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United States Department of Agriculture

Agricultural Research Service


Location: Citrus and Other Subtropical Products Research

2011 Annual Report

1a. Objectives (from AD-416)
1. Identify and characterize biologically active compounds in citrus peel and associated processing byproducts for potential as value-added products to promote human health. a) Advance the discovery, isolation, and structural characterization of compounds from peel, molasses, and ethanol conversion residues and from lipid-soluble constituents of peel oil residues for biotesting purposes. b) Discover new beneficial pharmacological actions of citrus byproduct compounds, validate these biological actions in animals, and characterize the associated modes of action, pharmacokinetics and bioavailability. 2. Identify citrus processing waste compounds that can be used as value-added products to control major citrus and other plant diseases. a) Discover anti-microbial compounds from citrus processing waste. b) Characterize the influences of citrus byproduct compounds on pathogen virulence and microbial ecology. 3. Develop economical recovery methods for biologically active classes of compounds in citrus processing waste. 4. Develop value-added food materials from polysaccharide constituents of citrus byproduct waste streams.

1b. Approach (from AD-416)
Develop new value-added uses of citrus processing byproducts by exploiting the bioactive constituents (secondary metabolites) and specialty food fibers of these citrus processing materials. Fractions enriched in specific phenolic compounds will be isolated and evaluated as potential value-add materials for food and health applications. New, untested compounds, novel compositions with other citrus compounds, and new biological applications will be pursued. To achieve this, research will be directed towards 1) discovering new beneficial biological actions of citrus byproduct compounds, 2) validating these biological actions in vivo, 3) characterizing modes of action, pharmacokinetics, and bioavailability of bioactive citrus byproduct derived materials, and 4) producing specialty fibers to fill a wide range of high value food applications. Approaches to discover new biological actions will extend to studies of the influences of citrus processing waste compounds on microbial pathogen virulence and ecology, with an aim towards controlling microbial pathogens in citrus production. Broad profiles of compounds will be tested against plant pathogens, with an emphasis on important citrus microbial pests. Another direction to this work is the development of new syntheses and analytical methods for the successful production of pectin materials possessing wide ranges in functionalities to fill high-value food and pharmaceutical applications. Emphasis will be placed on the production of these specialty fibers through selected actions of known hydrolytic enzymes and through site specific chemical modifications. Critical to any future commercialization of value-added citrus byproduct materials is the development of economical recoveries of the modified polysaccharide fibers and secondary metabolites. Effective fractionations of byproduct streams into specific classes of secondary metabolite compounds and structural polysaccharides will be developed.

3. Progress Report
Important progress was made in the isolation and identification of metabolites of citrus peel compounds, polymethoxylated flavons (PMFs), in a pharmacokinetic investigation in rats. Pharmacokinetic time courses of distinct profiles of metabolites of citrus peel PMFs were also measured. The PMFs are currently sold as food supplements, and their efficacy is dependent on their metabolism and oral bioavailability. This rat study will contribute to improved use of the PMFs, hence increasing the value of these citrus byproduct compounds. Progress was also made in enhancing the recovery of PMFs from peel waste, initially treated for ethanol production. The PMFs, are present in orange processing waste steams at approximately 0.02–0.03% of the wet weight. Research showed that after hydrolysis and fermentation of available sugars to ethanol for use as a biofuel, the available PMF content in the residue for isolation purposes increased by 150%. It was found that 1 1iter of the food grade resin (XAD 16) used for citrus debittering could remove the PMFs from approximately 200 Liters of hydrolyzed and clarified orange peel extract. However, with the low amounts (0.02–0.05%) of these components present and the need to clarify the orange processing waste extracts prior to treatment, recovery is unlikely to be economically viable. Progress was also made in understanding the chemical mechanisms of grapefruit-drug interactions caused by grapefruit juice phytochemicals. A new fungal metabolite of an important grapefruit furanocoumarin was detected and structurally characterized, and was found to lack CYP3A4 inhibitory activity. CYP3A4 inhibitory activity for most grapefruit furanocoumarins is responsible for the grapefruit/drug interactions that occur in humans consuming both grapefruit and certain prescription medications. The ability to modify these grapefruit compounds by fungi may lead to new technologies to eliminate these deleterious drug interactions in humans. Progress was also made in two separate projects to detect biomarker compounds for citrus canker and HLB diseases. Key differences were detected in the profiles of compounds of healthy versus Huanglongbing (HLB) affected leaves. This will aid in early detection of HLB and contribute to an understanding of the progression of this disease in citrus plant tissues, and to discover mechanisms by which citrus trees cope with the canker-causing bacterium. Work also continued with evaluating natural compounds for their antimicrobial activity against Xanthomonas citri and problematic decay pathogens. Progress was also made pertaining to value-added chemical research into the polysaccharide structures of citrus byproducts. Pectin methylesterase present in a commercial papaya enzyme extract was used to demethylate a model pectin molecule. The resulting modifications to the pectin nanostructure have been characterized. The results indicate that reaction conditions affect the nanostructural motifs which may lead to the development of novel pectin products. This project replaced project 6621-41000-014-00D which was a bridging project which continued work from project 6621-41000-012-00D.

4. Accomplishments
1. Chemical responses in grapefruit to bacterial plant pathogens. The bacterial disease in citrus, termed citrus canker, is devastating grapefruit and orange production in Florida for fresh fruit markets. Among the many different citrus cultivars there is a wide range in susceptibility to this disease, yet the chemical basis for this is poorly known. Extensive analyses of the susceptible grapefruit have provided key information about the changes in the chemical environments accompanying infection by the canker bacterium. Important marker compounds for canker infection have been found. These marker compounds may be plant defense compounds which may be useful in combating this disease. These discoveries will provide a baseline by which researchers can compare susceptible cultivars (i.e. grapefruit) against less susceptible cultivars, and gain an understanding of the chemical basis of tolerance to this pathogen. This will enable researchers to develop technologies to combat and mitigate this disease, and restore profitability to the Florida fresh fruit market.

Review Publications
Cameron, R.G., Luzio, G.A., Vasu, P., Savary, B.J., Williams, M.A.K. 2011. Enzymatic modification of a model homogalacturonan with the thermally tolerant pectin methylesterase from citrus: I. Nanostructural characterization, enzyme mode of action and effect of pH. Journal of Agricultural and Food Chemistry. 59:2717-2724.

Manthey, J.A., Cesar, T.B., Jackson, E., Mertens-Talcott, S. 2011. Pharmacokinetic study of nobiletin and tangeretin in rat serum by high-performance liquid chromatography-electrospray ionization-mass spectrometry. Journal of Agricultural and Food Chemistry. 59:145-151.

Manthey, J.A., Perkins-Viazie, P. 2009. Influences of harvest date and location on the levels of ß-carotene, ascorbic acid, total phenols, in vitro antioxidant capacity, and phenolic profiles of five commercial varieties of mango (Mangifera indica L.). Journal of Agricultural and Food Chemistry. 57:10825-10830.

Last Modified: 06/26/2017
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