2012 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.
This project is aimed at developing new products from citrus bioactive constituents and specialty food fibers derived from citrus processing waste streams. As part of these investigations, studies were performed to discover naturally occurring citrus plant response compounds capable of limiting the deleterious actions of microbial pathogens.
Objective 1: Improvements were also made in analytical techniques for the study of citrus by-product compounds. The use of fluorescence detection was explored for three major classes of bioactive constituents. Fluorescence detection allows for the detection of single peaks that were normally unresolved in ultraviolet wavelength monitored, highly overlapping chromatograms. The use of fluorescence detection enhanced the detection of a number of newly observed compounds in citrus, the identities of which are currently under investigation.
Objective 2: Studies on the biological and chemical ecology of the canker lesion were continued as well as screens for natural antimicrobials. Recent data show that when the citrus-causing bacterium is ‘stressed’ by treatments (e.g. exposure to antimicrobials) the colony changes. Whereas the Xanthomonas citri subsp. citri (Xcc) produces a yellow gummy substance (xanthin), the ‘stressed’ colony produces a companion cell that is white to opaque. This organism becomes the dominant cell form rather than the canker bacterium. Depending on the treatment (i.e. screens for antimicrobials) there are various ratios of the two cell types which may play a role in the success of the canker bacterium as an epiphyte. When comparing cell population of young and old cankers, young cankers are predominantly Xcc while older cankers are a mix of these and the opaque cells. Data show that the chemical make-up of the lesion changes as well as the canker ages. Data from our screens show that Xcc may be able to process toxic compounds in such a way that the bacterial cells are eventually able to overcome the toxicity and begin to grow.
Objective 4: Progress was 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 (i.e., pH and enzyme/substrate ratios) affect the introduced nanostructural motifs.
A predictive model for the relationship between pectin nanostructure and rheological properties. ARS Researchers at Ft. Pierce, FL have developed a predictive model for the relationship between pectin nanostructure and rheological properties. Pectin functionality is dependent on the distribution of charges along its polymeric backbone. Random versus ordered charge distribution affects pectin’s functionality and variations in the amount or topographical order of electrical charge within the polymer chain also affect rheology. We have modified pectin charge distributions using enzymes or chemical processes and developed technology to statistically describe the introduced nanostructural modifications. Rheological testing of the engineered pectins have enabled us to develop a predictive model relating nanostructural parameters to rheological properties. This accomplishment has the potential to allow food processors greater control over texture, gel formation and interaction of pectin with other food components related to flavor and aroma.
Vasu, P., Savary, B.J., Cameron, R.G. 2012. Purification and characterization of a papaya (Carica papaya L.) pectin methylesterase isolated from a commercial papain preparation. Food Chemistry. 133:366-372.
Pérez, C.D., Fissore, E.N., Gerschenson, L.N., Cameron, R.G., Rojas, A.M. 2012. Hydrolytic and oxidate stability of L-(+) -ascorbic acid supported in pectin films: Influence of the macromolecular structure and calcium presence. Journal of Agricultural and Food Chemistry. 60:5414-5422.
Manthey,J.A. 2012. Potential value-added co-products from citrus fruit processing. In: Bergeron,C., Carrier,D.J., Ramaswamy,S., editors. Biorefinery Co-Products. Hoboken, NJ: John Wiley & Sons, Ltd. p. 153-178.
Myung, K., Manthey, J.A., Narciso, J.A. 2012. Biotransformations of 6',7'-dihydroxybergamottin and 6',7'-epoxybergamottin by the citrus-pathogenic fungus Penicillium digitatum diminish cytochrome P450 3A4 inhibitory activity. Bioorganic and Medicinal Chemistry Letters. 22:2279-2282.