Introduction

Malnutrition and chronic diseases are a global issue, especially in developing countries and it contributes to mortality rates. Malnutrition has become an inhibition factor in the achievement of Millennium Development Goals 2015. The high prevalence of anemia caused by iron deficiency has become pregnant women, infants, and children under 2 years of age,  preschool children, school-age children, adolescents, and non-pregnant women¹. In Indonesia, malnutrition has been a public health issue with a wasting prevalence of 13% and stunting 36%, and anemia in the moderate category ². Low nutrient intake to be a significant determinant factor as a cause of micronutrient deficiencies, including iron deficiency ³.

Biofortification of micronutrient in rice (Oryza sativa L.), maize, and wheat was believed to be a sustainable strategy to overcome iron and zinc deficiency in humans ⁴. Maize grain has been the staple food of the world community after rice and wheat, including in Mexico, Sahara-Africa, and Indonesia^5-7. Maize biofortified iron has not been widely applied, and the interventions needed in the future to improve its quality and overcome micronutrient deficiencies ⁸. Previously studied show that biofortification through Pseudomonas putida IFO 14796 intervention has improved iron content in maize grain up to 18.79% ⁹, and intake 10% of the iron biofortified maize (IBM) was able to improve the erythrocyte formation in Wistar rats anemia¹⁰.

Therefore, to observe the effect of maize biofortified iron on the improvement of the body weigh also needs to be observed how it affects body weight. Wistar rats (Rattus norvegicus) were widely used in laboratory studies as an animal model. They are feasible enough to mimic all aspects of human abnormalities, develop all the main signs of metabolic syndrome, obesity, diabetes, dyslipidemia, hypertension, liver disorders, and kidney dysfunction as well as micronutrient deficiencies¹¹. This research aimed to assess the effect of IBM on the bodyweight of Wistar rat anemia.

Materials and Methods

Location and Time

The experiment was undertaken for 21 days from December 17^th, 2019 to January 5^th, 2020 in the center for an integrated laboratory of Dayanu Ikhsanuddin University, Baubau City, Southeast Sulawesi-Indonesia.

Experiment Materials

Female Wistar was accessed from Bandung, Indonesia (Certificate Veterine: No. 524.3/3873-Dispangtan/2019). Female Wistar rats anemia 150-200 g induced by injection 40 mg kg^-1 2.4 dinitrophenyl hydrazine (DPNH) for 4-7 days resulted in anemia (Hb<12 mg dL^-1). Iron biofortified Maize (IBM) 10.117 mg kg^-1 was produced from the previous project through an experiment using P. putida IFO 14796.

Design Experiment

The research was carried out with the randomized complete design, consisting of four treatment groups and one control group. The treatment group contained experiment units R1 (10%), R2 (12%), R3 (14%), and R4 (16%) and Ro (10%) body weight (BW). Iron biofortified maize (IBM) as an independent variable and formation bodyweight as the dependent variable. The sample size was 25 female Wistar anemia (Rattus novergicus ) randomly placed into 5 groups. Each group consisted of 5 animals were placed separately in a 30 x 25 x 30 cm cage.   The treatment group of Wistar anemia was supplied food from iron biofortified maize (IBM) 3 times a day as well as a control group by iron non-biofortified maize (INBM), and water drinking by add labitun ¹². Also, Wistar anemia was treated well in a room with good ventilation, normal sun exposure through the window, temperature 27-30^oC, and low humidity. Bodyweight of Wistar rats was measured at the beginning and end of the study using the SF-400A “Electronic Compact Scale” digital balance. While improving body weight rate (GR) was calculated using the formula¹³:

Data Analysis

The effect of intake of the iron biofortified maize (MBI) for the bodyweight formation rate of Wistar anemia was analyzed using statistical ANOVA-ONE WAY and Fisher’s LSD (Least Significance Difference) at p<0.05.

Ethical matters

The investigation was approved and that informed consent was obtained from the Health Research Ethics Committee, Faculty of Medicine, Hasanuddin University, Protocol number UH19010037.

Results                  

The mean body weight of female Wistar rat anemia was highest in R-3 (204.50 ± 3.96 g), and the lowest in R-2 (189.40 ± 8.56 g). Variation of MBI intake resulted in a different response to Wistar rat anemia. The IBM level in R-3 (14%) can increase body weight up to 17 g for 7 days and improving rate in 0.0109% per day (2.04 gd^-1), higher than R-o (control), and others. Nevertheless, the body weight in R-1(10%) and R-2 (12%) tends to decrease (Table 1).

Table 1: Average of Body Weight in Female Wistar Anemia (% d^-1)

There was an effect of the IBM on improving bodyweight of Wistar anemia (p < 0.03). While based on Fisher’s LSD (Least Significant Different) show that body weigh in R-3 significantly different with p=0.0109<0.05 (Table 1). However, the body weight tends to increase with MBI level (Figure 2):

Table 2: Distribution of the Different Effect of IBM to the Body Weight of Wistar Rat Anemia.

Note: * means significant different at p<0.05

IBM level in R-3 for Wistar rat anemia has a significant effect on improving the bodyweight (p= 0.001) (Table 2) and tends to increase with the IBM level following the equation Y=0.005x-0.0096; R² = 0.79, where Y is body weight, and x is IBM level (Figure 1).

Figure 1: The Correlation  Intake Iron  Biofortified Maize Level With Improving Body Weight  Rate  of Wistar Anemia. Click here to view Figure

Discussion

Female Wistar anemia (Hb <12 mg / dL) has been given a single intake of iron biofortified maize (IBM) can survive up to 95%. The body weight tends to increase (0.01% d^-1) with the IBM level. Therefore, IBM has the potential to be a single source of energy to maintain the body weight of female Wistar rat anemia. Improving the body weight of rats was 2.04 g per day lower than the normal growth of rat 5 grams per day ¹⁴.  Monotonous diet from cereal has a vulnerability in micronutrient deficiency, but the product of bio-fortification positively impact to improve the nutrition in human¹⁵.

Maize grains contained carbohydrate (79.15-80.64 g / 100 g), 1756 kinds of proteins, and the fat (3.21-7.71%) ^16-18. Also, the maize grain contained some amino acid such as leucine, lysine, tryptophan, methionine, isoleucine, valine, phenylalanine, glutamic acid, serine, alanine, tyrosine, and proline^{16, 19, 20}, and mineral of Fe, Zn, Mg, Cu, pro-vitamin A, vitamin E and antioxidants in fresh corn ¹⁸. However, vitamin A degradation occurs quickly around 45% in the first 3 months, even more than 85% can be lost if the storage conditions are not supportive harvest^{21 22, 23}. Therefore, the optimal values of nutrient content and sensory acceptance of corn are relatively higher (55.0-68.5%) compared to peanuts (27.5-35.0%), and malt barley (4.0-10.0%) ²⁴. Besides, maize also contains several of phytochemical compounds in the form of phenolic, flavonoid, anthocyanin. Even corn has higher carotene, tocopherol, and oils than rice and wheat^5-7. Corn has a fiber positive effect on metabolic processes and weight control of the bodyweight ^{25, 26}. Whilst phenolic, oligosaccharides, and flavonoids are not easily digested so relatively quite a lot reach the large intestine (10.24-64.4%)^{27, 28}.  Thus, corn consumption was considered to have significant to improve the bodyweight and even to prevent chronic diseases such as cardiovascular disease, obesity, diabetes, and cancer²⁹.

Biofortification in maize with cultivars that efficiently mobilize, extract and move iron into edible plant parts, increasing iron bioavailability in staple foods is one of the solutions expected to combat and prevent community iron deficiency, especially in areas with limited resources ^{30, 31}. The basis for calculating nutritional adequacy is bodyweight, basal metabolic energy (BMR), activities, additional needs for growth, additional energy for food digestion, and factors for body composition, age, and sex. The standard for nutritional adequacy of the diet in Indonesia 2012 to covering energy; protein; fats (including n-3 and n-6), carbohydrates, water, vitamins A, D, E, K, thiamine, riboflavin, niacin, pyridoxine, folic acid, vitamin B12, pantothenic acid, biotin, choline, and vitamin C; also includes minerals such as iron, calcium, phosphorus, magnesium, sodium, potassium, iodine, zinc, copper, chromium, selenium, manganese, fluorine. fiber, choline and biotin, nitrogen and essential amino acids, fatty acids, amino acids, cholesterol, and bio-active substances in food. ³² The challenge ahead is how to bio-fortified staple foods in maize grain to increase the amount of quantity and quality micronutrients and vitamins that can meet individual metabolic needs.

The low rate of weight gain in Wistar rat anemia fed with maize bio-fortified iron illustrates that this single food does not meet the body’s metabolic needs. Likewise in humans, a monotonous diet from cereals was susceptible to micronutrient deficiencies, although the maize bio-fortified products have a positive impact to improve the nutritional health in human¹⁵. The weakness of this studied has not measured the levels of each nutrient component integrated into the maize bio-fortified iron products.  While the perfection of the body’s metabolism requires many nutrients derived from diets ³³. However, the iron biofortified maize expected to increase the content of vitamins and essential micronutrients in food simultaneously.

Conclusion

Iron biofortified maize has influenced to improve bodyweight of female Wistar rat anemia. Improving the bodyweight of Wistar anemia tends to increase with IBM level.  IBM 14% was a maximum safe level for the growth rate of Wistar rat anemia but relatively lower than the normal growth.

Acknowledgment                           

The author would like to acknowledge to all of the staff of the Cereal Research Center-Maros, Indonesia, Food and Nutrition Laboratory of Gajah Mada University, Staff of Science Hayati of Institute Technology of Bandung.

Competing Interests

This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors so no conflict of interest includes my promoter and co-promoter.

Author Contribution

All authors contributed to establishing the topic of the research and design experiment. Saifuddin Sirajuddin focused on nutrition for iron deficiency anemia; M.Natsir Djide focused on the bacterial function in biofortification, and Anwar Mallongi focused on statistical analysis.

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