Introduction

Development of composite flour bakery products is the latest trend in bakery industry. The growing interest in these types of food products is due to their better nutritional properties. Exploitation of nutritious and medicinally-valued plant foods can play an important role in meeting consumer demands. Successful enrichment of staple cereal products with under-utilized foods like lotus rhizome can prove to be a sustainable strategy to meet the nutritional needs as well as bring healthiness to people.

Lotus (Nelumbo nucifera) is a rhizomatous aquatic, ornamental, edible and medicinal plant which is grown as a non-conventional vegetable commonly in Australia, China, India and Japan. Every part of the plant including flowers, seeds, leaves, stems and roots are consumed [1]. Tender rhizomes, stems and leaves of lotus are edible and can be cooked along with other vegetables, soaked in syrup or pickled in vinegar [2]. At present, most studies have been focused on physicochemical properties of starch from lotus rhizome [3–5], antioxidative capacity [6] and health benefits [7] of lotus rhizome. However, scientific information on the utilization of lotus rhizome in food products, particularly in bakery products is rare.

Bread sticks/soup sticks are also called as dipping sticks and are usually pencil-sized sticks, which are of crisp texture, dry bread which originated in Turin and surrounding area in Italy. They are offered as appetizer. Utilization of brewers spent grain as a functional ingredient in soup stick was reported, wherein incorporation of 15% brewers spent grain increased the dietary fiber content and had a shelf-life of 50 days [8]. The objective of the present study is to evaluate the potential of successful utilization of lotus (Nelumbo nucifera) rhizome powder in bread sticks and to analyze the nutritional composition. Rheological properties of wheat flour (WF) as influenced by the addition of 0–20% of lotus rhizome powder (LRP) were also studied.

Materials and methods

Commercial WF was procured from the local market. Other ingredients such as compressed yeast (Tower brand, AB Mauri, Chennai, India), hydrogenated fat (Bunge India Pvt Ltd., Mumbai, India), salt (common food-grade sodium chloride) and sugar from local market were also procured.

Processing of lotus rhizome

Fresh lotus rhizomes were procured from the local market of Manipur, cleaned, washed and sliced to 2–3 mm thickness. About 500 g of sliced rhizomes were placed in a stainless steel tray (40 × 80 cm) and kept for drying in an air circulated tray dryer (Sakav Drier, Shirsat Electronics, Mumbai, India) at 40 °C for 5–6 h. The dried rhizome was pulverized in a hammer mill (Falling Number, 3100, Australian Wheat Board) to obtain LRP of 140 µm particle size. The yield of LRP was about 6–7%.

Chemical characteristics of WF and LRP

The chemical characteristics of WF and LRP such as moisture (method 44-16), protein (method 46-10), crude fat (method 30-10), ash (method 08-01), protein (method 46-10), dry gluten (method 38-10), falling number (method 56-81B) and Zeleny’s sedimentation value (method 56-61A) were determined using standard methods of AACC (2000) [9]. Dietary fiber was determined using standard method of AOAC (2005) [10].

Preparation of extracts

WF, LRP and the bread sticks were homogenized with a food processor (Sumeet Mixie, Chennai, India) and dried at 45 °C in a hot air oven (Servell Instruments Pvt Ltd, Bengaluru, India) for 3 h. To extract the free compounds, 10 ml of HCl-acidified methanol (1:1000) was added to 5 g of WF/LRP/bread sticks separately, mixed and subjected to shaking at 40 °C for 1 h and filtered using Whatman filter number 40. The filtrate was further subjected to centrifugation (5000 rpm, 10 min) at ambient temperature. The supernatant was collected and stored in dark at −20 °C until further use for determination of total phenolic content (TPC), total flavonoid content (TFC) and DPPH scavenging activity. Extraction was done in triplicate for flours and bread stick samples.

DPPH free radical scavenging activity (RSA)

The radical scavenging activity was determined according to Brand-Williams et al. [11]. Sixty microlitres (60 μl) of the extract was taken in test tubes. To this 40 μl of methanol and 5 ml of 0.1 mM solution of DPPH were added. The test tubes were shaken and kept in dark at 27 °C for 20 min. The absorbance was read and the percentage free DPPH radical scavenging activity was calculated from the following equation:

$$\begin{aligned}{{\text{RSA}} } ( \% ) & ={{\text{Absorbance }} }( {\text{DPPH}} ) \\ & \quad -{{\text{Absorbance }} }( {\text{sample}} )/{ {\text{Absorbance }} }( {\text{DPPH}} ) \times 100 \end{aligned}$$

Total phenolic content (TPC)

The TPC was determined according to Kahkonen et al. [12] with slight modifications. The method was based on the coloured reaction of phenolics with Folin–Ciocalteu reagent. Upon the reaction with phenols, Folin–Ciocalteu reagent is reduced to a blue coloured oxide. The intensity of the resulting colour was measured with UV–VIS-1800 spectrophotometer (Shimadzu, Japan) at 765 nm. One milliliter of FC reagent (10%) was added to 0.2 ml of sample solution. After 2 min, 0.8 ml 7.5% Na2CO3 solution was added and made upto a final volume of 5.0 ml before 1 h incubation in darkness at room temperature. The absorbance was read against blank at 765 nm. A standard stock solution was prepared by dissolving 5.0 mg of gallic acid in 5.0 ml of ethanol and diluting to 50 ml with water. The TPC was expressed as gallic acid equivalents (mg GAE/g dry weight). All analyses were performed in duplicate.

Total flavonoid content (TFC)

Flavonoid contents of the extracts were assayed using the aluminum chloride colorimetric method of Chang et al. [13]. For total flavonoid determination, quercetin was used as the standard. Stock quercetin solution was prepared by dissolving 2.0 mg quercetin in 5.0 ml methanol. The appropriate dilution of extracts (0.2 ml) were mixed with 1.5 ml of 95% ethanol, followed by 0.1 ml of 10% aluminum chloride, 0.1 ml of 1 M potassium acetate and made upto a final volume of 2.5 ml with methanol. After mixing, the solution was incubated for 60 min at room temperature. The absorbance of the mixture was measured against a blank at 420 nm with a spectrophotometer. The concentration of TFC in the test samples was expressed as µg quercetin equivalent (QE)/g of the dry weight.

Rheological properties

Blends were prepared by replacing WF with LRP at 100:0, 90:10, 85:15 and 80:20 and effect of LRP on the rheological properties of WF dough was studied using farinograph (method 54-21) and amylograph (method 22-10) according to standard Approved Methods of American Association of Cereal Chemists (AACC, 2000) [9].

Preparation of bread sticks

Bread sticks were prepared according to Crassina and Sudha [14] using WF–LRP blends. Ingredients namely compressed yeast 2 g; salt 1 g; sugar 1 g; hydrogenated fat 5 g; ajwain seeds (as a spice) 0.5 g and water (optimum water absorption as determined with the farinograph) were mixed in a Hobart mixer (Model N-50, Hobart, GmbH, Offenburg, Germany) with a flat blade for 4 min at 61 rpm. The dough obtained was fermented in a fermentation/proving cabinet (National Manufacturing Co, Lincoln, USA) maintained at 30 °C and 75% relative humidity (RH) for 30 min, remixed, divided into 15 g piece each, molded into 19 cm long with approximate diameter of 10 mm using a bread moulder (APV, Australia), proofed for 15 min at 30 °C, 85% RH in a fermentation/proving cabinet (National Manufacturing Co, Lincoln, USA) and baked for 25 min at 200 °C in a rotary oven (APV, Australia), cooled and packed in polypropylene pouches for further analysis.

Chemical and nutritional composition of bread sticks

The control and bread sticks with different levels of LRP were analyzed for moisture content (method 44-15), total ash (method 08-01) and fat (3–10) according to the standard methods (AACC, 2000). Values reported were average of three determinations.The fiber content was estimated using the AOAC standard method [10].

Minerals namely iron and zinc were analyzed using flame atomic absorption spectrophotometer. The samples were ashed and digested using hydrochloric acid. Instrument parameters such as resonant wavelength, slit width and air–acetylene flow rate that are appropriate for each element was selected. The instrument was calibrated against a range of working standards of each element. Test solution was aspirated and the concentration of the element determined. The entire analysis was carried out in triplicates and the average values are reported on as-is basis [15].

Physico-sensory characteristics of bread sticks

The bread sticks were weighed using a weighing balance. The length was measured using a foot scale and volume of bread stick was measured by rape seed displacement method [16]. The breaking strength of soup stick was measured by following triple beam snap technique using a food texturometer (TAHDi, Stable Microsystems, Surrey, UK) according to the method mentioned by Gains [17]. The sample was rested on two supporting beams spread at a distance of 3 cm. Another beam connected to a moving part was brought down to break the soup stick at a cross head speed at 10 mm/ min using a 10 kg load cell. The peak force (g) at break, representing breaking strength, was recorded and the mean values of triplicates are expressed. Sensory attributes consisting of appearance, texture, taste and overall acceptability on a 9-point hedonic scale was measured according to the method of Hooda and Jood [18] by a semi trained 25 member panel of age group from 25 to 48 years.

Statistical analysis

Duncan’s New Multiple Range Test [19] was performed for the sensory parameters namely appearance, texture, taste and the overall quality using XLSTAT (Addinsoft, New York, NY, USA). A significance level of 5% was adopted for the comparison.

Results and discussion

The WF used in the study contained moisture 13.34%; ash 0.55%; dry gluten 9.47%; wet gluten 28.77%; Zeleny’s sedimentation value 23 ml; falling number 382 s; protein 9.12% and fat 1.98%. The above results indicate that the WF was of medium strong quality. LRP had moisture, ash, protein and fat of 10.28, 5.34, 8.25 and 1.19% respectively. The total dietary fiber of LRP was four times higher than WF (Table 1).

Table 1 Chemical and physical characteristics of WF and LRP
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Rheological properties of WF as influenced by LRP

Farinograph: effect of LRP on dough formation

The farinograph characteristic of WF as influenced by the addition of LRP (0–20%) is illustrated in Fig. 1a. Water absorption, which is the amount of water required for the dough to have a definite consistency, increased from 56% in the control flour to 58% with the addition of LRP. The dough development time and stability values increased from 1.3 to 2.27 and 5.58 to 7.43 min with replacement of 0–20% respectively. The increase in the stability indicate improved dough strength thereby good product making quality. Despite the dilution of wheat proteins which causes weakening of dough strength, the strength was compensated by the interaction of LRP fibers and starch molecules [20]. Crassina and Sudha [14] also observed a similar trend where mango ginger rhizome powder was used to replace WF. These results may be due to the delay in the hydration and development of gluten caused by the presence of fibers. The effect of lotus powder on the rheological characteristics of WF has not been reported yet; however, Balestra et al. [21] reported that addition of ginger powder in bread dough gave rise to a network with higher density of cross-links. A low effect of dietary fiber on dough viscoelastic behavior is desirable, because it may indicate minor changes in soup stick making performance. Mixing tolerance index is inversely proportional to the strength of the dough; higher values indicate lower strength or intolerance to mixing. Use of LRP increased the mixing tolerance index from 5 to 15 FU when LRP increased from 0 to 10% and further increased to 26 FU indicating poor tolerance of the dough to mixing. This may be due to the presence of fiber and other bioactives in LRP and dilution of gluten in WF. The data thus indicate that the dough properties are adversely affected above 10% incorporation of LRP.

Fig. 1
figure 1

Rheological characteristics of wheat flour-lotus rhizome powder blends. LRP lotus rhizome powder, DDT dough development time, DS dough stability, MTI mixing tolerance index, WA water absorption, GT gelatinization temperature, BD break down, SB set back, PV peak viscosity

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Amylograph: effect of LRP on the pasting characteristics

The effect of LRP on the pasting characteristics of WF is presented in Fig. 1b. The gelatinization temperature provides an indication of minimum temperature required to cook. It was observed that there was no significant change in the pasting temperature of the control WF and WF with added LRP (63.4–63.6 °C). This shows that the starch present in LRP swells at the same temperature as the starch in WF which is important during baking stage. The peak viscosity (the highest viscosity of the paste during the heating phase) represents the ability of the starch granules to swell freely before their physical breakdown and also α-amylase activity. The peak viscosity of WF was 699 BU and it showed an increase with the addition of LRP. In a typical amylograph profile, the viscosity increases to a maximum, followed by a decrease to a minimum value as the granules rupture (referred to as the breakdown). As the temperature decreases, the viscosity again increases from the minimum to a final value, which is referred to as the setback [22]. It was observed that the values of the amylographic breakdown marginally increased from 336 to 385 BU as the level of replacement of WF with LRP increased from 0 to 20%. The amylograph parameters have been correlated with texture and product quality. For example, larger breakdown value is considered to be an indicator of better palatability of cooked rice [22]. The setback values which indicate the retrogradation of gelatinized starch granules decreased from 496 to 388 BU with increase in LRP. This reduction may be attributed to the interaction of fibers from LRP with the starch making the swollen granules more fragile and the swollen starch granules gel into a semi solid paste while cooling. Higher setback values are usually correlated with the amylose content of the starch [22]. Amylose restrains swelling and maintains the integrity of swollen granules [23]. Lingshang et al. [5] has reported that the amylose content of lotus rhizome is 25.6–26.6%.

Composition of bread sticks

Nutritional evaluation of the bread sticks is summarized in Table 2. No marked changes in the moisture and fat content of the bread sticks were observed. The ash content which is an index of minerals content was at 1.69 ± 0.02% in control soup stick and it increased to 2.93 ± 0.13% with the incorporation of LRP to 20%. It was observed that control bread sticks had SDF of 0.10 ± 0.00%, IDF of 1.92 ± 0.00% and TDF of 2.02 ± 0.02%. The content of SDF, IDF and TDF increased from 0.56 ± 0.01% to 1.01 ± 0.01%, 2.39 ± 0.02% to 5.32 ± 0.05% and 2.96 ± 0.03% to 6.34 ± 0.12% respectively as the level of replacement increased from 10 to 20%. This is attributed to the fact that dietary fiber content of LRP is higher than that of WF as seen in Table 1. Crassina and Sudha [14] reported that addition of mango ginger (Curcuma amada L) powder increased the dietary fiber content in soupsticks. Control bread sticks had 7.75% total protein with percent in vitro digestibility of 76.59%. A gradual decrease in protein digestibility was observed as the level of LRP increased in the bread sticks. This may be due to protein–polyphenols interactions which led to low in vitro protein digestibility. Sweica et al. [24] studied the effects of addition of polyphenols rich onion skin in bread and observed a reduced protein digestibility.

Table 2 Composition of LRP incorporated bread stick
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Mineral content, total phenolics and total flavonoids

The minerals namely iron and zinc were evaluated for the raw materials and bread sticks (Table 3). The LRP had eight times higher iron content, whereas the zinc was present in double the amount. The control bread sticks had 1.02 mg of iron and 0.35 mg of zinc. With increasing levels of LRP in the bread sticks, the iron and zinc content increased by three times. The recovery of phenolic contents in different samples is influenced by the polarity of extracting solvents and the solubility of this compound in the solvent used for the extraction process [25, 26]. Dongmei et al. [27] have shown that methanol extract had the highest recovery rate against LRP. TPC (mg GAE/g DW) and TFC (µg QE/g DW) of WF, LRP and bread sticks are presented in Table 3. It was observed that the TPC of LRP and WF used in the work were 3.09 ± 0.19 and 0.94 ± 0.01 mg GAE/g respectively. Alvarez-Jubete et al. [28] reported that wheat and wheat bread contained 53.1 and 29.1 mg GAE/100 g of total phenols respectively, whereas Lilei et al. [29] reported 1.31 and 0.87 mg FAE/g in WF and bread respectively. In our study, the control bread sticks where no LRP was added had 0.91 ± 0.03 mg GAE/g of TPC. The TPC in the rhizome powder is higher than reported by Yang et al. [30] (20.1 mg catechin equivalents/100 g dry lotus) which may be partly due to the differences in genotype and extraction method. However, it increased from 1.06 ± 0.05 to 1.22 ± 0.20 mg GAE/g as the level of LRP increased from 10 to 20%. It is seen that LRP is a good source of polyphenols. Lim et al. [31] studied the quality and antioxidant activity of bread containing turmeric (Curcuma longa L.) at different levels (0–8%) and found that the phenolic compounds increased with increasing levels of turmeric powder.

Table 3 Mineral content, total phenolics and total flavonoids in raw materials and bread sticks
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The TFC of LRP (50.44 ± 0.86 µg QE/g) was higher than that of WF (20.56 ± 0.28 µg QE/g). Yang et al. [30] reported a TFC of 3.35 mg rutin equivalents/g for fresh lotus rhizome flesh wherein they have studied the phenolic profiles and antioxidant activity of different parts of the rhizome. Control bread sticks had 16.91 ± 0.57 µg QE/g of total flavonoids whereas the bread sticks prepared with 20% LRP had 44.55 ± 0.28 µg QE/g showing a significant increase in total flavonoids. Phenolics are heat unstable and reactive compounds [32] and the baking process may result in heat damage to phenolic compounds. A comparison of the measured TPC and TFC in WF and control bread sticks suggests that some degradation may have occurred during baking. However, despite the possible loss of bio-actives after baking, all bread sticks containing LRP showed significantly higher amounts of TPC and TFC when compared with the control.

DPPH radical scavenging capacity

Table 4, represents the DPPH radical scavenging capacity of the raw materials and bread sticks prepared from the blends of WF and LRP. The radical scavenging capacity of WF and LRP was 13.32 ± 1.90 and 87.54 ± 0.13% respectively. The process of baking involves thermal and moisture conditions that facilitate the Maillard reaction and, at the same time, the destruction–formation of natural-labile and thermally-induced antioxidant compounds, respectively [33]. Study by Lindenmeier and Hofmann [34] revealed that pronyl-l-lysine is the antioxidant compound that is being produced in bakery products. They observed high amounts of the antioxidant in bread crust, low amounts in the crumb and absence of the compound in untreated flour showing that the amounts of pronyl-l-lysine to be strongly influenced by the intensity of the thermal treatment. In our study also, a comparison of the measured DPPH radical scavenging capacity of the blends (40.50 ± 0.23 to 57.02 ± 1.02%) and the bread sticks (44.76 ± 2.01 to 60.62 ± 1.11%) showed that the antioxidant activity of the bread sticks had possibly increased due to the baking process. The antioxidant and radical scavenging activities are closely related to polyphenols [29]. The higher radical scavenging capacity of the bread sticks to their counterpart blends in our study may be attributed to the known phenomenon that fermentation helps in the release of fiber-bound phenolics. It is also in agreement with the studies of Gelinas and Mc Kinnon [35]; Han and Koh [36], which indicated that the antioxidant activities of phenolic acids increased during fermentation and the baking process. Similarly, Lilei et al. [29] reported that baking increased the antioxidant activities (ORAC values) of whole wheat and refined flour by 1.8 and 2.9 times and increased ferulic acid content from 18 to 35%. However, in contrast to this they also mentioned that baking decreased DPPH radical scavenging capacity by 32 and 30%. Earlier studies by Hansen et al. [37], Liukkonen et al. [38] and Jeffrey et al. [39] had revealed that fermentation increases soluble ferulic acid content in cereals. Jeffrey et al. [39] also concluded from their study that increased baking temperature and time increased all evaluated antioxidant properties (ORAC, DPPH, ABTS) in whole-wheat pizza crust. Recent study by Yang et al. [30] suggested that catechin and epicatechin were the primary contributors to the antioxidant activity of lotus root. A subsequent increase in the radical scavenging capacity was observed as the level of LRP increased from 0 to 20% in the bread stick. Control bread stick was found to have 37.04 ± 1.99% radical inhibition capacities. Bread sticks prepared form LRP blends were observed to have higher radical inhibition capacity compared to the control. This is due to the radical inhibition capacity contributed by the LRP which is higher than the WF alone.

Table 4 DPPH radical scavenging capacity of blends and bread sticks
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Physico-sensory characteristics

The data on physico-sensory characteristics of bread sticks is shown in Table 5 while Fig. 2 shows the picture of bread sticks with varying levels of LRP. The addition of LRP did not have any significant effect on the physical properties such as weight, length and volume of the bread sticks as compared to that of the control. However, there was a change in the viscoelastic behavior of the dough as seen in the decrease in the length of the bread stick with the addition of LRP. This indicate that resistance to deformation increased owing to the interactions between fiber structure of LRP and the wheat proteins. A decrease in the length of bread sticks from 18.50 ± 0.70 to 17.62 ± 1.10 cm was seen with increasing levels of LRP from 0 to 20%. The volume of the bread sticks marginally decreased. The texture of bread sticks measured in terms of breaking strength was 54.30 ± 1.10 g force for the control and on increase in addition of LRP, the bread sticks became crispy as seen in the decrease in breaking strength values from 40.42 ± 0.67 to 31.21 ± 1.37 g force. Crispiness is a desirable attribute for breadsticks. Alamprese et al. [40] observed that whole-wheat bread sticks became less crisp during storage due to moisture absorption.The sensory scores for appearance which was 9 for control, marginally decreased to 8 till 15% addition and at 20% addition of LRP, the scores decreased to 5. Increase in addition of LRP in the blend, decreased the sheen and increased the surface dryness due to higher fiber content in LRP. Similar trend was observed for texture values and taste. Increase in LRP increased the blandness in the taste, which reduced the sensory scores of the bread sticks beyond 15% incorporation. The texture of control breadsticks was hard as also indicated by higher breaking strength values and on increase in LRP, the crispiness increased. Overall quality scores showed that addition of LRP at 10–15% were nearer to control, whereas at 20%, the scores was quite low indicating the product unacceptability at 20%.

Table 5 Quality characteristics of bread sticks
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Fig. 2
figure 2

Photograph of bread sticks with varying levels of LRP. 1–0% LRP; 2–10% LRP; 3–15%LRP; 4–20% LRP; LRP lotus rhizome powder

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Conclusion

In this study, LRP blended wheat based bread stick was developed and analyzed for different physical characteristics, bioactive components and antioxidant activities as affected by different substitution levels of LRP. The results showed that up to 15% LRP could be included in a bread stick formulation without any significant interference with the sensory acceptability of the bread sticks. We may conclude that since lotus rhizome is a good source of bioactive components such as dietary fiber, polyphenols and flavonoids with high antioxidant activity, substitution of WF with LRP in WF based products like bread sticks can improve the health of consumers. Further, this formulation can be considered to widen the use of rhizomes in bakery industry.