Formulation and Characterization of Calcium-Fortified Jelly and Its Proximate Composition and Sensory Analysis.

Calcium is a dynamic mineral. Recent discoveries designate that low intake of calcium generates deficiencies and path to other diseases. Food fortification could play a key role to overcome this problem. To cope with this deficiency problem, jellies were formulated with food-grade calcium salts and chicken eggshell powder. In the present study, three different concentrations of calcium salts, as well as eggshell powder were used to formulate jellies. The results of the sensory evaluation indicated that the two jelly products (A&D) in the current study were suitable for consumers. Results of Atomic Absorption Spectrophotometer revealed Jelly A and jelly D had 151±0.05 ppm and 133±0.06 ppm calcium concentration, respectively. Proximate analysis of Jelly A showed that it has 6.0±0.01% ash, 9.2±0.1% moisture, 0.4±0.01 g crude protein, 82.79±0.001 g crude fiber, and 0.61±0.001 g crude fat, while the jelly D that was made with chicken eggshell powder exhibited 6.0±0.01% ash, 10.1±0.1% moisture, 0.5±0.01 g protein, 84.54±0.01 g crude fiber and 1.61±0.01 g crude fat. Therefore, these two jelly A & D were greatly appreciated among other attributes. In spite of naturally available calcium-rich sources, calcium-fortified jellies can be consumed by individuals who are incapable to take sufficient calcium from their diet.

consumption that can cause osteoporosis. There should be sufficient calcium in the diet of children before adulthood so that the bones are well firmed and adults do not develop osteoporosis 5 . Hypocalcemia is a condition in which there is a low calcium level in blood 6 . Hypocalcemia can present as an asymptomatic condition, or as a severe, life-threatening condition. This deficiency may be due to a variety of factors, including poor calcium intake over a while, treatments that may decrease calcium absorbance, dietary intolerance to foods rich in calcium, and many genetic factors also 7 .
Food fortification could play a key role to overcome this deficiency problem. Milk and other dairy food items such as yogurt, mozzarella & cheese, etc. are acknowledged as an ideal source of calcium in diet 8 . There are calcium choices of nondairy products such as broccoli, spinach, sesame seed, & almonds for those who do not consume dairy products. Despite these naturally available calciumrich sources, fortified foods are a very useful mode of helping those individuals who are incapable to consume satisfactory calcium from alimentary sources. From this perspective, food fortification is a good alternative approach 9 . Food fortification is proven a cost-effective and high-impact solution to micronutrients deficiencies. Staple food can be fortified with micronutrient, including iron, calcium, magnesium, folic acid, vitamin A, iodine, and others, without affecting taste and color 10 .
Several studies about the fortification of food with minerals have been published in recent years. Jelly was chosen for fortification because of its tempting nature and extensive consumption among children. A few countries added calcium to flour, but it is more usually added to other food. Calcium salts Calcium carbonate, Calcium gluconate were used to fortify the jellies 11 . On the other hand, the chicken eggshell powder was also used to fortify the jellies. Both types of jellies were formed to check their properties after preparation and during storage time too. In the end, the results were compared with each other to select the jelly with good perception. Food grade salts were used for jelly fortification.
Hidden hunger 12 that is, the second name of micronutrient malnutrition 13 , is affecting most countries of Central and South Asia 14 . The main objective behind the formulation of this fortified jelly was to produce food that can be a vehicle to provide sufficient levels of calcium and help offset this deficiency problem among the local population. Ending hunger regarding calcium is the aim of this research. This product will also help to understand the benefits of fortification later and more working on this idea will help to produce a high-quality calcium-fortified jelly.

Materials and Methods
Food-grade calcium salts, i.e., calcium carbonate, calcium gluconate, and brown chicken eggshells, were used for fortification. Calcium salts, brown chicken eggshells, and other ingredients gelatin, sugar, and citric acid were obtained from a local market in Lahore. Food flavor and color additives were purchased from Abkari market, Lahore. For the determination of mineral Ca , aqua regia was utilized for the digestion of samples that were prepared in the laboratory. For crude protein determination, digestion mixture CuSO 4 :K 2 SO 4 was also being used in a 1:8 ratio.

Preparation of calcium-forti ed jelly with calcium salt
Calcium-fortified jellies were prepared with three different concentrations of calcium salts. In this formation of calcium-fortified jellies, distilled water was used and brought to the boiling 15 . The mixture containing salts, gelatin, sugar, and citric acid was added in boiling water and stirred well for 3-4 minutes with a medium flame. Finally, food flavor and color were added as per taste 16 . Then the mixture solution was poured into a jelly mold and cooled at ambient temperature and stored in sterilized boxes 17 . The basic Formulations of calcium-fortified jellies are shown in Table 1. All three jelly formulations were subjected to sensory analysis and proximate analysis 18 .

Preparation of eggshell powder
The eggshell powder was formed by passing the brown chicken eggshells through three stages, i.e., washing, boiling, and drying. Eggshells were thoroughly washed twice under running tap water for the elimination of any infectious agent presence 19 . Then, the eggshells were boiled for 20 minutes to inactive any microbial contamination 20 . After boiling, the eggshells were dried in an oven and grounded in mortar & pestle to form a coarse powder 21 .

Preparation of calcium-forti ed jelly with chicken egg-
shell powder Three different concentrations of brown chicken eggshell powder were used in the production of jellies. In the preparation of these jellies, the same above stated method was applied. The distilled water was brought to boiling, after that mixture of eggshell powder, gelatin, sugar, and citric acid was added in the boiling water and stirred for about All fortified jelly samples were analyzed by using standard analytical methods. pH is an indication of the acidity of the solution. The pH value of 5-6 is considered optimal for gelling and taste reasons. The pH of the prepared jellies was measured by using a digital pH meter at room temperature 25 5 . For this purpose, 0.5 g of jelly was dispersed in 50 mL of distilled water to make a 1 solution, and then pH was noted 23 . The acid-base titration method was also used to calculate the acidity of jelly samples.

Brix value and refractive index
A refractometer was used to check the refractive index and Brix value of jelly samples. Brix is the sugar concentration in the product. Measurements should be made at 20 to get an accurate value.

Mineral Ca determination
All the fortified jellies were analyzed after preparation. Several methods are available for mineral content determination 24 . In the present study, the mineral content of jelly was determined by Atomic Absorption Spectrometry according to the standard methods. jellies samples 5 g were digested in aqua regia HCl:HNO 3 , 3:1 by heating and then diluted with double distilled water up to 25 ml . After digestion and filtration, mineral Ca was determined using Atomic Absorption Spectrophotometer AAS . The filtered solution was vaporized and atomized in the flame. The atoms then absorbed the light at a characteristic wavelength. The source of the light was a hollow cathode lamp, which was made up of the same element to be analyzed. The lamp produced radiation of a suitable wavelength between 190 and 320 nm, which while passing through the flame was absorbed by the free atoms of the sample. The absorbed energy was measured by a photodetector readout system. The amount of energy absorbed is proportionate to the concentration of the element in sample 25 .

Sensory evaluation
To determine the acceptance of fortified jellies with calcium salts and eggshell powder, a sensorial evaluation of all six samples was carried out by five semitrained panelists. Three samples of jelly with calcium salts and the other three with eggshell powder were evaluated. Jelly produced from a ready-made gelatin mixture was used as a control. Sensory evaluation was based on a simple hedonic scale of five points, 1 extremely disliked to 5 extremely liked. The evaluation was based on five attributes: appearance, taste, aroma, color, & overall acceptability 26 .

Proximate composition of calcium-forti ed jellies
The proximate content 27 of calcium-fortified jelly samples is presented in Table 2. There was no difference in ash content of both samples A & D . Both best attributes share the same ash content of 6.0 0.01 ash. A slight increase was observed in the moisture content of sample D 10.1 0.1 fortified with eggshell powder compared to sample A 9.2 0.1 i.e., fortified with calcium salts. The decrease in moisture content of Jelly A as compared to Jelly D suggests that the reduction in the moisture content would interrupt the proliferation of micro-organisms 28 . However, the crude fat content of jelly D increased and differed significantly with the addition of eggshell powder. Jelly A had a low crude fat content 0.61 0.001 g and jelly D had a high-fat content 1.61 0.01 g. Similarly, the crude fiber content increased with the addition of eggshell powder in jelly D 84.54 0.01 g. While jelly A had less crude fibers 82.79 0.01 g compared to jelly D. This observation indicates that the sample with low dietary fiber jelly A will digest easily and will promote good health 29 . There was no significant difference in the protein content of jelly A 0.4 0.01 g and Jelly D 0.5 0.01 g .

pH and acidity of jellies
The pH of the jelly samples was found out to be between the optimum range of 4.5-5.0. In foods, acidity indicates the content of free acids and other chemical compounds. The acid-base titration method indicated that the acidity of both jelly samples was in the acceptable range of 0.1 0.006 and 0.13 5.7 pH and acidity are important pa- rameters to assess the internal quality of the processed jellies for their potential use 30 .
Results of pH and acidity are given in Table 2. The overall results of pH and acidity for the samples showed no significant differences because the fortified jellies have the same behavior as other simple jellies.

Degrees Brix and refractive index
The refractive index and degree Brix were measured by using a refractometer. Measurement was made at 20 to get an accurate value of Brix 31 . Degree Brix for any jelly item should be 40. The fortified jellies were near to this value. Sample A has a degree Brix of 36 0.1 and sample D has 33 0.005 32 .

Mineral content of calcium-fortified jellies
The mineral content of the jelly samples was presented in Table 3. The mineral content of jellies fortified with a higher concentration of calcium salts and eggshell powder had significantly higher calcium content 33 . While samples with a low concentration of salts and eggshell powder have lower calcium content 34 . In all these samples, sample A and sample D had calcium content in a limited range that can be given in any fortified food. Jelly A and Jelly D have 151 ppm and 133 ppm respectively. While jelly C had the highest amount of calcium content 226 ppm among the jellies that formulated with calcium salts. While jelly F had the highest amount of calcium 170 ppm among other samples of eggshell powder 35 .

Sensory evaluation of calcium-forti ed jellies
The results of the sensory evaluation are summarised in Figs. 1 and 2 for jelly A and jelly D, respectively. The results showed that there was no significant difference in color and appearance of all fortified samples 36 . The rating for aroma showed that panelists placed preference to the sample with the least amount of salt and eggshell powder. Ratings for taste showed a slight difference among the samples 37 . Samples with less amount of salt and powder were preferred by the panelists. The scores of almost all the sensory attributes of fortified jellies were not different from each other Fig. 1 . The jelly samples with a higher concentration of calcium salts and the eggshell powder were bitter and crunchy in taste compared to other samples with low calcium concentration Fig. 2 . In all, the result of overall acceptability revealed that jelly A and jelly D had the best rating 38 .

Conclusion
It can be concluded from the present findings that the better quality of calcium-fortified jelly can be prepared by using calcium carbonate 0.3 g and calcium gluconate 0.24 g along with 50 g sugar and 25 g gelatin. Jelly sample A was far better than sample B & C in chemical analysis and sensory evaluation. Chicken eggshell calcium-fortified jelly sample D was of better quality among the other two eggshell samples with 0.3 g of eggshell powder. This enriched food was prepared for consumption by humans to fulfill their daily calcium needs. To determine the effect and differences between jellies fortified with calcium salts and fortified with eggshell powder, different analysis techniques were carried out. Fortification resulted in increased proximate and mineral content of fortified jellies. In general terms, the fortified samples were sensory well accepted, it was witnessed that the best scores were achieved by sample A and sample D of all aspects. This study shows that fortification could be of great advantage to developing countries, where many cannot afford highly enriched foods because of their cost. This food could help them in reducing calcium deficiencies.   Formulation and Characterization of Calcium-Fortified Jelly