Stability of Nano Encapsulated Rice Bran Derived Bioactive Pentapeptide inApple Juice

Abstract

Cereal grains and their components derived Bioactive compounds such as rice bran can promote health and can be derived from Rice bran contains 12–20% protein and could be a good source for extracting bioactive peptides. A pentapeptide with a sequence of amino acids Glu-Gln-Arg-Pro-Arg (EQRPR) has been prepared from heat stabilized defatted rice bran (HDRB) and has demonstrated anti-cancer properties in-vitro. This bioactive pentapeptide can thus be used as a nutraceutical by incorporating it into a suitable food system. Fruit juices can be vehicles to incorporate this pentapeptide. Fruit juices contribute to about 60% of the consumed beverages in the U.S. However, the stability of the pentapeptide in beverages can be a problem due to possible interactions with other components. Nano-encapsulation is a novel and promising technique that can be used to deliver bioactive ingredients into food systems. This study involves the use of a nano-encapsulating technique to protect the bioactive pentapeptide, incorporating the encapsulated pentapeptide into apple juice (model system), and testing for the stability of the peptide. The null hypothesis of the study: The Nano-encapsulated pentapeptide shall degrade over time when incorporated in apple juice and the alternate hypothesis that the Nano-encapsulated pentapeptide incorporated apple juice shall be stable over a storage period of 6 months or more. The specific objectives of this research were to: (1) prepare nanoparticles using polylactic-co-glycolic acid (PLGA) to encapsulate the rice bran pentapeptide, (2) incorporate the encapsulated pentapeptide into apple juice, (3) evaluate the stability of incorporated pentapeptide at 4°C for 6 months. Nanoparticles that can deliver three different concentrations (200/ 400/ and 600 µg/ml) of pentapeptide were prepared, and the particle size were measured using a laser particle size analyzer. Apple juice containing nanoparticles (loaded with pentapeptide) was ultra-centrifuged to separate nanoparticles, and the supernatant was analyzed by high performance liquid chromatography (HPLC) C18 column reverse phase (RP) to test the stability of pentapeptide. Physical properties of the apple juice were studied which included the evaluation of color, microbial count total, acidity (pH), and soluble solid (TSS) during storage period of 60 days. A particle size ranging from 81 to 83 nm was observed, and the results indicated that there were no significant changes in the size over the storage period (0–60 days). There was no microbial growth observed in the prepared apple juice samples. Total Soluble Solids content was 11.0 °Brix for the controls and 31.0 °Brix for the Nano-encapsulated pentapeptide. The stability of pentapeptide at prepared concentrations: 200, 400 and 600µg/ml in water at pH of 3.7 at 0th day was: 200ug/mL - 87%, 400ug/mL - 97%, 600ug/mL - 91%) and in apple juice was: 200ug/mL - 96%, 400ug/mL - 98%, 600ug/mL - 94 %. The stability of pentapeptide at 60th day in water was: 200ug/mL - 41%, 400ug/mL - 60%, 600ug/mL - 55%, and in apple juice was: 200ug/mL - 60%, 400ug/mL - 67%, 600ug/mL - 59%. The Nano-encapsulated pentapeptide in water at pH of 3.7 and apple juice was stable over the storage period of 60 days, which implies that the nanoparticles were effective in protecting the bioactive pentapeptide in the acidic environment of apple juice. The PLGA nanoparticles showed a remarkable effect in protecting and stabilizing the bioactive compounds (pentapeptide) during the shelf life at 4°C. Polylactic-co-glycolic acid nanoparticles can thus be a promising carrier for the bioactive pentapeptide when incorporated into a juice medium.