PhD Dissertation Proposal Defense by Maureen Otieno - "Optimizing the Antioxidant Properties of Cranberry Fruit and Pomace Extracts from Massachusetts"
Abstract:
The North American cranberry (Vaccinium macrocarpon) has garnered attention as a functional food due to its potential health benefits associated with its phytochemical content. The identified phytochemicals include phenolic acids (such as hydroxybenzoate and hydroxycinnamic acid), flavonoids (including type A proanthocyanidins, anthocyanins, flavonols, and flavones), and terpenes. Cranberry pomace, the solid byproduct generated during the production of cranberry juice, is primarily composed of skin, pulp residues, stems, and seeds. This pomace is a rich, natural source of pectin, fiber, and various phytochemicals. Cranberries and their derivatives have been linked to a decreased risk of several health conditions, including urinary tract infections, cardiovascular diseases, and certain cancers. Additionally, they exhibit notable antioxidant and anti-inflammatory properties.
This study aims to investigate the notable antioxidant potential of polyphenols derived from Massachusetts cranberry fruit and pomace. The first aim will assess the influence of different extraction solvents on the recovery and composition of phenolic compounds from cranberry pomace and their corresponding antioxidant activities. Antioxidant assays will be employed, including the measurement of total phenolic content (TPC) using Folin-Ciocalteu reagent, as well as DPPH (2,2-diphenyl-1-picrylhydrazyl), ABTS (2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid)), and ferric reducing antioxidant power (FRAP) tests. The proanthocyanidins will be characterized using p-dimethylaminocinnamaldehyde (DMAC), while high-performance liquid chromatography (HPLC) integrated with ultraviolet-visible (UV/Vis) detection, alongside ultra-high performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC–Q‐TOF–MS), will be utilized for the identification, isolation, and quantification of polyphenols and triterpenoids. Extracts yielding the highest concentrations will be further characterized and isolated for detailed analysis of specific bioactive compounds. Based on the antioxidant assays conducted, the solvent combination of acetone/methanol/water/formic acid (40/40/19/1) clearly demonstrated optimal antioxidant activities and phenolic content. Therefore, this extraction solvent will be utilized to analyze the fruit cultivars effectively.
The second aim will investigate various Massachusetts cranberry fruit cultivars' phenolic content and bioactive potential. By examining these distinct cultivars, we seek to deepen our understanding of cranberries as a powerful reservoir of health-promoting compounds. This study will not only illuminate the remarkable nutritional benefits of these fruits but also provide invaluable guidance to growers who seek to optimize the quality and market value of their crops. The fruit cultivars were analyzed in two phases. In the first phase, we examined three cultivars harvested in September 2023 at UMass Cranberry Station: “Early Black,” a native variety; “Stevens,” a first-generation hybrid; and “Mullica Queen,” a high-yield, low-rot second-generation hybrid. Among these, “Early Black” and “Mullica Queen” exhibited the best yields and phenolic contents, while “Stevens” performed less favorably. The second phase, which involves a comparative analysis of antioxidants in fourteen second and third-generation hybrid cranberry cultivars—including high-yield and low-rot varieties—harvested in October 2024, is currently underway.
Selected fruit and pomace extracts will be further investigated for antioxidant properties in both in vitro and in vivo models. Aim three will assess the effects of these cranberry polyphenols on the proliferation of HT-29 colon cancer cells. This will be achieved using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) and Alamar Blue cytotoxicity assays. Based on the results of these assays, the extracts from the three fruit cultivars and pomace showed inhibition of cell proliferation only at higher concentrations. We will evaluate oxidative stress levels to ascertain whether polyphenols act primarily as antioxidants or pro-oxidant agents, and how this impacts their anticancer properties. Intracellular reactive oxygen species (ROS) activity will be analyzed with DCFH-DA (2′− 7′- dichlorodihydrofluorescein diacetate) staining, the production of superoxide anions (O2•−) will be evaluated with the nitroblue tetrazolium (NBT) assay, and nitric oxide (NO) will be measured utilizing a nitrate/nitrite assay kit. Antioxidant enzymes, including superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GP), will be quantified using commercial assay kits, while lipid peroxidation will be assessed via the thiobarbituric acid reactive substances (TBARS) assay.
The fourth aim will investigate the protective effects of cranberry fruit polyphenol extract against oxidative stress induced by menadione in a zebrafish (Danio rerio) model. This will include examining morphological changes using fluorescent microscopy, assessing oxidative stress levels through biochemical markers, and conducting analyses of protein and gene expression. Findings from oxidative stress analysis on zebrafish indicated that embryos could survive at a menadione concentration of 1 µM, albeit with some deformities. In contrast, the EB DS (“Early Black” desugard) extracts at concentrations of 10 µg/mL, 5 µg/mL, and 1 µg/mL demonstrated no toxicity to the embryos after 96 hours.
In summary, Our proposed research seeks to investigate the effects of different Massachusetts cranberry cultivars on antioxidant activities and to elucidate the mechanisms behind the inhibition of proliferation and oxidative stress. This work aims to enhance the understanding and application of cranberries as rich sources of health-promoting compounds, providing valuable insights to growers looking to optimize crop quality and value. Additionally, this study will deepen our comprehension of how cranberry fruit extracts influence cellular viability and the crucial role of reactive oxygen species (ROS) in the anticancer properties of polyphenols. It will also underscore the potential of cranberry polyphenols as a natural intervention to combat oxidative challenges during early development. Therefore, this research is vital for facilitating the discovery and development of novel pharmacological agents.
SENG-311
Rachel White
508-999-8587
rwhite@umassd.edu