RICEO TM. Water Soluble Concentrate Powder Providing Natural Vitamin B Group and Minerals

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RICEO TM Water Soluble Concentrate Powder Providing Natural Vitamin B Group and Minerals RICEO TM is a water-soluble powder extracted from defatted rice bran. It is mainly composed of carbohydrate, dietary fiber, and protein, and also contains a high proportion of minerals including potassium, magnesium, and also retains a natural vitamin B group (B1, B2 and B6). Since RICEO TM contains about 30% of a physiological substance, IP 6 (phytic acid) (Table 1), it has a strong chelating effect, powerful antioxidant and anti-browning activities resulting in various applications in food and cosmetic products. RICEO TM is totally free from GMO. Table 1: Composition of RICEO TM. (Analyzed by Japan Food Research Laboratories) Composition Carbohydrate (non-fibrous) Inositol Phytic acid Dietary fiber Protein Potassium Magnesium γ-aminobutylic acid Calcium Sodium Iron Zinc Vitamin B6 Vitamin B1 Vitamin B2 Copper /100 g RICEO TM 30.0 g 11.0 g 27.8 g 11.2 g 8.2 g 5.3 g 2.7 g 237.0 mg 121.0 mg 30.9 mg 19.1 mg 13.2 mg 10.2 mg 7.2 mg 0.6 mg 0.4 mg Functional characteristic of RICEO TM 1. Source of Mineral Magnesium (Mg), one of many minerals in RICEO TM, has many health benefits. It is essential for energy production, protein synthesis and cellular replication (e.g., DNA, RNA). Magnesium works with calcium in muscle contraction and relaxation (calcium-contraction, magnesium-relaxation) in the human body. 1

Depletion of magnesium may result from high consumption of coffee, tea, chronic diarrhea, irritable bowel, laxatives, oral contraceptives, over exercising, emotional stress and in long-term use of diuretics. High doses of supplemental zinc and high protein intake interfere with magnesium absorption. Magnesium depletion may also result in people with gastrointestinal disorders, diabetes mellitus, old age, chronic alcoholism. Mg supply action of RICEO TM is shown in Figure 1 and Table 2. In the lab experiment, Wistar rats (male, age 3 weeks) were administered with normal feed (control diet Mg content 618 mg/kg), Mglack diet (Mg content 105 mg/kg), and Mg-lack diet + 1.7% RICEO TM (Mg content 606 mg/kg). After 15 days, the rate of weight gain (Figure 1 and Table 2) and amount of Mg in serum (Table 2) were measured. The result showed that both growth rate and levels of Mg in group of rats fed with control diet and group of rats fed with Mg-lack diet + RICEO TM were not significantly different, while the group fed with Mg-lack diet shows significant lower level weight gain and level of Mg. This result suggests that, although RICEO contains about 30% of phytic acid with a chelating effect, it does not inhibit the Mg supply action of RICEO. In addition, the growth of rats was improved by the addition of RICEO. Table 2: The growth promotion and Mg supply action of RICEO TM in rat. (In-house data) Group Control diet Mg-lack diet Mg-lack diet + 1.7% RICEO TM Rate of weight gain (g) 112 ± 1.00 b* 58 ± 2.00 a 115 ± 4.00 b The amount of Mg in serum (mg/100 ml) 0.93 ± 0.12 b 0.56 ± 0.17 a 0.87 ± 0.10 b *The significant difference was observed between the different letters, a and b (P<0.05, n=6). Figure 1: The growth promotion of RICEO TM in rats comparing with control diet (blue line), Mg-lack diet (red line), and addition of RICEO TM to Mg-lack diet (green line). (In-house data) 2

2. Antioxidant Effect Antioxidation is a way of prolonging the storage of foods. The addition of antioxidants to foods would help the human body reduce the losses from oxidation disturbances related to ageing and diseases, such as atherosclerosis, cancer, and cirrhosis [1]. The TBA (thiobarbituric acid) method was used to measure the antioxidant activity of RICEO TM in terms of inhibition on lipid peroxidation. To examine the role of RICEO TM on inhibition of lipid peroxidation, RICEO TM sample was subjected to the TBA assay. The result in Figure 2A shows the antioxidant effect of RICEO TM on lipid peroxidation by comparing the TBARS (2-thiobarbituric acid reactive substances) inhibition efficiency of RICEO TM at difference concentrations of 0.05%, 0.1%, and 0.2%. The result revealed that increasing RICEO TM concentration increased TBARS inhibition efficiency. RICEO TM of 0.2% gave the highest lipid peroxidation inhibition. Figure 2: A: TBARS value in homogenized rat brain (after leaving for 120 minutes) was measured with TBA method, B: The effect of heating (90 C for 15 min) on TBARS value of RICEO TM. (In-house data) 3. The heat stability of RICEO TM The effect of heat on lipid peroxide generation was examined at 90 C for 15 min. The result demonstrated in Figure 2B shows the antioxidative stability of RICEO TM against heat. 4. Comparison of antioxidant effect of RICEO TM RICEO TM exhibits the best antioxidant effect when compared with vitamin C and sodium phytate at the same concentration level of 0.05% (Figure 3). RICEO TM possesses superoxide anion or free radical scavenging and antioxidant effects in vitro. The inhibitory effect of the RICEO TM on in vitro lipid peroxidation is similar to that of vitamin C. 3

TBARS (nmol/g tissue) Figure 3: Comparison of antioxidant effect of RICEO TM compare with vitamin C and sodium phytate. (In-house data) 5. Effect of RICEO TM on vitamin C decomposition Ascorbic acid, vitamin C is a water-soluble antioxidant that can reduce radicals from a variety of sources. Vitamin C is well-known to be oxidized in the presence of both hydrogen peroxide and transition metal ions (Fe 2+, Cu 2+ ) through the Fenton reaction [2]. Fenton reaction is the iron-salt-dependent decomposition of dihydrogen peroxide, generating the highly reactive hydroxyl radical. Addition of a reducing agent, such as ascorbate, leads to a cycle which increases the damage to biological molecules. Fe 2+ + H 2 O 2 Fe 3+ + OH + OH The study in Figure 4 shows the protective effect of RICEO TM on vitamin C decomposition in the reaction which contains 150 µm hydrogen peroxide solution, 1.5 µm cupric sulfate solution and 75 µm ascorbic acid. Figure 4 shows the decomposition of vitamin C with reaction time. Comparing with control sample (without RICEO TM ), the decomposition of vitamin C decreased in 0.01% of RICEO TM, and in 0.05% of RICEO TM almost no decomposition of vitamin C in the reaction is observed. The result suggests that RICEO TM can protect vitamin C decomposition due to its chelating effect in hydrogen peroxide and cupric sulfate solution. 4

Figure 4: Protective effect of RICEO TM on vitamin C decomposition.(in-house data) 6. Whitening effect Tyrosinase is an essential enzyme in the biosynthesis of the skin pigment melanin. As shown in Figure 5, the effect of RICEO TM on inhibition of tyrosinase activity was investigated, comparing with control sample, higher concentration of RICEO TM exhibits lower tyrosine activity. The result suggests that RICEO TM can inhibit tyrosinase activity and therefore can potentially suppress melanin generation. Figure 5: Tyrosinase activity inhibition of RICEO TM (Tyrosinase activity was assayed using dopachrome as a marker). (In-house data) 5

Safety Profile: Acute Toxicity (LD 50 ) The acute oral toxicity of RICEO TM in mice was tested in accordance with OECD guidelines for the Testing of Chemicals 401 (1987). The guidelines recommend that if compound-related mortality occurs at a dose of 2,000 mg/kg, a full study may need to be considered. In the present test, oral administration of 2,000 mg/kg of the test sample caused neither death nor abnormalities at necropsy in any mouse. Consequently, we deduced from the test that the lethal dose is higher than 2,000 mg/kg in both male and female mice. Recommended dosage RICEO TM 1 g contains nutrient equivalent to that of 76.2 g of brown rice. White rice is a staple food in most countries, as well as brown rice which the recommended amount is 300 g per day per person. The equivalent nutrients can be obtained from only 4 g of RICEO TM, and it has a balanced composition that is healthy for everyone. Table 3: Recommended dosage of RICEO TM. Applications Dosage (%) Power drink 10 12 Cookie & bread 4 5 Beverage 0.4 0.5 Applications Increasing research and development efforts have been made on RICEO TM applications. Among numerous prominent properties is its use as a whitening agent and antioxidant agent. As a food additive, RICEO TM appears pale yellow to pale brown powder, it is easy to use in combination with other food materials. In terms of processing suitability, it is used in powder form and aqueous solution, and it helps stabilize of vitamin C, exhibits antioxidant action, and anti-fatty liver action. The sour taste makes RICEO TM a perfect flavor & nutrition additive to variety of foods with predominant sweet taste such as cookies, hot-chocolate, fruits juice, jelly, candies and etc. For cosmetic ingredient, such as whitening agent, emollient, moisturizer. Properties Product : RICEO TM INCI name : ORYZA SATIVA (RICE) BRAN EXTRACT Appearance : Light brownish yellow powder, faint characteristic odor Solubility : Soluble in water 6

References [1] Stoilova, I., Jirovetz, L., Stoyanova, A., Krastanov, A., Gargova, S., and Ho, L. (2008). Antioxidant activity of the polyphenol mangiferin. EJEAFChe., 7(13), 2706-2716. [2] Deutsch, J.C., Santhosh-Kumar, C.R., Hassell, K.L., and Kolhouse, J.F. (1994). Variation in ascorbic acid oxidation routes in H 2 O 2 and cupric ion solution as determined by GC/MS. Anal. Chem., 66, 345-350. 7