3 RESULTS AND DISCUSSION
The functional and pasting properties of the native and fermentedacha flour and cookies samples were presented in Table 1. The loose and tapped bulk density of Acha flour and cookie samples was observed to decrease significantly (P<0.05) with increased fermentation duration. The values ranged from 0.490 to 0.568 g/ml and 0.777 to 0.842 g/ml respectively for the flour sample, and 0.494 to 0.527g/ml and 0.710 to 0.867g/ml for the cookie samples. The highest value was observed in native acha flour and cookie samples while least value was observed in sample fermented for 72 h. The decrease observed in the loose and tapped bulk density might possibly result from increased particle breakdown of the fibrous materials during fermentation (Olapade et al., 2014). According to Nzigamasabo and Hui (2006), pectinolytic and cellulotic enzymes are elaborated by fermenting microorganisms and aid in the breakdown of cell membranes. The formation of carbon dioxide gas during fermentation, which causes the bubbles to grow in size and increase in volume may also be the reason of the decrease in bulk density of flour samples with longer fermentation durations (Elmehdi et al., 2007). The cookie sample’s low loose bulk density suggests that less food would be packaged in a constant volume, which ensures cheap packaging. The ability of flour to absorb water and swell, improving food consistency, is known as its water absorption capacity. In food systems, WAC is desired to increase yield, consistency, and food body (Osundahunsiet al., 2003). There was significant (P<0.05) decrease in the water absorption capacity of the flour (1.08 to 1.29g/g) and cookie samples (0.76 to 0.89g/g) with increase in fermentation duration. Highest value was observed in native acha flour while least value was observed in sample fermented for 72 h. The alteration of macromolecules during fermentation may be the cause of the samples’ decreased water absorption capacity (Oloyede et al. , 2016). The result is consistent with the pattern observed by Ogodo et al . (2018) in their research on lactic acid bacteria-fermented soybean flour and Ntsamo et al. (2020) on fermentation on sorghum flour properties. Giami and Bekebain (1992) also found that longer fermentation durations greatly reduced the microbial activity of food products that have low water absorption capacities. As a result, the product’s shelf life would be increased. The oil absorption capacity ranged between 0.72 – 0.64 g/g for the flour sample, and 0.65 to 0.77g/g for the cookie sample. With increased fermentation duration, there was no significant (P>0.05) increase in the samples’ oil absorption capacity. Acha flour fermented for 72 h had the highest oil absorption capacity value while native acha flour had the least value. Intrinsic variables like as protein structure, hydrophobicity or surface polarity may be responsible for the increase in oil absorption capacity with increased fermentation durations (Chandra and Samsher, 2013). The study of Oloyede et al. (2015) and Periago et al. (1998) on pea flour and moringa seed flour respectively, showed similar findings. Flour samples with good oil absorption capacity are suitable for food preparations that require oil mixing, such as bakery products, where oil is an essential ingredient (Sobowale et al ., 2021). The dispersibility of the flour and cookie samples varies significantly (P<0.05) as the values increased with increase in fermentation duration. Acha flour fermented for 72 h had the highest dispersibility value while nativeacha flour had the least value. Higher dispersability value enables the flour to reconstitute more readily in water (Adebowaleet al . 2008). This indicates that the flour samples’ capacity to reconstitute effectively in water has been improved by fermentation. This suggests that the cookies samples could digest easily (Adebowaleet al ., 2008). The swelling capacity and solubility index of the flour and cookie samples decreased with an increase in fermentation period. This might be related to the granules degree of association and binding force (Hoover, 2002). The values obtained in this study for the functional properties of cookies conform to the report of Razzaqet al. (2012).
It has been established that the most assessed set of flour quality attributes is its pasting properties, which accurately indicate the quality of the flour (Offia-Olua, 2014). With an increase in the duration of fermentation, there was a significant (P<0.05) decrease in the peak viscosity of the flour samples. The achaflour sample that was fermented for 72 h had the lowest value, while the native acha flour had the highest value. The similar phenomena have been reported for rice flour by Min et al. (2004), Yong and Tao (2008), Lu et al. (2005), and soybean flour by Olanipekun et al. (2009). The ability of the starch granules in the flour sample to bind water is indicated by the decrease in peak viscosity of the flour samples with an increase in fermentation duration. As shown in Table 1, the peak viscosity of the cookie sample increased significantly (P<0.05) with increase in fermentation duration. High peak viscosity also indicates the fragility of the swelled granules, which swell initially before disintegrating due to the Rapid Visco Analyzer constant mixing (Adebowale et al ., 2008; Sanni et al ., 2006). The trough viscosity values of the flour sample decreased gradually with an increase in fermentation duration. The decrease in trough viscosity with increase in fermentation duration may be due to the swelling capacity of the starch granules in flour samples (Ohizuaet al. , 2017). The flour samples generally had high trough values, indicating higher cooked paste stability and a higher ability to withstand shear at elevated temperatures (Farhat et al. , 1999). The trough viscosity values of the cookie samples increase significantly (p<0.05) with increase in fermentation duration. The breakdown viscosity values of the flour samples decreased with an increase in the fermentation duration. The highest break down value was obtained in native acha flour while the least was obtained in sample fermented for 72 h. A study by Chinma et al . (2010) also revealed that low breakdown values suggest that the starch in question has cross-linking characteristics whereas high breakdown values show that the swollen starch granules are relatively weak against hot shearing. The lower breakdown viscosity values of flour samples obtained in this study indicate high fermentation duration impact into the flour sample ability to possess cross-linking properties and also high stability against hot shearing. The breakdown values of the cookie sample increased significantly (p<0.05) as fermentation duration increased. Better palatability is indicated by a higher breakdown viscosity (Tren et al ., 2001). The final viscosity values of the flour was observed to decrease significantly (P<0.05) with increased fermentation duration. The simple kinetic effect of cooling on viscosity and the re-association of starch molecules in the samples may be the cause of the change in the final viscosity with longer fermentation durations (Ikegwu et al ., 2009). The relatively high value of final viscosity of the acha flour samples suggests the ability of the flour samples to form more viscous pastes. The final viscosity values of the cookie samples increased significantly (P<0.05) with increase in fermentation duration. The final viscosity changes as fermentation time increases and this was due to high amylose content of the cookie samples and hydrogen bonding during cooling. (Alais and Linden, 1986 cited by Iwe et al ., 2016). The retrogradation of the flour-based product during cooling decreases with increasing setback value (James and Nwabueze, 2014). As the duration of fermentation increased, the setback viscosity values of the flour samples decreased significantly (P<0.05) reduced. The setback viscosity value was highest in native acha flour while the least value was observed in sample fermented for 72 h. This indicates that increase in fermentation duration has improved the retrogradation tendency of the flour samples. When cooling paste made from starch or a starch-based product, low setback suggests increased resistance to retrogradation (Sanni et al., 2004), while high setback is linked to syneresis or weeping during freeze/thaw cycles (Adebowale et al., 2005). The setback values of the cookie samples increased with increased fermentation duration. It has been reported that setback viscosity has a major impact on how easily a food product can be digested when consumed. Higher setback values are correlated with lower digestibility (Shittu et al ., 2001). The result indicates that increased fermentation duration decreases the digestibility of the cookies samples. The peak time of the flour samples increased significantly (P<0.05) with increase in fermentation duration. This shows that increased fermentation duration also increases the cooking time of the acha flour samples. The peak time values are comparable to those of Adebowale et al. (2008) for instant yam-breadfruit composite flour and Chinma et al . (2007) for germinating tiger-nut flour. The peak time increased with an increase in fermentation duration. One of the properties that show the minimum temperature needed to cook a sample, the energy costs involved, and the stability of other components is the pasting temperature (Iweet al ., 2016). The pasting temperature of the flour was observed to increase with increased level of fermentation duration. A greater pasting temperature is indicative of a larger ability to bind water, a higher inclination to gelatinize, and a lesser swelling property in starch-based flour because of the strong associative forces that exist between the starch granules (Adebowale et al., 2008). Same observation was reported by Danbaba et al. (2012) for the pasting properties of ‘ofada rice’ varieties. Generally, the values obtained for the pasting properties of flour samples are within those recommended for commercial flour (Soni et al., 1993; Kemas et al., 2012). The pasting temperature of the cookie samples differs significantly (P < 0.05) as the values increased with increase fermentation duration. This implies that fermentation duration has increased the cooking temperature for cookies samples since pasting temperature is an indication of minimum temperature required to cook a sample (Iweet al ., 2016).
Table 2 shows the nutritional quality (proximate and mineral) of native and fermented acha flour and cookie samples. Significant differences (p<0.05) were observed in fat, fibre and protein content of the samples. The moisture content of the acha flour and cookies sample was observed to decrease with increase in fermentation duration. The samples’ decreased moisture content with longer fermentation times might likely be attributed to bacteria using some moisture for metabolic processes (Igbabul et al. , 2014). According to the work of Hwabejire et al. (2023), the moisture content of spontaneously fermented acha flour was observed to increase within 48 h of fermentation time and decreased by 72 h. The decrease in moisture content could be due to the breakdown of complex carbohydrates into simpler sugars, which are then used by microorganisms during fermentation. The moisture content of the cookie’s samples was observed to decrease generally after baking when compared to theacha flour samples. Food stability and value are indicated by the amount of moisture in the food (Nonaka, 1997). The moisture content values of the cookie samples found in this study fall within the World Food Programme’s (WFP) maximum moisture content for flour baked products for effective storage, which is set at 10%.
The protein content of the acha flour samples increased with increase in fermentation duration. It is possible that the increased microbial mass during fermentation led to the extensive breakdown of protein molecules into amino acids and other simple peptides, leading to why the protein content of the flour samples was reported to increase (Okechukwu et al. , 2015). The result of protein content observed corresponds to the observation of Michodjehoun-Mestres et al ., (2005) on increase in protein content during fermentation of sorghum, and the work of Igbabul et al. (2022) on mahogany bean flour. The protein content of the cookies was observed to decrease when compared toacha flour samples after baking and the values varies significantly (P<0.05). The decrease in protein content observed in cookies samples could be because of denaturation of protein due to baking temperature (Ogundele et al. , 2019).
The fat content of the acha flour samples was observed to decrease with increased fermentation duration. The breakdown of lipids during fermentation may have led to the low fat level found inacha flour samples (SefaDedeh and Kofi-Agyir, 2002). The reduced fat content contributes to an extended shelf life for acha flour by minimizing the likelihood of developing rancid flavors. The fat content of the cookie samples increased after baking when compared to the flour samples. The high fat content observed in cookies samples than the flour samples could be because of baking fat added as an ingredient during dough preparation and agreed with other authors (Caponio et al. , 2008). The ash content of the acha flour and cookie samples was observed to increase initially at fermented period of 24 h, after which the ash content decreased with increase in the fermentation duration. Highest value was observed in sample fermented for 24 h, while the least value was observed in samples fermented for 72 h shows to have no significant effect on the ash content of the acha flour samples, though numerical differences was observed. The reduction in ash content following a 24 h fermentation period may be attributed to the leaching of soluble mineral elements into the medium or to the general activities of the fermenting microorganisms, whose enzymatic activity broke down the food components into forms that could be absorbed (Igbabul et al. , 2014).
The crude fibre content of the acha flour samples and cookie sample values decreased with increase in fermentation duration. Highest value was observed in native acha flour/cookie while the least value was observed in samples fermented for 72 h. Due to the actions of microorganisms known for the bioconversion of carbohydrates and lignocelluloses, the fibrous tissues of the acha seeds may have softened during fermentation, which could account for the decrease in crude fiber content of the acha flour samples with longer fermentation periods (Igbabul et al ., 2014). This result correlates with that of Forsido et al., (2020) who observed a decrease in the crude fibre content of cereal-based complementary foods, as fermentation duration increased. The fibre content of the cookies varies significantly (P<0.05) and was not appreciably affected by baking. All the cookies’ fiber content, however, fell below the FAO/WHO (1994) suggested range of no more than 5 g of dietary fiber per 100 g of dry matter, which is in line with Okpala et al. (2013). The flour samples’ comparatively high carbohydrate content indicates that they may be an energy source. The result observed in this study correlated with those observed by Igbabul et al ., (2014) for fermented cocoyam flour.
The mineral composition revealed that calcium, sodium and zinc content of the acha flour and cookie samples varies significantly (P<0.05). The calcium, sodium and zinc content of the samples increased initially, but fermentation duration above 24 h cause a decrease in the mineral content of the samples with increase in fermentation duration. Highest value for calcium, sodium and zinc was observed in sample fermented for 24 h while the least value was observed in sample fermented for 72 h. The decrease in the mineral content of theacha flour samples with increased fermentation duration might be attributed to leaching of the mineral elements in the fermenting water during prolong fermentation (Assohoun et al., 2013). However, there was no significant (P<0.05) difference among the samples in terms of magnesium, copper and phosphorus. This shows that fermentation duration has no significant effects on the magnesium, copper and phosphorus content of the flour samples, although numerical differences were observed. The magnesium, copper and phosphorus content increased initially, with fermentation duration above 24 h, the mineral parameters decreased with increased fermentation duration. Assohounet al. (2013) also reported that possible microbial utilization and conversion could be responsible for the reduction in the levels of minerals during prolonged fermentation. Similar results were found in Samitya et al, (2021) on plant-based foods. The mineral content of the cookie samples was lower when compared to the flour samples. The baking process was observed to slight decrease the mineral content of the cookie samples except for the sodium and copper content which was observed to increase after the baking process. This implies that both Na and Cu are very heat resistant minerals, and that baking may have made them slightly more extractable. This observation can be attributed to the Maillard reaction that occurred during baking. Delgado-Andrade et al. (2011) stated that the Maillard reaction can result in disintegration or creation of compounds that may have an impact on the availability and solubility of minerals. Despite the trend observed in the mineral content, the cookies samples still contain appreciable quantities of mineral elements that can supply the recommended daily allowance of the mineral elements as stated above. The same results for extruded acha -soybean blends and fermented popcorn-African locust bean blends were obtained by Anuonye et al. (2010) and Ijarotimi and Keshinro (2013). The phenolic and flavonoid compounds of the fermented flour and cookies samples were shown in Tables 3. The values of phenolic and flavonoid compounds increased significantly (p<0.05) with increased fermentation duration. According to research, consuming phenolic compounds may have a significant impact on health, since it regulates metabolism, chronic disease, weight and cell proliferation (Cory et al ., 2018).
The Fourier transmission infrared (FTIR) spectra of fermentedacha flour and cookies are shown in Table 4. Fourier Transmission-Infrared (FTIR) spectroscopy is a technique for studying constituent functional groups in an analyte (Adebiyi et al.,2016). The infrared (IR) spectra of the native and fermented achaflours and cookies samples showed similar peaks with variations in intensity. The FTIR spectra of the flour and baked cookie samples reveal the changes brought about by the fermentation process. The peaks were between 3860 - 3650 cm-1 (Figure 1). The high peak range observed could be because of O–H bond stretching (Sun et al., 2014). The O–H absorption peak of native acha flour changed from 3650 cm-1 to 3670, 3830 and 3800 cm-1 for 24 h, 48 h and 72 h fermentation period, respectively. These changes could be attributed to better lipophilic and hydrophilic properties of fermented flour samples (Animashaun and Sobowale, 2024). A similar pattern was also noted in the cookie samples, where the O-H peak varied for the 24, 48 and 72 h fermentation periods, respectively, from 3610 cm-1 to 3800, and 3830 to 3860 cm-1. The degree of inter and intra-molecular bond formation is shown by the fluctuation in the O-H peak (Sun et al. , 2014). However, the increase in the width of the fermented samples spectrum in both flour and cookies could be as a result of characteristic presence of alcohol, produced during fermentation (Adebiyi et al. , 2016). Tightly bound water in the form of moisture was the cause of the asymmetric stretching of the C–H band in the region of 3030 – 2980 cm-1 and the vibration peak at 1490 – 1470 cm-1 observed in the spectrum (Shiet al., 2012). The carbonyl stretches peaks in the native and fermented acha flour samples ranged between 1760 and 1770 cm-1. However, the carbonyl peak values observed in the cookie samples were at higher intensities in the absorption range of 1780 –1770 cm-1. It was observed in Figure 1 that fermentation process reduces the carbonyl peaks which might be due to a decrease in the total lipids present in the samples as a result of fermentation (Correia et al. , 2005; 2008). The additional fat added during dough preparation may have contributed to the greater carbonyl peaks seen in the cookie samples. However, baking depolymerized the fat, resulting in a strong carbonyl peak in every cookie sample. C-O bond stretching and aliphatic C-N stretching are responsible for the peaks at 1180 – 1181 cm-1, whereas the amide I band formed in the 1550–1570 cm-1 range. The intensity of the bands increased from 1550 cm-1 in the nativeacha flour samples to 1560 and 1560 to 1570 cm-1 in the fermented flour samples after 24, 48 and 72 h, respectively. This indicates alterations in the samples crystallinity. There may have been a buildup of proteins in the processed samples as indicated by the fermented samples’ comparatively greater amide I peak intensities (Correia et al., 2005). Increases in the amide peak intensities were also visible in the cookie samples’ spectra. This might be due to the embedded proteins β-sheet structures (Duodu et al., 2001). This study is comparable to those published by Animashaun and Sobowale (2024) and Correia et al. (2008) on the IR spectra for maize and sorghum respectively. Figure 2 shows the X-ray diffraction (XRD) pattern for native and fermentedacha flour and its resultant cookie. The variations in the crystallinity of the flours and cookies affected by processing are examined using X-Ray Diffraction (XRD) patterns. Peak intensity relates to the number of semi-crystalline structures that are arranged and/or the variation in electron density between lamellae that are crystalline and those that are amorphous. (Hamley, 2013), while the sharp peaks correlated to crystalline region, the diffused peaks amounted to the amorphous region of the flour and cookies samples (Hamley, 2013; Adebiyiet al., 2016). The diffraction patterns for the native and fermented acha flour samples shows that all the flour samples exhibited an A-type XRD pattern (Sun et al., 2014). Nativeacha had a diffraction pattern with major reflections at 2θ of 16.2o, 17.9o, 19.0o, and a diffused peak at 22.8o. Flour samples fermented for 24, 48 and 72 h had similar peaks with stronger diffraction intensities with peaks occurring at 2θ of 15.3o, 17.4o, 20.2o and 22.9o, 15.5o, 17.5o, 19.4o and 23.2o, and 15.3o, 18.0o, 19.4o and 23.1o respectively. The fermented flour blends’ XRD patterns showed variation, which might be attributed to hydrolysis that occurred during the fermentation process (Lu et al., 2005). Other cereal flours exhibit the observed A-type structure (Amadou et al. , 2014; Adebiyi et al., 2016). The X-ray diffraction plots show that the crystallinity in cookies samples has little variation when compared to the flour samples. The diffused V-type peaks seen in all samples are commonly found in cereals that have been processed (Dharmaraj et al.,2014). This explains why all of the cookie samples showed increased thermal degradation and the disintegration of the crystalline region of the flour samples to an amorphous zone. The disruption of crystalline region is due to heat processing resulting to a partial or complete reorganization in the crystalline alignment (Sun et al. , 2014). Similar result was reported by Adebiyi et al., (2016) and Dharmaraj et al ., (2014) for malted and fermented pearl millet biscuit and during heat treatments resulted in a decrease in crystallinity of acha flour and cookies, respectively.
The scanning electron microscope (SEM) image for native and fermentedacha flour and corresponding cookies samples is presented in Figure 3. The internal structure of food samples is commonly evaluated and characterized using scanning electron microscopy (SEM). SEM has been utilized by numerous authors to characterize the morphology of starch granules, explain the endosperm’s structural arrangement, and examine how distinct treatments affect the granules (Lauro et al., 2000; McPherson and Jane, 2000; Perera et al., 2001; Naito et al., 2004; Wilson and Betchel, 2004). The scanning electron microscope (SEM) images of native and fermented acha flour samples were compared to observe the morphological changes in them. It was observed according to flour micrographs that the granules of native acha flour have a non-uniform, smaller, lenticular or disk-like shape, which is spherical or ellipsoidal in shape. Regular and smoother structural networks were observed with shape and structure almost the same to native achaflour in the fermented samples. The granules had smaller pores and seemed to be round or polygonal in shape. This suggests that fermentation process had an effect in forming a regular shape in their corresponding samples. The effect of osmotic stress during the fermentation process is responsible for the fermented samples’ increased porosity, disorientation, and less condensation as seen in the SEM images (van der Weerd et al., 2001). The spherical or ellipsoidal shape of the granules is like the shape of granules in wheat and corn starches reported by Horstmann et al. (2017). The scanning electron microscopy images of the cookie samples give a more varied arrangement of the granules when compared to the flour samples. The baking process has been shown to disrupt the granular structure as all the cookies samples had larger granular sizes with a “honeycomb like” structure. Similar observations were reported by Dhamaraj et al.(2014) and Adebiyi et al. (2016) for high temperature processed finger millet and pear millet respectively. The high structural deformation of the samples may be because heat applied during baking causes a higher diffusion rate. High-temperature levels, while baking led to intensified water evaporation and melting of starch granules (Izli and Polat, 2019). In general, the baking procedure stabilized the colloidal batter system, enabled the formation of an enveloping crust, and altered the structural nature of the cookie samples. This situation may undermine the starch-protein matrix and hence give rise to structural damage and finally rupture the compact structure of the samples. The micrograph of native and fermented acha cookies was in close conformation with the findings of Vega-Gálvez et al . (2015) for high temperature processed products structure.
The sensory attributes of cookies produced from native and fermentedacha flour is presented in Figure 4. There were significant (P<0.05) difference among all the samples produced from native and fermented acha flour. In terms of appearance and crispness, the mean score shows decrease with an increase in the fermentation duration. Highest mean score was observed in cookie produced from nativeacha flour, while the least mean score was observed in sample produced from acha flour fermented for 72 h. Perhaps, decrease in the mean score for appearance and crispness with increased fermentation duration could be attributed to dark colouration developed because of mallard reaction during baking (Hussain et al., 2006) and increase in mechanical force required require breaking the samples due to increase in hardness of the samples facilitated by decreased moisture content of the flour samples, respectively (Guinard and Mazzucchelli, 1996; Luyten et al ., 2004). Also, it was observed that there is decrease in taste and aroma with increased fermentation duration. The panelist rated the cookie produced from acha flour fermented for 24 h and sample produced from native acha flour the most preferred in terms of taste and aroma, respectively. There was significant difference (P<0.05) among samples in terms of color. The panelists preference for the colour of the cookies decreased with increase in fermentation duration. The decrease was as a result of increase fermentation duration which might be attributed to more rapid mallard reaction between protein and carbohydrate which produced cookies with darker colour which may not be appealing to the consumer. Cookies sample produced from native acha flour was more preferred in terms of colour. The textural properties of the samples decreased with increased fermentation duration. This was in line with the report of Luyten et al . (2004). The panelists overall acceptability for the cookies samples decreased with increase in fermentation duration. Generally, cookies produced from native acha flour were rated most acceptable while fermented acha flour for 72 h was rated the least (Figure 5). The result obtained for sensory attributes in this study was similar with the work of Okpala et al. (2013) for cookies samples made from blends of germinated pigeon pea, fermented sorghum, and cocoyam flour.
CONCLUSIONS
This study provides valuable insights into the impact of fermentation duration on the functional, pasting, nutritional composition, phenolic, flavonoid microstructure, and sensory attribute of acha flour and cookies. The results indicate that there were significant differences (p<0.05) in the functional and pasting properties of the flour and cookies samples. An increase in fermentation duration was observed to enhance the oil absorption capacity and dispersibility of both flour and cookies samples. Overall, the values obtained for the pasting and functional properties of the flour samples fall within the recommended range for commercial flour. Regarding the proximate composition, as the fermentation duration increased, the protein and ash content of theacha flour samples increased. The mineral elements such as calcium, magnesium, sodium, zinc, copper, and phosphorus initially increased with fermentation up to 24 hours. Cookies with improved nutritional composition and acceptable sensory attributes could be produced from both native and acha flour fermented for durations between 24 and 72 h. This suggests that fermented acha flour holds promise for various food applications, including preparations, complementary foods, and gel formulations with reduced syneresis.