Effect of grain fermentation and malting time on nutrient and anti nutrient composition of biscuits from Aksum finger millet (Eleusine coracana)

Abstract

Finger millet grain is healthy and high in protein, fat and minerals especially calcium after proper processing. However, antinutrients represent a major obstacle to the usage of finger millet grains, which have restricted dietary applications. Biscuits are typically made from wheat flour, which increases wheat demand despite restricted supplies. The study aimed to explore how the length of fermentation and grain malting affected the color, nutritional content and antinutrients value of Aksum finger millet biscuits. Fermentation times were 24, 36 and 48 hours while the malting times were 24, 48 and 72 hours. The raw grain flour biscuit was used as a control sample to compare with each six samples of both factors. The moisture content of biscuits ranged from (5.64–8.00%), crude protein (6.60–10.09%), crude fat (12.12–17.30%), crude fiber (3.20–3.95%), total ash (1.55–2.19%), carbohydrate (70.81–75.50%) and energy content (439.50–479.51 Kcal/100 g) showing significant (P < 0.05) differences due to fermentation and malting duration. The results of zinc, iron and calcium were 1.80–2.36, 2.30–3.31 and 358.00–377.09 mg/100 g, respectively. The tannin and phytic acid content were significantly reduced from 1.21 to 0.78 and 435.50 to 430.78 mg/100g. The control biscuit had a good color value, but the biscuit samples exhibited significant (P < 0.05) reductions in their L*, a* and b* values due to fermentation and malting time. The result showed that grain fermentation and malting periods increased the proximate composition and mineral contents while decreasing the antinutrient content of the biscuits. In general, the acceptance of biscuits were enhanced by fermentation and malting.

Keywords:

• Aksum finger millet
• biscuits
• fermentation and malting times
• nutrient composition

Reviewing Editor:

• M. Luisa Escudero-Gilete, Universidad de Sevilla, SPAIN

subjects:

• Science
• Food Science & Technology
• Food Additives & Ingredients
• Food Chemistry
• Food Engineering

1. Introduction

Cereals have contributed to human population growth since ancient times and play an important role in the daily diet of billions of people around the world (Deshpande et al., 2015; Chinenye et al., 2017). Small millets, and a grain such as finger millet, are better able to grow in unsuitable lands with small inputs, so they are an important food source for low-income farmers in the tropics (Opole et al., 2019).

Finger millet (Eleusine coracana) is a tropical and subtropical grain that can grow in drought and high-temperature environments where corn, rice, wheat and other grains cannot grow (Rathore et al., 2019). Compared to rice, finger millet is richer in protein, lipids and minerals such as calcium and iron. It is also acid-free, easily digestible and one of the most digestible and least allergenic grains (Ambre et al., 2020). The existence of antinutrients in raw finger millet constitutes a significant limitation hindering the utilization of finger millet, implying that oxalic acid and phytic acid hinder their ability to provide nutrients. Tannins inhibit the digestion of proteins and calories. In finger millet, phytates interfere with the absorption of minerals, especially calcium, iron and zinc; Oxalate interferes with calcium and magnesium metabolism and combines with proteins to create centers that block gastric digestion (Abioye et al., 2018).

Simple processing steps commonly used on millets, such as shelling, sprouting and fermentation, have a significant impact on millet starch digestibility and protein digestibility in vitro (Verma & Patel, 2013). A combination of steeping, germination, drying, grinding and sieving is called malting, and its benefits include improved nutritional quality, increased starch digestibility, better sensory qualities and decreased antinutritional activities (Swami et al., Swami et al., 2013; Ramashia et al., 2019). As compared to malted sorghum and maize, finger millet grains are thought to be of the highest quality, hence malting is a widespread technology utilized in Africa (Ramashia et al., 2019). Producing enzymes and dissolving the cell walls around starch granules is the primary goal of malting (Swami et al., 2013). Natural fermentation is a process through which microorganisms like bacteria and yeast aid in the extraction of energy from carbohydrates in the absence of oxygen. Due to its decreased antinutrients content, finger millet is widely used as one of the main ingredients in many fermented food products, which not only enhances the dish’s flavor but also adds fiber, calcium and protein (Ambre et al., 2020). The antinutrient content of cereal grains can be significantly reduced by using traditional food processing techniques such as soaking and malting/sprouting, which also increase the bioavailability of nutrients (Hejazi & Orsat, 2016).

Several authors have researched the nutritional properties and health benefits of finger millet seeds, but it has been discovered that only traditional consumers eat it as food which limits the extent of utilization of the crop, especially in the East African region (Adebiyi et al., 2016; Owheruo et al., 2023). Studying the potential of finger millet is necessary considering on its nutritional and health benefits, and the rising demand for gluten-free food to overcome the grain’s decline (Adebiyi et al., 2016). This can be done by appropriately modifying the grain through malting and fermentation, increasing its uses by creating new products far beyond its normal range of uses (Akpoghelie et al., 2022). Value-added products suitable for rural and urban consumers can now be processed and prepared with advances in processing and value-added technology. In addition to having a higher fiber and micronutrient content than other cereals, finger millet also has an opportunity to enter the baking industry and produce several higher value-added products (Ambre et al., 2020). The use of scientific processing techniques can overcome current limitations, including the presence of antinutritional components and low sensory acceptability of conventionally produced millet products (Kumar et al., 2018). It has been demonstrated that cereal goods baked with various gluten-free cereals aside from oats have a slower rate of staling, a lower volume and a worse physical texture than those containing wheat (Hosseini et al., 2018).

Today, snacks such as cookies and crackers have become an essential part of the diet of people in cities through the country. Biscuits are usually made from wheat flour; imports require foreign currency due to limited supply on the domestic market. Indigenous grains can be used to completely or partially replace wheat in bread products (Friday et al., 2017). Biscuits are one of the instant snacks that have many attractive characteristics, such as a wider consumer base, relatively high stability, more convenience and better taste quality (Owheruo et al., 2023). According to the literature on gluten-free biscuits, finger millet is a healthy option in the commercial product range for people with celiac disease and diabetes (Singh & Kumar, 2018).

The Aksum finger millet variety (ACC #229355) is one of the varieties released by Melkasa Agricultural Research Center in 2016 G.C. approved for Shalla, Arsi Negelle, Assosa, Bako, Adet, Pawe and Axum adaptation zones. Therefore, this study is necessary to study the influence of processing conditions (malting and fermentation time) on the ability to provide nutrients in biscuits prepared from the Aksum finger millet variety which is now commercially grown in Ethiopia. The objective of this study was to study the effect of fermentation and malting time on the nutrient and antinutrient content of Aksum finger millet biscuits. Generally, the current focus has been on encouraging the commercial use of finger millet grains in food formulations and attaining a higher food security feature, with increased nutraceutical value and improved stress tolerance.

2. Materials and methods

2.1. Experimental materials

The raw material, 15 kg Aksum (ACC #229355) finger millet (Eleusine Coracana), was collected from Fedis Agricultural Research Center located in Harar, Ethiopia. The samples were placed in sacks and stored in a dry and cool place at Haramaya University Food Technology and Process Engineering Department laboratory. Sodium hypochlorite solution for surface sterilization and distilled water was prepared for sample preparation.

2.2. Experimental design

The experiment was conducted with a completely randomized design. It was planned with two independent factors: fermentation time (F) of the seed with three levels (24, 36 and 48 hours) and malting time (M) of the seed with three levels (24, 48 and 72 hours) and raw grain milled finger millet as a control (Adebiyi et al., 2017; Udeh et al., 2018). The treatments were done in triplicates and analyzed for all response variables.

2.3. Aksum finger millet raw grain flour preparation

Damaged grains, stones and other foreign objects were removed, and the normal grains were sorted and cleaned. Samples of 500 g were dried in a hot air oven drier for 8 hours at 50 $°\mathrm{C}$ to reduce the moisture content to roughly 11% (Akinsola et al., 2018). To produce raw flour, the dried samples were winnowed, ground in a laboratory mill, and then sieved at a 710 µm size. The flour was then placed in a plastic bag and kept in a dry, cool location until they were used (Adebiyi et al., 2018).

2.4. Malting of Aksum finger millet

The finger millet grains were initially surface sterilized for 30 minutes in a one percent sodium hypochlorite solution using a portion of 500 g. The finger millet grains were soaked in water for 24 hours at 25$°\mathrm{C}$ in a growth chamber after being rinsed five times with distilled water (1:3 w/v). The grains were placed on a nylon fabric and wrapped to keep them moist at room temperature. Water was then sprinkled on the grains occasionally every 24 hours. The sprouted grains were dried to increase the milling efficiency for 8 hours at 50$°\mathrm{C}$ after the required malting period was reached. The dried grains were then milled into fine light brown flour in a laboratory mill machine (3303 Perten Instrument, Finland), sieved through 710 µm, and stored in polyethylene bags until analysis (Udeh et al., 2018; Adebiyi et al., 2016; Abioye et al., 2018).

2.5. Fermentation of Aksum finger millet

The naturally occurring microbes such as yeast and bacteria on the grain surface were used to ferment the grains (Griffith et al., 1998). The grains of 500 g were cleaned by manual blowing, sorted, and separately steeped in deionized water at a grain-to-water ratio of 1:4 (w/v) and allowed to ferment at ambient temperature in a previously sterile beaker at 28 °C for 24, 36, and 48 hours (Adebiyi et al., 2017). Measurement of pH by digital pH meter from 24–48 hours was carried out (5.2–3.6) to control fermentation conditions. Yeast grows and multiplies much more rapidly at a pH higher than 5.0 (5.5 is optimal), but fermentation is more efficient if the pH reading is below 5.0. After each fermentation time, the foam water was drained, and the grains were dried for 8 hours at 50 $°C$ in an air oven drier, milled by a laboratory mill (3303 Perten Instrument, Finland) to get fine light brown flour followed by passing through a sieve of 710 µm (Onweluzo & Nwabugwu, 2009). The samples were stored in polyethylene bags until used for analysis.

2.6. Biscuit preparation

The baking process was done following the methods and formula described by Serrem et al. (2011), Shebabaw (2013) and Pedersen et al. (2004). The baking ingredients formulation were flour 100 g, sugar 25 g, shortening 29 g, baking powder 0.70 g, water 60 g, skimmed milk powder 0.30 g and salt 0.40 g.

All ingredients except flour were gently mixed in a planetary mixer to make homogenized cream (Table 1). The creamy mixture was manually poured into another mixer with flour, to make dough. The flours of fermented, malted grain and raw were separately prepared for biscuits. After thorough mixing the dough was molded manually and cut into circular shapes. The biscuits were baked in a previously heated oven at 210 $°\mathrm{C}$ for 18 minutes with little modification. The biscuits were allowed to cool for about 15 minutes on a rack before further analysis. Finally, raw grain biscuits, fermented grain biscuits at 24, 36 and 48 hours, and malted grain biscuits at 24, 48 and 72 hours were obtained. Totally seven formulations of biscuits were produced as revealed in Figures 1a–c.

2.7. Proximate analysis of biscuits

2.7.1. Crude protein

Based on the Kjeldahl method proposed by AACC (American Association of Cereal Chemists) (2000), official method 923.03, protein content was calculated. The crude protein content was estimated using the formula: $$\text{Crude\hspace{0.33em}Protein\hspace{0.33em}content\hspace{0.33em}}\left(g/100g\right)=\%N\times 6.27$$

Where 6.27 is the conversion factor and %N is the total percentage of nitrogen.

2.7.2. Crude fat

The crude fat of the finger millet biscuits was determined by the method described by AACC (American Association of Cereal Chemists) (2000), official method 923.03, using Soxhlet apparatus and extraction by petroleum ether.

2.7.3. Crude fiber

The crude fiber content of biscuit flour was determined according to the study of Gemede and Fekadu (2014).

2.7.4. Ash content

According to AACC (American Association of Cereal Chemists) (2000), official method 923.03, the total ash content of the biscuits was determined by the gravimetric method.

2.7.5. Carbohydrate

Utilizable carbohydrate (CHO) was calculated by subtracting some other contents from 100. The mathematical expression is as follows: $$\left(g/100g\right)\text{CHO}=100-\left(\begin{array}{l}\%\text{\hspace{0.33em}moisture}+\%\text{\hspace{0.33em}crudeprotein}\\ +\%\text{\hspace{0.33em}fiber}+\%\text{\hspace{0.33em}crudefat}+\%\text{\hspace{0.33em}ash}\end{array}\right)$$

2.7.6. Total energy

The mean values of crude protein, crude fat, and total carbohydrate were multiplied by the factors of 4, 9 and 4 correspondingly to produce the energy content (kcal/100g). Below is a formula for calculating energy. (1) $$\begin{array}{l}\text{Total energy in kcal}/100g\\ =\left(4\times \text{\%}P\text{rotein}+9\times \text{\%Fat}+4\times \text{\%CHO}\right)\end{array}$$(1)

2.8. Mineral analysis of biscuits

2.8.1. Iron

The UV–VIS spectrophotometer method of AACC (American Association of Cereal Chemists) (2000) was used to measure iron contents. The UV–VIS spectrophotometer read the absorbance of the sample, the standard and the blank at 510 nm. The following formula was used to compute the iron content: (2) $$\text{Iron Content}(\mathrm{\text{mg}}/100\mathrm{g})=\frac{C\times \mathrm{\text{DF}}\times 10}{\text{mass of sample}\left(\mathrm{\text{db}}\right)}$$(2) where C is the concentration of the sample in ppm,

DF is the dilution factor and 10 is a conversion factor since 10 mL were analyzed from 100 mL.

2.8.2. Calcium

Calcium was analyzed based on the AACC (American Association of Cereal Chemists) (2000) method. The atomic absorption spectrometer was used to measure the absorbance of the samples as well as the standard solutions at 422.7 nm. The concentration of samples was read from the plot of absorption against µg/mL of Ca. (3) $$\text{Calcium Content\hspace{0.33em}}\left(\text{mg}/100g\right)=\frac{\left(\mathit{\text{Cs}}-\mathit{\text{Cb}}\right)V\times D}{S}$$(3) where Cs and Cb are the analyte and blank concentrations in g/mL, respectively.

V = original volume (100 mL)

The ratio of the dilution volume V (mL) and the original aliquot volume (mL) used for dilution determines the dilution factor (D) if the original solution is diluted. S stands for sample mass in grams.

2.8.3. Zinc

Zinc was analyzed based on the AACC (American Association of Cereal Chemists) (2000) method. Both the standard solutions and the samples were analyzed using the atomic absorption spectrophotometer and absorbance was measured at 213.8 nm. The concentration of samples was read from the plot of absorbance against µg/mL Zn. (4) $$\text{Zinc content}(\mathrm{\text{mg}}/100\mathrm{g})=\frac{(\mathit{\text{Cs}}-\mathit{\text{Cb}})V\times D}{S}$$(4)

Where: Cs and Cb are concentrations in µg/mL of analyte and blank, respectively.

V is the original volume (100 mL) and D is the dilution factor = dilution volume (mL)/original aliquot volume (mL) used for dilution.

S is the sample mass in g.

2.9. Anti-nutritional factors analysis of biscuits

2.9.1 Tannin content

Using catechin as a tannin standard, the Gemede and Fekadu (2014) approach was used to determine the amount of tannin in the sample. A screw cup test tube containing 2 g of samples from each treatment in triplicates was weighed before being extracted for 24 hours at room temperature with mechanical shaking using 10 mL of 1% HCl in methanol. After 24 hours of shaking, the solution was centrifuged at 1000 RPM for 5 minutes. One milliliter of supernatant was combined with 5 mL of vanillin-HCl reagent (prepared by combining the equal volume of 8% concentrated HCl in methanol and 4% Vanillin in methanol).

For the condensed tannin determination, D-catechin was used as a standard. As a stock solution, 40 mg of D-catechin was weighed and dissolved in 1000 mL of 1% HCl in methanol. In test tubes, 0, 12, 24, 36, 48 and 60 mL of the stock solution were placed, and the volume of each test tube was adjusted to 1 mL with 1% HCl in methanol. Each test tube received 5 mL of vanillin-HCl reagent. The absorbance of the sample solutions and the standard solution was measured at 500 nm using a spectrophotometer after 20 minutes of incubation at 30$°C$. The absorbance of a blank sample was subtracted from the absorbance of the sample. The calibration curve was built using a series of standard solutions. A standard curve was created by plotting absorbance versus catechin (1 mg/mL) and the slope and intercept of the curve were used to calculate concentration. (5) $$\mathrm{\text{Tannin}}\left(\mathrm{\%}\right)=\frac{\mathrm{C}\times 10}{2000}\times 100$$(5)

Where: Concentration corresponding to the optical density.

2000 Volume of the extract (mL).

2000 = Sample weight (mg).

2.9.2. Phytic acid content determination

The phytic acid concentration of Aksum finger millet biscuit flour was determined using the method described by Wheeler and Ferrel (1971). About 0.25 g of grind flour sample was extracted with 12.5 mL of 3% trichloro acetic acid for 45 min in a water bath (GLS 400 water bath, 0 England) with vortex mixing (REAX top, Germany) at an ambient temperature (23$°C$) and centrifuged (4000 rpm/10 minutes) (Centurion Scientific Model 1020 DE, UK). The supernatant was used to calculate phytate levels. The sample solution was centrifuged after 4 mL of FeCl₃ was added. After carefully decanting the clear supernatant, the precipitate was washed with 20 mL of 3% TCA, 0.2 M of HCl and 20 mL of distilled water. Concentrated H₂SO₄ and H₂O₂ were used to digest the precipitate (30%). Phosphorus is converted into phosphate during digestion. The phosphate generated was analyzed by measuring the absorbance of phosphomolybdate blue generated on the addition of ammonium molybdate [(NH4)₆Mo₇O₂₄.4H₂O] (Morrison, 1964). Stock P solutions were prepared by taking 0.1 mg KH2PO4 into 250 mL H2O. For the calibration curve, a series of solutions (0, 0.2, 0.4, 0.6, 0.8, 1.0 and 1.2 ppm) were prepared from the stock solution. A UV/vis spectrophotometer was used to measure absorbance at 822 nm (Model 6505, U.K, and GENWAY). The sample absorbance was subtracted from the blank absorbance, and the phosphorus level was estimated using the calibration curve. Then phytate was estimated from phytate phosphorus (i.e., phytate = P × 3.55) (6) $$\mathrm{\text{Phytate}} \left(\frac{\mathrm{\text{mg}}}{100\mathrm{g}}\right)=\frac{\mathrm{C}\times 3.55}{\mathrm{S}}$$(6)

Where C = concentration (µg/mL) of phosphorus,

S = sample weight (db.).

2.8. Data analysis

One-way analysis of variance (ANOVA) was used to examine the data, and it was done using Statistical Analysis System version 9.0, SAS computer software (TS Level 00 M 0 XP-PRO platforms). A significant test was accepted with a probability value of P < 0.05. The Fisher’s least significance difference test was used to compare means. Results were provided in mean and standard deviation (mean ± standard deviation).

3. Results and discussions

3.1. Effects of fermentation and malting times on proximate composition of biscuits

The proximate composition of control, fermented, and malted Aksum finger millet-based biscuits is given in Table 2.

Table 1. Baking ingredients formulation.

Ingredients	Quantity (g)
Flour	100
Sugar	25
Shortening	29
Vanilla essence	6
Baking powder	0.70
Water	60
Skimmed milk powder	0.30
Salt	0.40

Table 2. Effects of fermentation and malting times on proximate composition of Aksum finger millet biscuits.

Factors		Proximate composition (g/100g)
FT	MC	Ash	Crude protein	Crude fat	Crude fiber	CHO	Total energy (kcal)
F1	6.60 ± 0.68^b	1.91 ± 0.10^b	8.81 ± 0.72^b	13.92 ± 1.25^b	3.62 ± 0.13^c	73.93 ± 0.79^b	456.32 ± 10.7^c
F2	6.51 ± 0.35^b	1.55 ± 0.06^c	9.57 ± 0.68^a	17.20 ± 0.58^a	3.89 ± 0.04^b	70.81 ± 0.50^c	476.45 ± 6.43^b
F3	5.64 ± 0.78^c	1.57 ± 0.23^c	9.40 ± 0.72^a	17.30 ± 0.60^a	3.95 ± 0.08^a	71.52 ± 0.80^c	479.51 ± 8.12^a
RFB	8.00 ± 0.50^a	2.19 ± 0.19^a	6.60 ± 0.67^c	12.12 ± 0.35^c	3.20 ± 0.16^d	75.5 ± 0.50^a	439.5 ± 0.50^d
MT
M1	6.73 ± 0.73^b	1.74 ± 0.18^b	8.58 ± 0.38^c	15.41 ± 1.93^c	3.74 ± 0.20^c	72.35 ± 1.63^b	462.51 ± 12.98^c
M2	6.38 ± 0.63^b	1.70 ± 0.15^b	9.12 ± 0.52^b	16.20 ± 1.77^b	3.81 ± 0.14^b	71.89 ± 1.63^b	469.92 ± 11.5^b
M3	5.65 ± 0.47^c	1.58 ± 0.29^c	10.09 ± 0.34^a	16.81 ± 1.61^a	3.91 ± 0.13^a	72.03 ± 1.44^b	479.85 ± 10.39^a
RFB	8.00 ± 0.50^a	2.19 ± 0.19^a	6.60 ± 0.67^d	12.12 ± 0.35^d	3.20 ± 0.16^d	75.5 ± 0.50^a	439.5 ± 0.50^d

Note: All values are means of triplicates ± standard deviation. FT = fermentation time (F1, F2 and F3 = 24, 36 and 48 hours, respectively. MT = malting time (M1, M2, and M3 = 24, 48 and 72 hours respectively. RF = raw flour biscuit, MC = moisture Content, CHO = carbohydrate.

3.1.1. Moisture content

Finger millet biscuits showed a significant decrease (P < 0.05) in moisture content with values ranging from 6.60% to 5.64%. According to Table 2, the raw grain flour biscuit exhibited a value of 8.00%. The primary goal of malting is to create enzymes and break down the cell walls around starch granules (Swami et al., 2013). Throughout the germination process, amylase enzymes break down starch into low molecular weight carbohydrates (oligo and disaccharides). The product that results from germinated grain has low water holding capacity and high energy after baking. The product can attain the desired crispness and extended shelf life due to the product’s low value. The level of moisture in food items indicates how prone they are to being invaded by microorganisms, which eventually leads to their decay. The moisture level is crucial in determining how long the final product can be stored (Sani et al., 2020). Since the standard moisture level for biscuits from wheat flour was 2–5% this biscuit is nearly in the acceptable range.

The moisture content of biscuits varied significantly (P < 0.05) depending on the duration of grain malting. When the malting time increased from 24 to 72 hours, the values declined from 6.73 to 5.65%. Adebiyi et al. (2016) obtained comparable findings, indicating that the moisture content of biscuits made from raw pearl millet (5.77 g/100g) was slightly greater than that of samples derived from malted and fermented grains (5.09%). In general, the levels of moisture recorded in this research were significantly lower than 10.0 g/100g, suggesting that they could offer a favorable storage benefit.

3.1.2. Ash content

The quantity of inorganic minerals found in a specific food is connected to its ash content. The lengthier the fermentation process, the total amount of ash content shows a considerable decrease (P < 0.05), as illustrated in Table 2. The ash content values of biscuits made from fermented grains for 24–48 hours varied between 1.91 and 1.57 g/100g. The biscuit sample made from raw grain flour had a higher value of 2.19/100 grams. Results are similar to Mudau et al. (2022) findings, which showed that the ash content of light brown and dark brown finger millet biscuits decreased from 1.66 to 0.93 and 1.52 to 1.17 g/100g respectively, as the duration of spontaneous fermentation increased from 0 to 72 hours. The probable reason for the reduction in ash content in biscuits is either the release of soluble mineral elements into the fermenting medium (acid liquid) or the activity of fermenting microorganisms (Mudau et al., 2022).

Table 2 reveals that extending the duration of malting from 24 to 72 hours resulted in a significant reduction (P < 0.05) in ash content, with values ranging from 1.74 to 1.58 g/100g. In the study of Kumar et al. (2021), it was reported that the ash content of finger millet decreased from 2.27 g/100g to 1.24 g/100g in nongerminated samples and samples that underwent a germination period of 96 hours, respectively. Similar decreases were also reported by Adebiyi et al. (2016) due to the leaching of soluble inorganic salts during the fermentation and malting processes.

3.1.3. Crude protein

The amount of crude protein in biscuits made from fermented finger millet flour samples showed a significant increase (P < 0.05) as the fermentation time increased from 24 to 48 hours. The crude protein content ranged between 8.81 and 9.57 g/100g during this period (Table 2). Biscuits from fermented grain for 48 and 72 hours had the highest protein content, while biscuits made from raw grain flour had the lowest crude protein content of 6.60 g/100g. To put it differently, the biscuits experienced a notable rise in their crude protein content as a result of fermenting the grains. Mudau et al. (2022) have also noted a comparable pattern of protein increase following fermentation. Nkhata et al. (2018) also reported a significant rise in protein content of pearl millet grains, amounting to 15.32%, after undergoing 16 hours of fermentation. According to Akinola et al. (2017), the increase in protein content is probably caused by the conversion of insoluble proteins into soluble proteins during fermentation.

Similarly, the duration of the malting process had a significant impact (P < 0.05) on the protein composition of the biscuits. According to the data, there was an increase in protein content as the malting times progressed over 24 to 72 hours, with corresponding values of 8.58, 9.12 and 10.09 g/100g, respectively. The findings of Bolarinwa et al. (2016) align with this study, as they also observed an increase in the protein content of malted-sorghum biscuit samples, ranging from 7.28 to 11.74 g/100g. The greater crude protein values of biscuits from fermented and malted samples were caused by the accumulation of proteins and the creation of some extra amino acids with an increase in fermentation and malting duration (Adebiyi et al., 2017). As a result, Aksum finger millets show great potential as a nutritious grain for creating value-enhanced food products that provide an adequate amount of protein when combined equally with fermented and malted flour. Consuming millet biscuits regularly can potentially raise protein consumption for both children and adults.

3.1.4. Crude fat

Table 2 displays the average fat content values of the biscuits, which significantly increased (P < 0.05) as the fermentation and malting period of the grains increased. The biscuits from grains that underwent fermentation for 36 and 48 hours had the highest fat content, with values of 17.20 and 17.30 g/100g respectively. The bioavailability of nutrient composition can be enhanced through malting and fermentation, which are efficient ways to eliminate antinutritional effects (Igbabul et al., 2014). On the other hand, the biscuits made from raw flour (control) had the lowest fat content, with a value of 12.12 g/100g. Mudau et al. (2022) also noted an increase in the fat content of fermented finger millet biscuits, with reported values ranging from 18.83 to 22.11 g/100g for light brown biscuits and 16.69–20.75 g/100g for dark brown biscuits. Enhancing food appearance, flavors, aroma, proteins, vitamins, vital amino acids, and reduction of anti-nutrients can all be achieved naturally by fermentation (Sharma et al., 2020).

Malting of finger millet grain had a significant difference (P < 0.05) in the fat content of biscuits. The fat content increased with values ranging from 15.41 to 16.81 g/100g when the duration of malting increased from 24 to 72 hours. The biscuits in this study contained a high amount of fat due to the use of vegetable oil and milk powder in their preparation and depolymerization during baking. However, the fat content increased significantly (P < 0.05) when the fermentation and malting times of Aksum finger millet grain were extended. Fat plays a crucial role in enhancing both the texture and overall quality of the final product.

3.1.5. Crude fiber

The fiber content of the biscuits significantly increased as the fermentation time increased, and the biscuits made from fermented samples at 48 hours had the highest average value (3.95 g/100g). On the other hand, the lowest average value (3.20 g/100g) was found in the biscuits made from raw Aksum finger millet flour (control). As the duration of fermentation for finger millet biscuits increased from 24 hours to 48 hours, the values of the biscuits increased ranging from 3.62 to 3.95 g/100g. According to research conducted by Adebiyi et al. (2016), it was found that the carbohydrate, fiber, and energy content of fermented and malted biscuit samples were considerably greater (P < 0.05) than that of biscuits made from regular grain flours. Finger millet sets itself apart from other millet, such as foxtail millet, pearl millet, kodo millet, and proso millet, because it possesses a unique five-layered testa. According to Chandra et al. (2016), this could potentially be among the factors responsible for the increased amount of dietary fiber in finger millet biscuits. The complex protein is hydrolyzed into simpler proteins throughout the fermentation process, and the phytates and tannins are also reduced. This improves the bioavailability of major and minor nutrients as well as protein and decreases the amount of antinutrients, all of which aid in a larger absorption of essential nutrients (Gowthamraj et al., 2020).

The amount of fiber in the biscuits differed significantly (P < 0.05) as a result of the duration of malting. The fiber content ranged from 3.74 to 3.91 g/100g as the fermentation time increased. During germination, structural components like as hemicelluloses and cellulose are synthesized, whereas starch is broken down (Obadina et al., 2017). Tamilselvan and Kushwaha (2020) suggested that fiber content is increased during germination due to changes in polysaccharides present in the cell wall and increased cellular structure in plants while sprouting. The fiber levels found in the malted sorghum and soybean biscuit samples, as reported by Feyera (2020), were similar to those mentioned here, ranging from 2.56 to 3.46 g/100g whereas the fiber content of wheat biscuit was 3.54 g/100g.

3.1.6. Carbohydrate content

As the fermentation time of the grains increased, there was a significant reduction (P < 0.05) in the amount of carbohydrates present in the biscuits. The values obtained for biscuits made from grains that were fermented for 24, 36 and 48 hours were 73.93, 70.81 and 71.52 g/100g, respectively. Even though the carbohydrate content shows a significant increase at 48 hours than 36 hours duration, the reason for the decline can be attributed to higher levels of crude protein, crude fat, and crude fiber content, since carbohydrate was calculated by subtracting some other contents from 100 which are reflected in the increased levels of both factors as shown in Table 2.

There was no significant (P > 0.05) variation in the amount of carbohydrates found in the biscuits, regardless of the duration of malting. Nevertheless, the values of all the samples were significantly lower than those derived from raw grain flours, with a significant difference (P < 0.05). The raw grain flour biscuits achieved the highest recorded value of 75.5 grams per 100 grams. The findings are similar to a report by Adebiyi et al. (2017), which demonstrated that the carbohydrate content of fermented (67.82 g/100g) and malted (65.19 g/100g) pearl millet biscuit samples was significantly higher (P < 0.05) than that of the raw samples (64.48 g/100g). The explanation might be that during germination, there is partial breakdown of amylopectin and an increase in amylose content, which contributes to the total carbohydrate of malted finger millet (Adebiyi et al., 2017).

3.1.7. Total energy

The significant increase in the fat content of fermented sample biscuits, as shown in Table 2, may have played a role in the elevated energy values, which ranged from 456.32 to 479.51 kcal per 100 grams. The raw biscuits had the lowest recorded result (439.5 g/100g). According to Okache et al. (2020), the reason for the high energy levels can be ascribed to the conversion of carbohydrates (specifically starch) into simpler sugars, thus increasing calories. There was a significant variation (P < 0.05) in energy values as the fermentation time increased, as it is derived from the combined quantities of protein, fat, and carbohydrate (4 times crude protein + 4 times carbohydrate + 9 times fat). As the fermentation and malting period extended, the higher presence of fat became a contributing factor to the increase in values (Forsido et al., 2020). This is because an increase in processing time led to an increase in both fat and protein, while simultaneously causing a decrease in carbohydrates (Adebiyi et al., 2016).

The energy content of biscuits varied significantly (with a statistical significance level of P < 0.05) depending on the duration of malting. The malting times between 24 to 72 hours resulted in calorie content ranging from 462.51 to 479.5 Kcal per 100 grams. The findings of the research indicated that the energy values obtained from samples of weaning food, which fell between 381.25–435.72 Kcal/100g, were lower than the results of the study conducted by Sani et al. (2020). Results are in line with the report of Adebiyi et al. (2017) who observed that the energy content of raw, fermented, and malted pearl millet biscuits was 447.84, 452.05 and 458.67 kcal/100g correspondingly.

3.2. Mineral content of Aksum finger millet biscuits

Table 3 presents the mineral content data of biscuit samples influenced by fermentation and malting duration. The mineral levels in the biscuit samples made from malted and fermented grain were generally much higher. The aforementioned observation can still be attributed to the Maillard reaction that takes place while the baking process is happening.

Table 3. Effects of fermentation and malting times on the mineral content of Aksum finger millet biscuits.

Factors	Mineral contents
Fermentation time	Zn (mg/100 g)	Fe (mg/100 g)	Ca (mg/100 g)
F1	1.82 ± 0.10^b	2.42 ± 0.46^b	368.57 ± 2.79^c
F2	1.81 ± 0.03^b	2.30 ± 0.36^d	374.98 ± 2.02^b
F3	2.36 ± 0.20^a	3.31 ± 0.34^a	377.09 ± 3.29^a
RFB	1.80 ± 0.01^b	2.34 ± 0.05^c	358.00 ± 2.64^d
Malting time
M1	1.87 ± 0.18^c	2.35 ± 0.70^c	370.72 ± 4.21^c
M2	2.02 ± 0.28^b	2.59 ± 0.42^b	373.38 ± 3.48^b
M3	2.09 ± 0.36^a	3.08 ± 0.41^a	376.53 ± 4.26^a
RFB	1.80 ± 0.01^d	2.34 ± 0.05c	358.00 ± 2.64^d

Values are means ± Standard deviation and values in the same column with different superscript letters are significantly different from each other (P ≤ 0.05). FT = fermentation time (F1, F2, and F3 = 24, 36 and 48 hours respectively. MT = Malting time (M1, M2 and M3 = 24, 48 and 72 hours respectively. RFB = Raw flour biscuit.

3.2.1 Zinc

There was a significant difference (P < 0.05) in the zinc content of biscuits (2.36 mg/100g) at the fermentation time of 48 hours as indicated in Table 3, but the zinc content of biscuits was not significantly (P > 0.05) affected by the fermentation time, as evidenced by the related values of 1.80, 1.82 and 1.81 mg/100g for biscuits made from raw, fermented grain for 24 and 36 hours, respectively. Nevertheless, the biscuits that were prepared using flour derived from grains that underwent a fermentation process lasting 48 hours exhibited the highest recorded zinc content of 2.36 mg/100g.

In contrast, the grains that underwent malting resulted in flours that produced biscuits with increased higher levels of zinc (P < 0.05). For each sample that underwent malting at 24, 48 and 72 hours, the zinc content measured 1.87, 2.02 and 2.09 mg/100g, respectively. The control sample contained 1.80 mg of zinc per 100 g. The results of zinc content found in these biscuits were similar to the zinc content (1.02–1.16 mg/100 g) found in biscuits made from malted sorghum observed by Bolarinwa et al. (2016). Zinc deficiency in young children can lead to stunted growth, diarrhea, and pneumonia. Hence, regular consumption of fermented finger millet biscuits can play a significant role in eliminating mineral deficiency, particularly in children, due to their enhanced mineral content.

3.2.2. Iron

The iron content of the biscuits was significantly affected (P < 0.05) by increasing the fermentation time. According to Table 3, the biscuits had the highest value of 3.31 mg/100g after 48 hours of fermentation, while the lowest value (2.30 mg/100g) was observed after 36 hours of fermentation. Adebiyi et al. (2017) suggest that the Maillard reaction has the potential to create complexes or break down compounds, ultimately impacting the solubility and accessibility of minerals during baking.

As the malting period increased, there was a significant difference (with a statistical significance of P < 0.05) in the increase of iron content. The biscuit samples, produced from grain malting periods of 24, 48 and 72 hours, were found to have levels of 2.35, 2.59, and 3.08 mg/100g respectively. Rani et al. (2018) found that the level of phytic acid in pearl millet grain decreased, while the ability to extract minerals such as calcium, iron, zinc, phosphorus, iodine, copper and manganese significantly improved as the germination time progressed. The reason for this could be the presence of phytase, an enzyme produced during germination that breaks down phytic acid, a substance that binds minerals and makes them accessible to the body (Rani et al., 2018).

3.2.3. Calcium

The calcium content of the biscuits varied significantly (P < 0.05) depending on the fermentation period of the grain. The values of the biscuits made from fermented grains increased from 358.00 mg/100g in the control sample to 368.57, 374.98 and 377.09 mg/100g when the fermentation period was 24, 36 and 48 hours, respectively. Each of the values displayed significant differences from one another (P < 0.05). Mudau et al. (2022) also acquired comparable data which indicated that finger millet grains had calcium content (386.35, 398.39 and 411.33 mg/100g) after being fermented for 24, 48 and 72 hours, respectively.

The variations in the calcium content of biscuits were found to be significant (P < 0.05) based on the duration of grain malting. The values varied between 370.72 and 376.53 mg/100g as the malting periods increased from 24 to 72 hours. It can be inferred from the statistical data that the calcium availability in grain products was improved through the malting process, as all the measured values were higher than the control. Fermentation and malting improve the bioavailability of calcium, phosphorous, and iron, most likely due to the breakdown of oxalates and phytates that are complex with minerals and hence reduce their bioavailability (Nkhata et al., 2018). Osteoporosis, osteomalacia, and rickets are known to be caused by a deficiency of calcium in the body, particularly in Africa and Asia (Awuchi et al., 2020).

3.3. Effects of fermentation and malting times on the ant-nutritional factors of Aksum finger millet biscuits

3.3.1. Tannin content

The biscuits showed significant variations in condensed tannin content caused by fermentation time, which was significant (P < 0.05) different (Table 4). The biscuits made from raw Aksum finger millet flour had the highest value of 1.21 mg per 100 g, while the biscuits made from flour where grains were fermented for 48 hours had the lowest value of 0.78 mg per 100 g. The tannin content of biscuits decreased from 1.04 to 0.78 mg/100g as the fermentation period increased from 24 to 48 hours.

Table 4. Effects of fermentation and malting times on the antinutritional factors of Aksum finger millet biscuits.

Factors	Anti-nutritional factors
Fermentation time	Phytic acid (mg/100g)	Tannin (mg/100g)
F1	434.03 ± 0.36^b	1.04 ± 0.07^b
F2	431.54 ± 0.89^c	0.97 ± 0.07^c
F3	430.78 ± 0.61^d	0.78 ± 0.08^d
RFB	435.50 ± 0.50^a	1.21 ± 0.14^a
Malting time
M1	432.74 ± 1.34^b	0.99 ± 0.10^b
M2	432.00 ± 1.63^c	0.96 ± 0.12^c
M3	431.60 ± 1.59^c	0.85 ± 0.14^d
RFB	435.50 ± 0.50^a	1.21 ± 0.14^a

Values are means ± standard deviation and values in the same column with different superscript. Letters are significantly different from each other (P ≤ 0.05). FT = fermentation time (F1, F2 and F3 = 24, 36 and 48 hours, respectively. MT = malting time (M1, M2 and M3 = 24, 48 and 72 hours respectively. RFB = raw flour biscuit.

Likewise, the duration of the malting process had a significant effect (P < 0.05) on the tannin levels found in the biscuits. The longer the grains were malted, the lower their tannin content became. Specifically, the tannin content decreased by 0.99, 0.96 and 0.85 mg/100g for malted times of 24, 48 and 72 hours, respectively. In their study, Syeunda et al. (2021) conducted research on finger millet that had been malted for varying durations. They successfully demonstrated a favorable decrease in tannin content, which dropped from 0.82 to 0.53 mg/100g. According to Hejazi and Orsat (2016), the process of malting can greatly decrease the levels of anti-nutrients in cereal grains and enhance the absorption of nutrients in them.

3.3.2. Phytic acid

The phytic acid content was impacted by the fermentation time, as indicated by the significant effect observed (P < 0.05) in Table 4. The biscuits made from raw grain, which consisted of unfermented grain, had the highest value of 435.50 mg/100g. On the other hand, the biscuits made from grain that had been fermented for 48 hours recorded the lowest value of 430.78 mg/100g. When the fermentation time increased, the levels of phytic acid decreased. The fermentation process hydrolyzes the complex protein into simpler proteins while decreasing the phytates (Gowthamraj et al., 2020).

Similarly, the duration of malting exhibited a significant decrease (P < 0.05) in the phytic acid level found in the biscuits. As the malting duration increased to 24, 48 and 72 hours, the amount of phytic acid in biscuits was measured to be 432.74, 432 and 431.60 mg/100g, respectively. Phytase activity rises during the process of germination, leading to the breakdown of phytate. Sihag et al. (2015) stated that the increase in phytase activity during the germination of grains was identified as an additional factor contributing to the reduction of phytic acid levels. This increased activity is responsible for the hydrolysis of phytate, breaking it down into phosphate and myoinositol phosphates.

3.4. Effects of fermentation and malting times on the color of Aksum finger millet biscuits

The L*, a*, and b* values of the biscuit samples were significantly (P < 0.05) affected by fermentation time (Table 5). The L* values of the biscuits made from fermented grains ranged from 33.06–33.85 due to different times. The L* values of the control biscuit had the largest value (38.92) of all the data. The redness value of the control was 8.90 significantly the largest of all the other values. The L* value of the biscuits produced from flours fermented grains was 8.18, 8.44, and 7.29 for those with fermentation times of 24, 36, and 48 hours, respectively. The yellowness (b*) values were observed as 12.45, 12.64, and 12.29 for 24, 36 and 48 hours fermentation times which were lower than the value (15.91) of the control biscuit. Similar trends for pearl millet biscuits obtained L* values ranging from 74.347 to 55.351 (Kulthe et al., 2017). The comparable increasing trend in L*, a*, and b* values was also found by Mudau et al. (2022) for biscuits made from fermented finger millet flour. The species of the grains and the phytochemicals that leak during malting may be the cause of the elevated level of L*. Because of the times involved, the content of a* was lower during the malting and fermenting period. The presence of phenolic chemicals, such as tannin, at the grain’s pericarp and testa, which were lessened by leaching (during malting) and polyphenol oxidase during fermentation, maybe the cause of this color shift, particularly in a* (Olamiti et al., 2020).

Table 5. Effects of fermentation and malting times on the color of Aksum finger millet biscuits.

Factors	Color
Fermentation time	L*	a*	b*
F1	33.06 ± 1.66^c	8.18 ± 0.57^cb	12.45 ± 1.10^b
F2	34.06 ± 2.06^b	8.44 ± 0.57^b	12.64 ± 0.57^b
F3	33.85 ± 1.77^b	7.99 ± 0.44^c	12.29 ± 0.46^b
RFB	38.92 ± 0.01^a	8.90 ± 0.02^a	15.91 ± 0.01^a
Malting time
M1	34.54 ± 2.43^b	7.71 ± 0.28^d	11.95 ± 0.52^d
M2	33.46 ± 0.88^c	8.28 ± 0.56^c	12.44 ± 0.63^c
M3	32.97 ± 1.63^c	8.62 ± 0.34^b	12.99 ± 0.74^b
RFB	38.92 ± 0.01^a	8.90 ± 0.02^a	15.91 ± 0.01^a

Values are means ± standard deviation and values in the same column with different superscript. Letters are significantly different from each other (P ≤ 0.05). FT = fermentation time (F1, F2 and F3 = 24, 36 and 48 hours, respectively. MT = malting time (M1, M2 and M3 = 24, 48 and 72 hours, respectively. RFB = raw flour biscuit.

The L*, a* and b* values of the biscuit samples were significantly (P < 0.05) affected by malting time. The L* values of biscuits decreased to 34.54, 33.46 and 32.97 for samples with 24, 48 and 72 hours of malting times, as compared to the 38.92 of the control biscuits. Likewise, the redness (a*) and yellowness (b*) values were decreased as malting times increased, below the control biscuits.

4. Conclusions

The nutritional composition and mineral content of biscuits were enhanced through fermentation and malting techniques. The study results indicate that Aksum finger millet biscuits contain considerable quantities of carbohydrates, crude protein, overall energy, crude fiber, calcium, zinc and iron. In addition, the anti-nutritional substances were further diminished during the processing stage. The alterations improved the color attributes and capabilities of the biscuit, making it more suitable for use in various baked food items. In this investigation, it was discovered that fermentation and malting methods, compared to raw grain biscuits, demonstrated superior nutrient and mineral levels. Additionally, malting was found to have lower levels of antinutritional factors which were considered in this study. Potential gluten-free products can be made using fermented and malted Aksum finger millet biscuits as raw materials. The optimal period for fermentation was observed at 48 hours whereas 72 hours for the malting period showed favorable nutritional enhancements and a decrease in anti-nutritional components that impede the solubilization of nutrients.

Figure 1. (a–c) Photo of raw, fermented and malted grain biscuits, respectively.

Authors’ contributions

Ayele Assefa was responsible for the first draft, data collection, research, formal analysis, and drafting of the manuscript. Getachew Neme and Solomon Abera were responsible for project management, consulting, and draft editing. Girma Daba was responsible for project support, drafting and editing.

Acknowledgments

Haramaya University deserves a huge thank you for providing me with financial assistance. Great thanks to Fedis, Melkasa and the Holetta Agricultural Research Centers for their material and laboratory support and encouragement. The Ministry of Education is also highly acknowledged for the study sponsorship.

Disclosure statement

The authors state that they have no conflicts of interest.

Data availability statement

This publication incorporates all of the generated data that support the study’s conclusions.

Additional information

Notes on contributors

Ayele Assefa Adugna

Ayele Assefa has been a chief technical assistance and researcher at Haramaya University, Ethiopia, for the last five years. His field of specialization is in Food Engineering. His research interests are mostly in the fields of food biotechnology, food processing technology, the production of nutritional and fortified foods for improved life , health and the quality and safety of food. His ultimate focus is to use current technology and scientific research to introduce his people’s traditional food to the rest of the world in a modern style.

Getachew Neme Tolesa

Getachew Neme Tolesa (PHD) is an associate professor (researcher and lecturer) of Food Science and Postharvest Technology at Haramaya University with over 15 years of experience in teaching and research. He specializes in Food Science and Postharvest Technology. He mainly works and publishes in the areas of postharvest food preservation, postharvest handling, food value addition, food science, food processing, food engineering, food value chain, food safety and nutrition intervention developments research.

Solomon Abera

Solomon Abera (D.Eng) is an associate professor (researcher and lecturer) of Food Engineering at the Department of Food Technology and Process Engineering, Haramaya University, with over 30 years of experience in teaching, research, and development. He specializes in Food Engineering and mainly works and publishes in food engineering, postharvest food preservation, postharvest handling, food value addition, food science, food processing, food engineering, food value chain, food safety and nutrition intervention research and developments.

Girma Daba Deme

Girma Daba is a lecturer and researcher at the Department of Food Technology and Process Engineering, Haramaya University, with over 10 years of experience in teaching, research, and development. He specializes in Process Engineering and mainly works and publishes in food engineering, waste utilization, food processing and food engineering.