Report an error   

HERDIN Record #: R07-CIM-18020718010325 Submitted: 08 February 2018 Modified: 04 August 2018

The effect of storage on refrigerated temperature on the antioxidant property of Citrofortunella microcarpa (Calamansi) using 2, 2-diphenyl-1-picrylhydrazyl (DPPH) assay.

Gail Ann Bongalos,
Arvin Dela Vega,
Mark Gavin Dinsay,
Kenn Carlson Dy,
Mikaela Adeliz Gorme,
Bea Angeli  Kuizon,
Lalaine Mayol,
Esther Ann Nacita ,
Jane Marie Valmores,
Lenie May Vaporoso

See More

Introduction: Antioxidant compounds are the substances, compounds or nutrients found in food which can prevent or slow oxidative damages in the body. Antioxidants play an essential role against free radicals by neutralizing free radicals through donation by one of their electrons. Among the plant sources of antioxidant-rich compounds, citrus fruits like Citrofortunella microcarpa (Calamansi) are widely consumed by Filipinos. The antioxidant content of citrus fruits is affected by storage conditions, particularly temperature and method of preparation. Objective: This study aims to detect the decrease in the antioxidant property of Citrofotunella microcarpa (Calamansi) extracts after storage intervals of 24,48, 72 hours at 4°C (±0.5°C). It further aims to determine the absorbance values of the extracts and correlate these to the standard Vitamin C absorbance values, the extent of the decrease in absorbance of the mentioned storage intervals at 4°C (±0.5°C) of the extracts. Design and Setting/ Population: A quantitative-experimental study was used to determine the extent of absorbance of the different time intervals. The study was performed at the CIM Biochemistry and Histology Laboratory. The population included Citrofortunella microcarpa (Calamansi) fruits with green intact skin that are firm to touch. Fruits that were worm-infested, overripe (yellow to orange in color) with bruised and damaged skin and discolored with black blotches were excluded in the study. Methodology: Approximately one hundred Citrofortunella microcarpa (Calamansi) were used to collect 60 ml extract of Calamansi. The collected extract was further subdivided into 12 equal aliquots containing 5 ml each. The aliquots were divided into four groups for the 0, 24,  48 and 74-hour antioxidant activity assay. All vials except the 0-hour group were stored in a refrigerator set at 4°C (± 0.5 °C). Water was used as negative control - 60 ml was divided and stored under the same conditions as the test samples. Sodium ascorbate (vitamin C) preparation from a capsule (Fern-C), at an initial concentration of 0.25 mg/ml, was used as a positive control and a standard to derive the calibration curve. 2,2-diphenyl-1-picrylhydrazyl (DPPH) prepared by dissolving 10 mg in 250 ml of 80% ethanol, was used to determine the radical scavenging activity of each test group. Absorbance was measured at 517 nm. A quantitative decrease in antioxidant content was hypothesized to occur with increased storage duration. A line graph was made by plotting the time against the absorbance. The value reflects the change in antioxidant activity in relation to time. Results: The results showed no trend in the vitamin C standardization. The results showed that there was an increase in absorbance with increase duration of storage of calamansi extract. Negative control values from 0-72 hours had lower absorbance values than those of the test sample. Discussion: Since the expected trend of the vitamin C standardization curve was not obtained and the relation to negative control absorbance values was anomalous, the significance test results cannot be determined. The researchers postulated that different factors in the experiment could have affected the results. Conclusion: The test sample results cannot be evaluated because there was no linearity observed in the vitamin C standardization assay and the negative control values does not conform to the principles stated in the related literature. Recommendations have been made to improve the outcomes of future studies.

Publication Type
Thesis Degree
Publication Date
April 2016
LocationLocation CodeAvailable FormatAvailability
Cebu Institute of Medicine Abstract
1. Murray, R , Bender , D , Botham, K , Kenelly, P , Rodwell, V , Weil, P , Bender , D , Botham, K , Kenelly, P , Rodwell, V , Weil, P . Harpers Illustrated Biochemistry 29th edition . McGrawHill,
2. Murray, R , Bender , D , Botham, K , Kenelly, P , Rodwell, V , Weil, P , Bender , D , Botham, K , Kenelly, P , Rodwell, V , Weil, P . Harpers Illustrated Biochemistry 29th edition . McGrawHill,
3. Kondoa , S , Katayamaa , R , Uchino, K . Antioxidant activity in meiwa kumquat as affected by environment and growing factors. Japan: Elsevier B.V. Kagoshima 895-0041, 2004.
4. Kondoa , S , Katayamaa , R , Uchino, K . Antioxidant activity in meiwa kumquat as affected by environment and growing factors. Japan: Elsevier B.V. Kagoshima 895-0041, 2004.
5. Kumar, V , Lemos, M , Sharma , M , Shriram, V . (2012, November 30). Antioxidant and DNA damage protecting activities of EulophianudaLindl. Free Radicals and Antioxidants . . Retrieved from:
6. Sen, S , Chakraborty , R , Sridhar, S , Reddy, R Y. S., De, B . (2009, November 30). FREE RADICALS, ANTIOXIDANTS, DISEASES AND PHYTOMEDICINES: CURRENT STATUS AND FUTURE PROSPECT. . Retrieved from:
7. Halliwell, Barry . "Free Radicals and antioxidants " Nutrition Reviews 70(5): 257-265, May 2010
8. Valko, Marian , Leibfritz, Dieter , Moncol , Jan , Cronin, Mark T.D., Mazur , Milan , Telser, Joshua . "Free radicals and antioxidants in normal physiological functions and human disease." The International Journal of Biochemistry and Cell Biology 39(1): 44-84, 2007
9. Kumaraswamy, M.V , Satish, S . "Antioxidant and anti-Lipoxygenase Activity of Thespesia lampas Dalz nad Gibs" Advances in Biology Research 2(3-4): 56-59, 2008
10. USDA-ARS, . (2013, November 30). National Genetic Resources Program . Germplasm Resources Information Network- (GRIN). . Retrieved from: [URL:] cited
11. Mandadi, K , Ramirez, M , Jayaprakasha, GK , Faraji, B , Lihono, M , Deyhim, F , Patil, BS . "Citrus bioactive compounds improve bone quality and plasma antioxidant activity in orchidectomized rats" Phytomedicine 16(6-7): 513-20, 2008. Retrieved from: http://doi:10.1016/j.phymed.2008.09.0001
12. Rekha, C , Poornima, G , Manasa, M , Abhipsa, V , Pavthira, D , Kumar, V , Kekuda, P . "Ascorboc acid, total phenol content, and antioxidant activity of fresh juices of four ripe and unripe citrus fruits " Chemical Science Transactions Journals 1(2): 303-310, 2012
13. Swaran, JSF . "Structural, chemical and biological aspects of antioxidants for strategies against metal and metalloid exposure" Oxidative Medicine and Cellular Longevity Journals 2(4): 191-206, 2009
14. Nakajima , VM , Macedo , GA , Macedo , JA , Macedo , GA , Macedo , JA . " Citrus bioactive phenolics:Role in the obesity" Citrus bioactive phenolics 59(2): 1205-1212, 2014
15. Gattuso , G , Barreca , D , Garguilla , C , Leuzzi, U , Caristi, C . "Flavonoid composition of Citrus juice" Molecules 12(8): 1641-1673, 2007
16. Manners , GD . "Citrus limonoids: analysis, bioactivity, and biomedical prospects. " Journal of Agricultural and Food Chemistry 55(21): 8285-94, 2007. Retrieved from: http://DOI: 10.1021/jf071797h
17. Igwe , OU . "Quantitive Estimation of Ascorbic Acid Levels in Citrus Fruits at Variable Temperatures and Physicochemical Properties" International Journal of Chemical and Biological Sciences 5, 67-71, 2014
18. Kumar, R , Vijay, S , Khan, N . "Comparative Nutritional Analysis and Antioxidant Activity of Fruit juices of some Citrus spp." Octa Journal of Biosciences 1(1): 44-53,
19. Molyneux , P . "The use of stable free radical DPPH for estimating antioxidant activity" Songklanakarin Journal of Science and Technology 23(1): 211-219, 2004
20. Garcia, E . "Antioxidant Activity by DPPH Assay of Potential Solutions to be Applied on Bleached Teeth" Brazilian Dental Journal 23(1): 22-27, 2012
21. Syamaladevi, RM , Andrews , PK , Davies , NM , Walters, T , Sablani, SS . "Storage effects on anthocyanins, phenolics and antioxidant activity of thermally processed conventional and organic blueberries" Journal of the Science of Food and Agriculture 15(4): 916-24, 2011. Retrieved from: http://10.1002/jsfa.4670.Epub2011 Oct 3
22. Paolo, R , Marisol , LB , Paolo, P , Nicolina, T . "Effect of cold storage ob vitamin C, Phenolics and antioxidant activity of five orange genotypes [ Citrus sinenis (L.) Osbeck)" Postharvest Biology and Technology Journal 49(3): 348-354, 2008
23. Jasenka , PZ , Dunja, S . "Antioxidant stability of small fruits in postharvest storage at room and refrigerator temperatures" Food Research International Journal 44(1): 345, January 2011
24. Nicklisch, SCT . "Optimized DPPH assay in a detergent-based buffer system for measuring antioxidant activity of proteins " Methods X Journal 1, 233-238, 2014

Copyright © One Window Project 2020. All rights reserved.