Influence of Pectin and Guar Gum Composite Flour on Plasma Biochemical Profile of Streptozotocin-Induced Diabetic Male Albino Rats

Abstract

Dietary fiber sources like pectin and guar gum have gained immense importance because of their constructive role of releasing sugars and absorbing sugars slowly in the intestinal tract; consequently, they reduce the severity of diabetes mellitus. In this article, we explore the hypoglycemic and hypocholesterolemic worths of pectin and guar gum by adding them to wheat flour to prepare chapaties, a staple in the diet of those in the Indo-Pak region. For this purpose, four different treatments of composite flour were prepared by adding wheat flour with 3% pectin (T₁), 3% guar gum (T₂), a combination of pectin and guar gum 2% of each (T₃), along with a control (T₀). Composite flour samples were stored for 60 days and analyzed for proximate analysis on monthly basis. Protein and fat contents of different treatments were affected significantly by various storage intervals. Likewise, chapaties were prepared after every storage interval and evaluated for sensoric attributes. Chapaties prepared from treatment T₂ gained highest score with respect to all sensory parameters (color, flavor, taste, texture, chewing ability, and foldability) followed by T₀, T₃, and T_1. The chapaties prepared from composite flour samples were fed to Streptozotocin-induced diabetic male albino rats to find out their impact on plasma biochemical profile. After decapitating the rats, the blood sample was collected and analyzed for blood glucose, total cholesterol, and triglycerides level. Rats fed on T_2, showed highest reduction (17.09%) in blood glucose followed by T₁ (10.25%) and T₃ (9.24%), as compared to T₀. Blood cholesterol was reduced up to 18.88, 16.76, and 12.23% by T₂, T₃, and T₁, respectively than that of control. Similarly, T₂ showed the highest reduction (16.33%) in blood triglyceride level followed by T₃ (12.31%) and T₁ (2.76%). We concluded from our research study that dietary fibers from guar gum and pectin are helpful in reducing blood glucose and cholesterol levels. It is further suggested that guar gum is more effective than pectin to control these diseases. For this reason, wheat flour should be supplemented with dietary fiber sources to combat the existing dilemma of diabetes and hypercholesterolemia.

Keywords:

• Composite flour
• Pectin
• Guar gum
• Plasma glucose
• Streptozotocin-induced diabetes
• Albino rats

INTRODUCTION

Diabetes mellitus (DM) is so common today that the professionals refer it to as an epidemic. There are about 100 million diabetic patients in the world and the number is likely to increase by two-fold in the year 2010. Pakistan falls in the area of high prevalence of diabetics.[1] About 9 million people in Pakistan are diabetic with causes not fully known. This figure, owing to lack of any comprehensive strategy to check its growth, is likely to take a frightening leap to 14.3 million in the next 25 years, as predicated by the World Health Organization.[2]

Hypercholesterolemia is associated with diabetes and is a risk factor for cardiovascular diseases.[3] Cholesterol is a vital substance that is synthesized by the liver. The body uses cholesterol as a building block for essential organic molecules such as steroid hormones, cell membranes, and bile acids. However, its excessive accumulation leads to coronary heart diseases.

Dietary fiber has established benefits for health maintenance, disease prevention, and as a component of medical nutrition therapy. It consists of the storage and cell wall polysaccharides of plants that cannot be hydrolyzed by human digestive enzymes. Consumption of viscous dietary fibers lowers blood cholesterol levels and helps to normalize blood glucose and insulin levels.[4] A high intake of dietary fiber, particularly of the soluble type, above the level recommended by the ADA, improves glycemic control, decreases hyperinsulinemia, and lowers plasma lipid concentrations in patients with type-2 diabetes.[5]

The addition of viscous dietary fibers slows gastric emptying rates, digestion, and the absorption of glucose to benefit immediate postprandial glucose metabolism[6] and long-term glucose control[7,8] in individuals with diabetes mellitus. The long-term ingestion of 50 g of dietary fiber per day for 24 weeks significantly improved glycemic control in diabetic individuals.[9,10] The role of fiber in the prevention and treatment of cardiovascular disease is related to its effect on blood cholesterol levels.[11] Soluble fiber, like guar gum and pectin, may improve glucose tolerance to carbohydrates, satisfy appetite without supplying energy due to more bulk (of benefit to the obese), reduce the risk of peptic and duodenal ulcers, lower plasma cholesterol levels (potentially reducing the risk of heart disease), and protect against the potential adverse actions of toxins.[12,13]

Composite flours have proven to have practical uses in the cereal industry in many parts of the world. People accept cereal-based products easily made from composite flours. Keeping in mind the beneficial effects of fiber, we designed the present article to fortify wheat flour with guar gum and pectin to prepare different composite flours higher in dietary fiber. In Pakistan, wheat is mainly used in the form of flour for the preparation of unleavened flat bread called chapaties—a staple of the diet in the region.[14] This can be an ideal vehicle for the distribution of fiber-enriched composite flours to improve the nutritional and functional properties of the wheat flour and ultimately to achieve the goal of controlling diabetes and high cholesterol.

MATERIALS AND METHODS

Procurement of Raw Materials

To make the rats diabetic, Streptozotocin injection was purchased from Pakland Chemicals, Lahore, Pakistan. Commercial wheat flour (atta) produced by roller flour mills as a resultant product, pectin, and guar gum were procured from the local market.

Preparation of Composite Flours

The particle size of pectin and guar gum was reduced into fine flour using a Cyclotec- 1093 Sample Mill (Tecator, Sweden). After conducting some preliminary trials, wheat flour was blended with pectin and guar gum in different combinations as mentioned in Table 1. Each treatment of composite flour was thoroughly mixed and sieved in order to achieve a uniform dispersion of flour particles with wheat flour. Three homogenous replicates for each composite flour were taken and stored at room temperature in woven polypropylene bags for 60 days.

Table 1 Different treatments used to prepare composite flours

Treatments	Wheat flour (%)	Pectin %	Guar gum %
T0	100	–	–
T1	97	3	–
T2	97	–	3
T3	96	2	2
T0 = Commercial wheat flour without any supplementation (acts ac control treatment); T1 = wheat flour supplemented with 3% pectin; T2 = wheat flour supplemented with 3% guar gum; and T3 = wheat flour supplemented with a combination of guar gum and pectin, 2% each.

Proximate Analysis of Composite Flours

Each treatment was analyzed for the moisture (method No. 44-15 A), crude protein (method No. 46-10), crude fat (method No. 30-10), crude fiber (method No. 32-10), total ash (method No. 08-01), and NFE contents by following their respective procedures described in AACC.[15] Flour stored for 0, 30, or 60 days was analyzed and made into chapaties.

Preparation of Chapaties

Chapaties were prepared from each composite flour.[16] The dough was made by mixing 200 g atta with predetermined quantity of water for 3 minutes and kept for 30 minutes as resting period. A dough piece weighing 80 g was rolled with a wooden roller pin on a specially designed wooden platform in a sheet of 3 mm thickness. This was cut into circle of 17 cm in diameter. The chapaties were baked from both sides on thermostatically controlled hot plate at a temperature of 210°C for 1.5 minutes.

Sensory Evaluation of Chapaties

Sensory evaluation of chapaties was carried out for color, flavor, texture, chewing ability, appearance, and folding ability using hedonic score system.[17] Sensory evaluation was done immediately after baking by a trained taste panel, using a 9-Point Hedonic Score System.

Efficacy Studies

Efficacy studies were conducted using Streptozotocin-induced diabetic male albino rats to find out the impact of various composite flours on plasma glucose, total cholesterol, and triglyceride levels. Twenty rats were randomly divided into four groups of five. Each rat was placed in a separate cage. All rats had been injected intravenously with Streptozotocin (STZ) @ 60 mg/Kg body weight in 0.01M citrate buffer having 4.5 pH.[18] The blood sample of each rat after injecting Streptozotocin was analyzed to check whether all the rats had become diabetic or not. All rats in each group were fed on a particular diet. The chapaties prepared from individual composite flours were mixed with other ingredients of the diet (corn oil, corn starch, vitamin mixture, salt mixture, Chapatties) such that 12% of the protein was made available from the final diets to meet the rat's nutrient requirements. The temperature and humidity were recorded daily throughout the study. After 40 days, all the rats were decapitated, and their blood samples were collected to analyze the plasma biochemical profile.

ANALYSES OF PLASMA BIOCHEMICAL PROFILE

Procurement of Plasma

Twelve drops of EDTA (anticoagulant) was put in labeled centrifuge tubes. The blood from each rat was collected in the centrifuge tubes. The blood samples were then put in the centrifuge machine and were centrifuged at 3000 rpm for 10 minutes.[19] Plasma of each sample was then collected in already labeled plasma cups (ependrof tubes) with the help of micropipette, which was then frozen (−20°C) for plasma glucose, cholesterol, and triglycerides analysis by conducting the following analyses.

Glucose, Cholesterol, Triglycerides

Blood glucose concentration was calculated by GOD-PAP method[20] using the commercial laboratory kit. Cholesterol in the collected plasma of individual rats of all groups was measured by liquid cholesterol CHOD–PAP method[21] to find out the effect of individual diet on the cholesterol level of respective groups. Triglycerides in individual rats were measured by liquid triglycerides GPO-PAP method.[22]

Statistical Analysis

Analysis of variance for each parameter was carried out to determine the level of significance by using analysis of variance (ANOVA) technique. The significant differences were compared using Duncan's Multiple Range Test.[23]

RESULTS AND DISCUSSION

Proximate Analysis of Composite Flour

Composite flours, along with control, were statistically analyzed and means for moisture content, crude protein, crude fat, crude fiber, total ash, and NFE content are given in Table 2 and Table 3. The moisture content is of great importance for many scientific, technical and economic reasons. The lower the flour moisture, the better its storage stability. The results regarding means for moisture content of various composite flour treatments clearly indicated that the moisture contents of different treatments varied significantly. It is obvious from the results that the moisture content is significantly higher in T₂ (11.94%) followed by T₁ (11.63%), and T₃ (11.61%) as compared to T₀ (commercial wheat flour, i.e., 11.34%). There was gradual increase (11.34–11.95%) in moisture content during 60 days storage. The higher moisture content in composite flours containing fibers is probably due to higher water-holding capacity of these fibers. It may be concluded that greater the percentage of dietary fibers in composite flour, greater is the amount of moisture content.[24] Storage has decisive significant effect on moisture content of composite flour samples. Minimum moisture content 11.34% was observed at the initiation of the study while maximum moisture content 11.95% after 60 days storage (Table 3). There was a gradual increase in moisture contents in various composite flour samples with an increase in storage period. The water content also influences the storage stability of wheat flour. It should be below 14% to prevent microbial growth and chemical changes during storage.[25] Increase in moisture content was observed in resultant atta during five months storage. In this article, commercial wheat flour was used with moisture content 11.61% that also followed a similar trend. Higher moisture content in composite flours might be due to variation in the processing conditions, method of grinding, as well as the variations in temperature and climatic conditions.[26]

Table 2 Means∗ for effect of various treatments on proximate composition (%) of composite flours

	Proximate composition (%)
Treatments	Moisture	Crude protein	Crude fat	Crude fiber	Ash	NFE
T0	11.34^b ± 0.13	11.25^d ± 0.05^d	1.30 ± 0.03	0.58^c ± 0.01	1.40^d ± 0.080	74.10^a ± 0.19
T1	11.63^ab ± 0.15	11.35^c ± 0.19	1.32 ± 0.03	0.64^b ± 0.01	1.44^c ± 0.03	73.70^ab ± 0.31
T2	11.94^a ± 0.15	11.62^b ± 0.10	1.35 ± 0.03	0.70^a ± 0.01	1.56^b ± 0.03	72.73^c ± 0.28
T3	11.61^ab ± 0.11	11.66^a ± 0.15	1.34 ± 0.03	0.71^a ± 0.01	1.60^a ± 0.03	73.08^bc ± 0.22
Mean values for treatments carrying same letters in a column are not significantly different; ∗n = 9.
Note: Superscripts indicate the implementation of statistical technique DMR (Duncan's Multiple Range test) values.

Table 3 Means∗ for effect of different storage intervals (days) on proximate composition of composite flours

	Proximate composition (%)
Storage (days)	Moisture	Crude protein	Crude fat	Crude fiber	Ash	NFE
0	11.34^c ± 0.00	11.72^a ± 0.09	1.38^a ± 0.02	0.65 ± 0.02	1.50 ± 0.07	73.50 ± 0.25
30	11.63^b ± 0.12	11.42^ab ± 0.07	1.33^ab ± 0.02	0.66 ± 0.02	1.49 ± 0.04	73.47 ± 0.26
60	11.95^a ± 0.09	11.26^b ± 0.15	1.27^b ± 0.02	0.66 ± 0.19	1.50 ± 0.04	73.26 ± 0.30
Mean values for treatments carrying same letters in a column are not significantly different; ∗n = 12.
Note: Superscripts indicate the implementation of statistical technique DMR (Duncan's Multiple Range test) values.

Protein content is an index of flour quality. Mean squares for protein content (Tables 2, 3) indicated that the crude protein was significantly affected as a function of storage and treatment while the storage × treatment interaction was non significant differences. The crude protein contents of T₃ (11.66%) was significantly highest followed by T₂ (11.62%) and T₁ (11.35%), while T₀ (11.25%) showed the lowest significant value for crude protein content. In previous studies, it has been found that the crude protein contents of commercial wheat flour ranged between 7.98–13.16%.[27,28,29] Storage has significant effect on protein content in various composite flours. It is evident from the means that there is a significant decrease in protein content during storage. At day 0, the mean value for protein was 11.72%, which decreased significantly to 11.42% and 11.26% after 30 and 60 days of storage, respectively. In the present investigation, the decrease in protein content in the commercial flour and remaining samples was due to the absorption of moisture from the atmosphere that accelerated the proteolytic activity of the enzymes. The proteases are responsible for the degradation of protein during storage. It has been observed that that activity of proteases and lipases in the resultant atta is higher (2 to 2.5 times) than chakki atta.[25] The higher enzyme activity might be due to the presence of higher amount of enzyme rich parts like germ and aleurone layer of the wheat.

The mean squares for crude fat content of various composite flours (Tables 2, 3) showed that treatments had non-significant where as storage had significant effect on the fat content of various treatmenst. The overall range for the crude fat contents of control along with the different composite flour samples was found to be 1.30%–1.35%. The results concerning crude fat contents in the present investigation are closely associated with the earlier results[28,30,31] that the crude fat contents in different commercial wheat flours ranged between 0.80–1.42%. Storage has highly significant effect on various treatments of composite flours (Table 3). The mean for fat content was 1.38% at day 0, which decreased to 1.33% and 1.27% after 30 and 60 days of storage, respectively. The results showed that there was a gradual decrease in fat contents with the passage of time. This decrease in fat contents may be attributed to the development of rancidity.[25] Fat deterioration during storage might be due to the activation of lipase enzyme, which might split off the fat into free fatty acids and glycerol in the presence of moisture and other proxidants like light and heat. The condition conducive to lipolysis was high moisture content of the flour during storage as in present case. The fat content was gradually split up into glycerin and fatty acids by the enzyme lipase, and then acids were oxidized by taking up oxygen. This process also happens during the storage of flour, rendering it sour and rancid. The lower the flour moisture, the better its storage quality, especially in silos.[32]

Crude fiber is an insoluble and combustible organic residue which remains after the sample has been treated with dilute acid and alkali under prescribed conditions. The mean squares for crude fiber content of different composite flours (Tables 2, 3) indicated that significant differences exist in the crude fiber due to treatments however it was remained unaffected by storage. The means for the crude fiber content of different composite flours exhibited that T₃ and T₂ had the highest significant value (0.71%) followed by 0.64% in T₁ (pectin 3%), while T₀ (wheat flour 100%) showed the lowest value (0.58%) for this trait. Addition of guar gum at different levels in commercial wheat flour increases the value for crude fiber content. The results for crude fiber contents in the present article are in concordance with the earlier findings[33] that commercial flour contained 0.59%, while composite flour of 3% guar gum contained 0.84% fiber contents. Storage has non-significant effect on various treatments of composite flours. The fiber content has non-significant influence due to storage intervals confirmed the present results.[34]

The ash content in the food stuff represents inorganic residue remaining after the organic matter has been burnt away. It is generally correlated with the amount of bran and is considered as rough indicator of flour yield. The analysis regarding mean squares for composite flours (Tables 2, 3) indicated that it differed significantly with treatments. However, the ash content was non-significantly influenced by storage. The means for ash content of various flour samples revealed that it ranged from 1.40–1.60%. The highest value (1.60%) was noted in the T₃ (pectin 2% + guar gum 2%), while T₀ (commercial wheat flour) showed the lowest value (1.40%). The ash content of composite flour having 1–2% guar gum ranged from 1.51–1.58%.[31] Commercial wheat flour had minimum ash content because of absence of any type of enrichment with guar gum and pectin. The ash content in food stuff not necessarily accounts for exactly the same composition as the mineral matter present in the original food, there may be some losses due to volatilization or some interaction between the constituents.[35] Means for ash content showed non significant effect of storage in different composite flour samples.[36]

Mean squares for NFE content of composite flours (Tables 2, 3) indicated that various treatments of composite flours showed significant differences on NFE content where as storage and interaction remained non-significant. Highest significant NFE content was found in T₀ (74.10%) followed by T₁ (73.70%) and T₃ (73.08%). The lowest value for NFE was found in T₂ (72.73%). Whole wheat flour of Pakistani varieties contained 80.43% NFE contents.[26] Means for NFE revealed non significant differences due to storage. At 0 day, the means for NFE were 73.50% followed by 73.47% and 73.26% after 30 and 60 days storage, respectively. The significant results regarding treatments were due to significant changes in crude protein and crude fat contents during storage of different flour samples. In previous studies on composite flours, NFE contents of commercial flour and composite flour of 3% guar gum were to be 73.14 and 73.69% respectively.[30]

Sensory Evaluation of Chapaties

Sensory evaluation is usually performed towards the end of the product development or formulation cycle. It is carried out to assess the reaction of judges towards the product, and they rate the liking on a scale. Chapati is the staple diet of the region; the consumer favors a chapati of light brown and creamy in color, which can be smashed to pieces of the desired size and folded to form a scoop. The sensory evaluation of chapati for various sensory attributes was carried out by trained taste panel. Mean squares for sensory evaluation are presented in Table 4 and Table 5, with respect to various sensory attributes. The results indicated that the quality characteristics of chapati such as color, flavor, taste, texture, chewingability and foldingability scores differed significantly due to storage and various treatments of composite flours. Outcomes regarding each sensory attribute are discussed following.

Table 4 Means for effect of different composite flour treatments on sensory attributes of chapaties

	Sensory attributes of composite flour chapaties
Treatments	Color	Flavor	Taste	Texture	Chewing ability	Folding ability
T0	6.46^b ± 0.26	6.36^b ± 0.26	7.16^b ± 0.28	6.63^b ± 0.24	6.16^b ± 0.26	6.53^b ± 0.22
T1	5.00^d ± 0.34	5.10^d ± 0.28	5.96^d ± 0.30	5.36^d ± 0.31	5.00^d ± 0.32	5.06^d ± 0.27
T2	7.10^a ± 0.24	7.10^a ± 0.22	7.86^a ± 0.22	7.20^a ± 0.23	6.70^a ± 0.27	7.03^a ± 0.23
T3	5.66^c ± 0.27	5.56^c ± 0.30	6.56^c ± 0.28	6.10^c ± 0.27	5.50^c ± 0.30	5.73^c ± 2.26
Note: Superscripts indicate the implementation of statistical technique DMR (Duncan's Multiple Range test) values.

Table 5 Means for effect of different storage intervals (days) on sensory attributes of chapaties

	Sensory attributes of composite flour chapaties
Storage (days)	Color	Flavor	Taste	Texture	Chewing ability	Folding ability
0	6.95^a ± 0.23	6.87^a ± 0.22	7.77^a ± 0.20	7.20^a ± 0.19	6.77^a ± 0.21	6.92^a ± 0.22
30	6.02^b ± 0.28	6.05^b ± 0.24	6.90^b ± 0.24	6.27^b ± 0.22	5.82^b ± 0.22	6.02^b ± 0.24
60	5.20^c ± 0.28	5.17^c ± 0.28	6.00^c ± 0.24	5.50^c ± 0.25	4.92^c ± 0.21	5.32^c ± 0.25
Note: Superscripts indicate the implementation of statistical technique DMR (Duncan's Multiple Range test) values.

In baking, color serves as a cue for the doneness of foods and is correlated with changes in aroma and flavor. There was a highly significant variation among different treatments for color and during storage. It is apparent from the results that the chapatti prepared from T₂ had the highest significant value (7.10) for color. The lowest score for color (5.00) was obtained by chapaties prepared from T₁ (composite flour with 3% pectin). Decrease in color scores was observed with increase in the level of replacement in composite flours.[37] Addition of guar gum in different combinations for the preparation of chapaties showed a significant role towards the color of chapaties.[33] Storage has significant effect on color of chapaties prepared from different composite flours. At the beginning, means for color of chapaties during storage was 6.95 which decreased to 6.02 and 5.20 after 30 and 60 days storage. The chapati prepared with 3% guar gum gave whiter look and puffiness to the chapati. The deterioration in color of chapaties with storage might be due to the absorption of moisture, oxidation of fats & carotenoids, rancidity, and progressive increase in the mold count of composite flour samples with the passage of time. In previous studies, chapaties prepared from 3% guar gum were found to have the highest significant value (8.00) for color.[33]

Perceptions of flavor are a synthesis of taste and smell impressions, along with texture and are even influenced by appearance. The mean scores for flavor of chapaties prepared from different flour samples revealed that chapati prepared from T₂ (guar gum 3%) obtained the highest flavor score (7.10) followed by T₀ (commercial wheat flour) and T₃ (2% guar gum + 2% pectin). Flavor of chapaties prepared from composite flours were affected by the nature of the fiber sources used for mixing. Particularly pectin has a typical flavor that resulted in lowering scores in chapati. Better score for flavor of chapaties prepared from guar gum flour blends may be attributed due to the good look, refinement in color and texture by the addition of guar gum. Storage has significant effect on flavor of chapaties prepared from different composite flours. At day 0, mean for flavor of chapaties was 6.87, which decreased to 6.05 and 5.17 after 30 and 60 days of storage. During storage, increase in free fatty acid, alcoholic acidity, and lipase activity was observed during five months storage that is the key factors leading towards flour deterioration.[25] In the present article, an increase in the flour moisture during storage also favor hydrolytic rancidity—all these are the contributory factors towards the decline of flavor score.

The taste is a sensation perceived by the tongue and influenced by the texture, flavor and composition of the foods. Mean scores for taste of chapaties prepared form different flour samples showed that the highest taste score (7.86) gained by T₂ (guar gum 3%) while T₁ was assigned the lowest taste score (5.96). Decrease in taste scores of Egyptian Balady bread was noted when wheat flour was supplemented with processed cotton seed flour;[38] even biscuits produced from blended flours have been found to loose score for taste and tang.[39] There was gradual decease in taste mean sores with increased level of supplementation.[40] Storage has momentous effect on taste of chapaties prepared from different composite flours. At the initiation of the study, mean for taste of chapaties during storage was 6.77 which decreased to 6.90 and 6.00 after 30 and 60 days of storage. In a similar study, deterioration in the taste score was observed at the end of 3 month studies. Furthermore, they concluded that decrease in taste scores were due to mold growth and insect infestation during storage. Similarly in the present study, taste scores decreased during storage that might be due to the rancidity of fat in flour owing to oxidation.

The International Standards Organization defined texture of a food product as all the rheological and structural attributes of the product perceptible by means of mechanical, tactile, and where appropriate, visual and auditory receptors.[41] Food texture may be extremely important to the consumer. Yet, unlike color and flavor, texture is used by the consumer not as an indicator of food safety but as an indicator of food quality. The mean scores for texture of chapaties prepared form different composite flours revealed highly significant effect of legumes blended in different proportions on texture of chapaties. Highest texture score (7.20) was assigned to T₂ (guar gum 3%) flour sample followed by T₀ and T₃ which obtained 6.63 and 6.10 scores, respectively. In the present work, flour samples of different composition were used, varying amount of protein, dietary fiber, and other characteristics. This deviation is the main factor for different sensory rating of texture scores. In another study, it was observed that that the texture of chapati significantly varied due to its composition.[42] Storage has significant effect on texture of chapaties prepared from different composite flours. Initially, mean scores for texture were 7.20 which subsequently decreased to 6.27 and 5.50 after 30 and 60 days of storage. Increase in moisture, peroxide value, and acidity of flour samples has an inverse correlation with regard to texture. All these factors contribute towards deterioration in the quality of stored flour samples and ultimately influence the texture of chapati.

Mean scores for chewing ability of chapaties prepared from different composite flours revealed momentous effect of fortification in different proportions on chewing ability of chapaties. Chapaties prepared from T₂ (guar gum 3%) flour sample acquired the highest chewing ability score (6.70) followed by T₀ (wheat flour 100%) and T₃ (guar gum 2% + pectin 2%) that got 6.16 and 5.50 scores, respectively, while least scores (5.00) were assigned to T₁ (pectin 3%). Storage also has momentous effect on chewing ability of chapaties prepared from different composite flours. Mean scores for chewing ability were 6.77 at the start which decreased to 5.82 and 4.92 after 30 and 60 days storage.

One of the preferred textural features of chapaties is ease with which it can be split into smaller pieces and folded to form a scoop for picking up the curry. Mean scores for folding ability of chapaties prepared from different flours revealed that chapaties prepared from T₂ (guar gum 3%) flour sample attained the highest folding ability score (7.03) while minimum score (5.06) was assigned to T₁ (pectin 3%). Decrease in the mean score was observed with the increase in level of supplement. The results also exposed that apart from the commercial flour, chapaties prepared from composite flour containing guar gum at 3% level gave a product, which received even higher scores. Addition of pectin produced chapaties that got lowest score regarding this attribute. The findings of the present study agreed with regard to hand feel, which was smooth and highly pliable particularly in case of chapaties containing guar gum, which may partly be due to higher moisture absorption capacity of the guar gum. In a prior study, a decrease in the folding ability score of chapati was observed with an increase in the level of supplementation with winged bean.[43] Storage has significant effect on folding ability of chapaties prepared from different composite flours. In the beginning, the mean score for folding ability was 6.92, a decreasing trend in scores i.e., 6.02 and 5.32, was observed after 30 and 60 days storage, respectively. Similar trends have been observed in previous studies in which dietetic flour was prepared by supplementing with guar gum[33] and chickpea.[30]

Efficacy Studies

The diets prepared from composite flour chapaties were fed to the male albino rats for a period 40 days to determine the effect of individual diet on plasma biochemical profile including glucose, cholesterol, and triglycerides. Rats were used because of their convenient handling, close supervision, and feasibility of controlled environmental conditions, which would have been difficult in case of humans. Moreover, it was really difficult to find out the volunteers who could be restricted only to chapati made from selected composite flours. The results regarding each parameter are summarized following.

Glucose

Regarding plasma glucose, it is apparent that there are highly significant variations among different groups of rats fed on various diets of composite flours (Table 6). Maximum glucose concentration was found to be 316 mg/dL in control group (T₀) followed by 286.8 (T₃) and 283.6 (T₁) mg/dL in groups fed on a combination of guar gum 2% + pectin 2% and pectin 3%, respectively. Lowest glucose concentration (262 mg/dL) was observed in rats fed on guar gum 3% (T₂). Percent decrease of glucose concentration in different groups of rats with reference to control showed that guar gum 3% showed maximum reduction (17.09%) followed by pectin 3% and guar gum 2% + pectrin 2% induced 10.25% and 9.24% reduction in glucose as compared to control (Fig. 1). From the present investigation, it was concluded that there is 9–17% decline in glucose by the addition of selected treatments.

Table 6 Means∗ for plasma biochemical profile (mg/dL) of diabetic rats

	Treatments
Plasma bio-chemical profile	T0	T1	T2	T3	Std. Error
Glucose	316.00^a	283.60^c	262.00^d	286.80^b	0.71
Cholesterol	75.20^a	66.00^b	61.00^d	62.60^c	0.63
Triglycerides	79.60^a	77.40^a	66.60^c	69.80^b	0.62
∗n = 5.
Note: Superscripts indicate the implementation of statistical technique DMR (Duncan's Multiple Range test) values.

Figure 1 Percent decrease in plasma glucose of diabetic rats with reference to control.

The results found in present investigation are corroborated with the previous findings of various workers, that dietary fiber sources are helpful in the management of plasma glucose. The blood glucose level of STZ induced diabetic rat was found to be > 325 mg/dL.[18] Guar gum in the diet (10% and 20% concentration) decrease blood glucose significantly.[44] In a study, blood glucose response to a test meal was significantly reduced by guar gum compared with the placebo after a 4-week guar-gum supplementation.[45] In another study, 24.56% reduction in plasma glucose in humans, by incorporating guar in bread.[46] The effectiveness of guar gum is authenticated by the fact that the hyperglycemia and hyperlipidemia can be controlled by a herbal powder consisting of guar gum, methi, tundika and meshasringi. The herbal powder was given twice a day before major meals for four weeks.[47] Metabolic response to short and long-term guar gum consumption were studied in adolescent and adult rats. Reduced plasma glucose was measured for only the guar gum group.[48] Hypoglycemic effect of guar gum indicated by the addition of guar to an intragastric glucose load (1 g/kg) markedly delayed the rise in plasma glucose levels when the concentration of the gum was adequate (10 mg/ml). Guar gum can reduce fasting blood glucose from 205.42 ± 66.6 mg/dL (11.4 ± 3.7 mmol L⁻¹) to 171.19 ± 70.28 mg/dL (9.5 ± 3.9 mmol L⁻¹) in rats.[49,50] Various other researchers also proved the phenomena of hypoglycemia due to guar and guar by-products (GBP) which affected carbohydrate tolerance in rats. Both 1% and 10% GBP suspensions administered immediately before a glucose challenge (1 g/kg body weight) caused a 31% reduction in the integrated plasma glucose response area during a 180-minute test.[51] Guar gum showed greater viscosity than the other gums during acidification and/or alkalinization and also showed larger effects on plasma glucose levels (35% reduction in maximum rise in plasma glucose), and on the total area under the curve of plasma glucose.[52]

The acute effect of a single dose of guar gum has been verified to reduce the peak postprandial whole blood glucose levels (about 10%). Following long-term treatment, a further reduction was seen in the obese subjects with the highest postprandial glucose levels.[53] Clinical impact of fiber supplementation for the reduction of postprandial blood glucose showed 17% decrease in mean daily plasma glucose concentrations while the area under the curve for 2-hour plasma glucose concentrations reduced by 36%.[54] Inclusion of guar gum into a high carbohydrate diet, compared with wheat bran, is effective in lowering plasma glucose concentrations in STZ diabetic rats that were food-deprived overnight. Furthermore, the guar diet improved insulin sensitivity in STZ diabetic rats, with the majority of the improvements resulting from increased insulin sensitivity of glucose disposal in peripheral tissues.[55] From the present facts, it is concluded that it is desirable to incorporate guar gum at the mandatory levels as proved in the study in the staple diet, i.e., chapati, a good attempt to reduce plasma glucose level of hyperglycemic persons.

Cholesterol

Means for plasma cholesterol in different groups of rats showed highly significant variations among groups (Table 6). Maximum plasma cholesterol was found to be 75.20 mg/dL in control group (T₀) followed by 66.00 and 62.60 mg/dL in rats fed on T₁ and T_3, respectively. Lowest plasma cholesterol was observed in rats fed on T₂ (61.00 mg/dL). The results indicated that there is a subsequent diminishing trend in the cholesterol concentration with progressive increase in guar gum concentration. It was further observed that T₃ showed a significant reduction (18.88%) in plasma cholesterol followed by T₃ (16.76%) and T₁ (12.23%) as contrast to control (Fig. 2).

Figure 2 Percent decrease in plasma cholesterol of diabetic rats with reference to control.

Cholesterol lowering effects are most often associated with gelling, mucilaginous, viscous fibers such as guar gum as well as pectin; the only cardiovascular risk factor that has been shown to be modifiable by dietary soluble fiber is the level of plasma cholesterol.[56,57] The outcomes of the present study are in close line with the previous findings. The cholesterol level in STZ diabetic rats was found to be 69 ± 7.[18] Guar gum in the diet (10% and 20% concentration) decreased blood total cholesterol significantly.[44] Dietary fiber is inevitable for hypercholesterolemic and hyperglycemic individuals because 1g/ Kg of soluble fiber from guar gum can lower total cholesterol by about 0.045 mmol/L.[58] Guar gum has shown 10–15% reduction in plasma total cholesterol after short-term treatment. However, daily consumption of 15 g guar gum considerably reduces plasma total cholesterol.[59] In another study, 16.6% reduction in total cholesterol was observed by using guar gum in capsule form.[60] Approximately 40% lower cholesterol concentration was achieved with daily consumption of 8–36 g guar gum.[61,62] It has been proved through clinical trials that, when used alone, guar gum may reduce plasma total cholesterol by 10–15%.[63,64,65]

Pectin has significant effect on cholesterol and bile acid metabolism in rats. Male Wistar rats fed on pectin-supplemented (7 g/100 g) diet for 4 weeks resulted in lower serum and liver cholesterol concentrations as compared to fiber free diet.[66] Pectin and gums bind with bile acids, and decrease cholesterol levels and fat absorption.[67] Guar gum supplementation in diet of type-I diabetic patients, decreased serum total cholesterol level by 21%.[45] It is concluded from our article that the chapaties prepared from composite samples lowered plasma cholesterol from 9–18%, which would be helpful for hypercholesterolemic individuals.

Triglycerides

Means for triglycerides in different groups of rats explored significant variations among different groups of rats fed on various diets prepared from composite flours (Table 6). Maximum triglycerides concentration was found to be 79.60 mg/dL in control group followed by 77.40 mg/dL in T₁. In case of rats fed on guar gum 3% (T₂), triglyceride concentration of 66.60 mg/dL was observed, which showed highly significant differences as compared to rats fed on control. Percent decrease of triglycerides in different groups of rats showed a maximum reduction (16.33 %) in T₂ followed by T₃ (12.31%) and T₁ (2.76%), with reference to control (Fig. 3).

Figure 3 Percent decrease in plasma triglyceride of diabetic rats with reference to control.

The present research findings are supported by the earlier studies in which 22% decrease in triglycerides was observed in guinea pigs by feeding soluble fiber diet containing 2.5g/100g of guar gum, 5g /100g of psyllium, and 5g/ 100g of pectin diet.[68] Triglyceride level in STZ diabetic rats were 98 ± 32.[18] Water-soluble fibers, such as pectin found in fruit, guar gum and other gums found in beans, and beta-glucan found in oats, are particularly helpful in lowering triglycerides.[62] In several clinical trials, it has been investigated that diets high in fiber have reduced TG levels.[69] Guar gum fed diets (10% and 20%) decrease blood triglycerides significantly.[44] It is concluded from the present findings that use of chapaties prepared preferably from the selected treatments is a good attempt to lower the triglycerides level.

CONCLUSION

The number of hyperglycemic and hypercholesterolemic individuals is increasing rapidly and demands some strategic measures for their control. Diet management is an effective tool to control the plasma hyperglycemia and hypercholesterolemia. In order to improve the health of the people, especially diabetic and heart patients, it is imperative to mix wheat flour with suitable edible materials containing dietary fiber, which lower the serum cholesterol and glycemic index. Guar gum and pectin are vital to augment the dietary fiber in the diet of vulnerable group. When a staple food that is consumed regularly by the masses is used as a vehicle, high population coverage can easily be achieved. From the present research article, we conclude that blood glucose, cholesterol, and triglyceride levels can effectively be controlled by introducing these fiber sources. Further research is also needed to explore some new dietary fiber sources with special reference to soluble fibers. The addition of guar gum and pectin increases the dietary fiber intake for chapaties. Community-based research trials are needed to demonstrate a significant health effect.