solution mixed with 1.4 ml of water and 4ml of glucose oxidase/peroxidase (5.6g/100 ml) as described by [14]. The mixture was incubated for 1hr at room temp and the absorbance read at 450nm.
The different sugar concentrations were calculated with the empirical deduced equations

C 0 =Free concentration of glucose (without adding α-glucosidase or Invertase):
Cm= Glucose concentration after adding α- glucosidase
Cs =Glucose concentration after adding invertase

Fourier Transform infrared Analysis (FT-IR): The sample pellets were prepared by mixing 100 mg of KBr (sigma Aldrich, FT-IR grade) with 5 g of the sample. The samples were placed in an evaluable KBr dye and a 13 mm clear disk was pressed in a hydraulic press which formed KBr pellets. The pelletized samples was placed in cell holders (universal demountable cell) and were inserted into the FT-IR (Model, Shimazu FTIR-8400S Fourier transform infrared spectrophotometer) and scanned at a range of 350cm-1 -4000cm-1.

Starches were sieved using a No 60 fisher sieve mounted and coated with gold (1 nm) using a polaron sputter coater and analysed using Burker scanning Electron Microscope. Starch granules for birefringence test were suspended in water and viewed with a Leica DMRME polarized microscope.
Data analysis: The results of the triplicate experiments were analysed and expressed as mean +/- standard deviation (SD). Mean was compared by one-way analysis of variance (ANOVA) followed by Duncan's multiple range test, and least significant differences were carried out accepted at p≤0.05.

Proximate composition (Table 1) of the processed and unprocessed (control) tubers revealed that the moisture content ranged from 9.59 g/100g in fresh white yam to 11.86g/100g in boiled cocoyam. There was slight difference between the processed and the control. These values were lower than those of sweet potatoes 14g/100g reported by [17]. The tubers had low moisture content, below the value of 15% which was reported by [17] not to favour microbial activities during storage. The low moisture content of the sample is an indication of good storage property with minimum fungal and bacterial attack. The crude fibre (CF) was highest (3.35g/100g) in the fresh sweet potato and lowest (1.73g/100g) in fresh cocoyam; these values were low in all the processed samples except in cocoyam where the fresh had (1.73 g/100g) compared to (1.99 and 2.41g/100g) in boiled and roasted respectively. The values in the present study were lower to 7.20g/100g in sweet potato reported by [18].Fibre is important in food because it aids digestibility, maintains internal distension of the intestinal tract thereby aiding peristaltic movement. The recommendation daily allowance (RDA) is 18-35g, [19]. This  indicates that 100g of the tubers under study can only provide 1.73 to 3.35g of daily fibre to the body. Crude (CP) protein content in the tubers ranged 4.45g/100g in the boiled cocoyam to 9.34g/100g in the fresh white yam. The result was lower than that on white yam Germplasm (15.55g/100g) reported by [20]. A report by [21] stated that heating food that contain protein causes several structural changes to the protein as well as the linkages, however, the nutritional values and the digestion of the food does not change. The recommended dietary allowance of protein as reported [22], for children, adult male, adult female and pregnant women were 28, 63, 50, and 60g respectively .Therefore, for every 100g each of the tubers analysed will give a range of 4.45 to 9.34g of proteins, which indicates that these tubers especially boiled are poor sources of daily proteins.
The lipid ranged (0.63g/100g) in roasted white yam to (2.98g/100g) in fresh white yam. These were lower than those of taro (Colocasia esculenta) 3.50g/100g but within the range of (1.97g/100g) for sweet potato reported by [18]. Consumption of dietary fat and oils are the principle sources of energy but should not exceed the daily recommended dose of 30 calories/day [23] in order to prevent obesity and other related fat diseases. One gram of lipid provides 8.37Kj, which indicates that 100g of each tuber provides in the range (0.08-0.37 Kcal).

The ash content of the sample is a measure of its inorganic mineral content. The ash of the tubers ranged between 2.08g/100g in boiled sweet potatoes to 5.53g/100g in roasted cocoyam. The observed increase in ash especially in roasted may be due to concentration due to dry heating. The values obtained were higher compared to (1.8g/100g) reported [24] on white yams (Dioscorea cayenesis-rotundata) but lower than that in cassava (8.22g/100g) reported by [25].
The nitrogen free extract (NFE) of all the tubers analysed were not significantly different p≤ 0.05 except in fresh white yam. The carbohydrate in all tubers were high and ranged between 72.08g/100g in fresh white yam to 76.46g/100g in fresh cocoyam, these values were lower than 82.8g/100g reported by [25] for cassava. The main function of carbohydrates and lipids is to provide the body with energy.
In general, among all the parameters measured processing significantly P≥ 0.05 reduced moisture, fibre and crude protein while the ash content in roasted samples increased Results on anti-nutritive and phytochemicals (Table 2) revealed that tannin ranged 0.52mg/g in boiled cocoyam to 1.54mg/g in roasted white yam; these values were lower than 17.0mg/g reported by [4] in wild yam. Processing especially boiling significantly p≥0.05 reduced the tannin content in the samples.
Tannins in food impose an astringent taste affecting palatability, reduce the intake of the food and consequently body growth. Tannins can bind to both exogenous and endogenous proteins for tannin-protein complexes. Tannin-protein interactions are most frequently based on hydrophobic and hydrogen bonding. Tannin phenolic group is an excellent hydrogen donor that forms strong H-bonds with the protein's carboxylic group. To have high protein affinity, tannins must be small enough to penetrate interfibrillar region of protein molecule but large enough to crosslink peptide chains at more than one point. However, tannins act as defence mechanism in plants against pathogens, herbivores and hostile environmental conditions.
Saponins were observed to be the most abundant Antinutrients in all the tubers analysed as high as 169.27mg/g in fresh cocoyam. Saponins are glycosidic compounds composed of steroid (C-27) or triterpenoid (C-30) nucleus with one or more carboxylate branches and most Saponins are bitter in taste and toxic substances, [26]. Concentrations of 3-7% are represented as powerful poisons while at 1% they are known as inoffensive; at 1.5% some biological activities on damaged mucus membranes are observed. Saponins are highly heat labile as shown by its remarkable reductions especially in boiled sweet potato and boiled white yam (93.36 to 17.36 mg/g and 144.73 to 95.27mg/g respectively).
Terpenoid showed a decrease upon processing as shown by the ranged 1.95-1.31 mg/g; 4.09-1.74mg/g; 5.22-3.04mg/g in sweet potato, cocoyam and white yam respectively, however, highest reduction was found in all the boiled samples. Terpenoids are primary constituents of the essential oils of many plants and flowers; it displays a wide range of biological activities against cancer, inflammation and variety of infectious diseases [27]. However, despite the health benefit of these Terpenoids it becomes toxic at the levels higher than 20ppm in food samples [27].
The oxalate content of the tubers ranged 0.54-1.08 mg/g in sweet potato, 0.72-1.26 mg/g in cocoyam and 0.36-0.86 mg/g in white yam. These results were lower when compared to 5.34mg/g in wild yam as described by [4], but similar to 1.20mg/g in sweet potato reported by [18]. The levels of oxalate in all the tubers analysed were within physiological tolerance of 2.5g [4].
Oxalate can bind to calcium present in foods thereby rendering calcium unavailable for normal physiological and biochemical roles. Oxalate present in food is insoluble; it may also precipitate around soft tissue such as kidney causing kidney stones, [28]. Remarkable reduction was found in all the samples after boiling, this may be as a result of cell rupture which facilitated the leakage of soluble oxalate into cooking water.
Phytate content in all the tubers analysed was highest 22.25 mg/g in fresh cocoyam and lowest value of 6.59mg/g was found in roasted white yam. These results were lower than those reported by [29] on Anchote (Cossinia abyssinica) tubers 33.36mg/g (boiled) while this range compared well with 8.72-20.2mg/g reported by [30], on differently processed T.leontopetaloides flour. Phytates are common anti-nutrients in plants foods, which complexes Ca, Mg, Fe and Zn and decrease their bioavailability. Meanwhile roasting and boiling processes significantly (P≤0.05) reduced the phytate content in all the tubers with highest reduction observed in roasted samples.
Flavonoid ranged (2.23mg/g) in roasted sweet potato to (3.21mg/g) in raw of the same sample; it was below detection level in all the cocoyam samples and the roasted white yam, while the raw sample had (3.90mg/g) and boiled (3.07mg/g). Flavonoids are widely distributed group of polyphenolic compounds with health related properties, which are based in their antioxidant activity. Interestingly, there is no evidence of side effects associated with dietary intake of flavonoids. The reason could be attributed to their low bioavailability, however, flavonoids in the supplement form do have side effects and sometimes severe ones and therefore at times referred to as Antinutrients, and for example, they can cause nausea, headache or tingling of the extremities in some people when taken in doses of 1000mg/day [31].
Decreased in phlobatannin was observed after processing in cocoyam and white yam flours while it was below detection level in sweet potato flours. The values in cocoyam flour ranged 1.3 to 1.98mg/g and 1.53 to 1.69mg/g in white yam flours, however, the least values were found in all the boiled samples .Phlobatannin is tannin which is formed under action of dilute acids to yield phlobaphene.

Alkaloids were observed to be most representative antinutrients in tubers; interestingly levels were below detection limit in the studied raw and processed sweet potato while cocoyam and white yam ranged 15- 44.58mg/g, 15.20-32.04 mg/g respectively. These values were below the maximum limit of 200mg/kg released by [32]. Processing methods lead to about 40% reduction in cocoyam and white yam. Alkaloids generally include those basic substances which contain one or more nitrogen atoms, usually in combination as part of a cyclic system. Although some biological activities are attributed to some secondary metabolites, alkaloids are often toxic to humans and many have dramatic physiological changes such as trembling, shaking, excitation and convulsion due to loss of co-ordination and muscular control [32].
(Table 3) shows the extracted starches before addition of enzyme (undigested).The concentration of the glucose ranged 2.09 g/100g in fresh white yam to 10g/100g in roasted cocoyam. In the digested starch, glucose was lowest (6.06 g/100g) in fresh sweet potato and the highest value of 15.25 g/100g was found in roasted cocoyam. Sucrose level in the undigested tubers starches as well as the digested starches were not significantly different p≤0.05 for example fresh, boiled and roasted values in undigested and digested sweet potato were 1.24 and 1.25 g/100g; 4.66 and 4.69 g/100g ;6.12 and 6.10 g/100g respectively. The roasted samples of all the tubers (digested and undigested) had highest values of glucose. This same trend was found in sucrose and maltose and the values were significantly (p≥0.05) different. These results are in agreement with [33] on sweet potato, but, higher than those reported by [24] on Dioscorea cayensis rotunda. The results on the sugar compositions showed that sucrose, glucose and maltose concentrations were lower in the raw samples, and after processing (heat application) these values increased. This may be due to the fact that at higher temperature range, complex carbohydrates in the starches granules which are made up of long chains of glucose units are broken down and this may lead to an increase in amylose levels [34]. In addition, alpha amylase enzyme breaks down starch into sugars, generally into monosaccharide glucose and disaccharide maltose (double glucose). Slight changes was found in sucrose digested and undigested because sucrose is a disaccharide of glucose and fructose, the amylase enzymes are not keyed for the pair and thus cannot split it up [35]. Increase in the level of sugars in roasted compared to the boiled may be due to the effect of water vaporisation because no external water added during roasting which lead to the increase in the concentration of the amylose [35] .The decrease in the values of amylose in the boiled samples compared to roasted may be as a result of hydrolysis and leaching reactions.
The infrared spectrum analysis (Figure 1a-1i) revealed that both raw and processed tubers had similar band positions which meant that identical compounds were observed which were mainly alcohol, alkane, alkene, alkyne, Alkyl, aliphatic amine and ether at 3415cm-1, 3410 cm-1, 2933-1364 cm-1,1644-1643 cm-1, 2268-2264 cm-1, 1249-1022cm-1bands respectively. Broad band of carbonyl compounds 1717-1786cm-1 which was observed in the fresh and boiled tubers disappeared in the roasted this changes may be adduced to the milliards reaction and the formation of brown colour in roasted tubers. The results on spectrum showed that no high molecular compounds formed due to heat reactions, which may lead to increased levels of radicals in the system based on iron and copper-catalysed Fenton reactions.

Scanning electron microscopy (SEM) (Figure 2a-2i) was used to visualise and monitor the structural morphology of the processed tubers extracted starches. The result showed that fresh tuber starches had smooth surface, without any pore, except for occasional cracks, reason being that fresh tuber samples contained moisture content that maintained and supported the structure of the granules; it was therefore, smooth and had more granules. All the boiled samples had an irregular shape with smooth surface while the roasted had an irregular shape with rough surfaces; these findings are in agreement with properties of starches from cocoyam and cassava reported by [25]. Starch granules ordinarily is resistant to water and hydrolytic enzymes due to hydrogen bonding. Heat treatment due to boiling and roasting weakens the inter and intra hydrogen bonding and the starch granules swells this is known as gelatinization. Particle size and specific surface area were observed to play key roles in the digestibility of tuber starches [36]. From all the samples, it was observed that all the processed samples especially starches from boiled samples had more surface area followed by the roasted, therefore, higher digestibility may be achieved in boiled samples compared to roasted.

Result on proximate showed no pronounced changes among the processing methods. Antinutrients and phytochemical reductions were more pronounced in the boiled tubers. FTIS analysis showed no formation of higher molecular compounds while results on SEM showed increased surface area in boiled tubers. However, the research concluded that it was preferable and healthier to consume boiled tubers due to higher digestibility and reduction in sugars after processing. This may reduce cases of blood sugar related diseases such as diabetes when consumed.

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