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ARS Home » Southeast Area » Fort Pierce, Florida » U.S. Horticultural Research Laboratory » Citrus and Other Subtropical Products Research » Research » Research Project #429418

Research Project: Enhancing Utilization of Citrus Processing Co-products

Location: Citrus and Other Subtropical Products Research

2019 Annual Report

1. Enable, from a technological standpoint, new commercial products from pectic hydrocolloids derived from citrus processing. 2. Characterize and quantify bioactive flavonoid compounds from byproducts of citrus processing, determine their in vivo pharmacokinetics and bioavailability; and enable a new commercial delivery of bioactive flavonoids in food and feed by encapsulation with pectic hydrocolloids. 2A. Characterize and quantify bioactive flavonoid compounds from byproducts of citrus processing, determine their in vivo pharmacokinetics and bioavailability. 2B. Enable a new commercial delivery of bioactive flavonoids in food and feed by encapsulation with pectic hydrocolloids. 3. Enable a novel immunologically-based assessment of structural quality and functional properties of citrus pectin in raw and processed foods and industrial products.

Experimentation is required to determine the necessary time, temperature and concentration conditions to enable pilot-scale functionalization of the released pectic hydrocolloids from steam explosion of peel material. Response surface methodology will be used to determine these parameters using analytical methods. Consequently, the variables of temperature, time and concentration of steamed peel waste will be manipulated to determine optimal conditions for functionalizing the released pectic hydrocolloids. Functionality will be assessed by measuring resulting calcium induced viscosity using a concentric cylinder viscometer and/or oscillatory measurements using a stress controlled rheometer and related to final degree of methylation, charge distribution and molecular weight (MW) of the modified pectic hydrocolloids. Compositional analysis and structural properties will be characterized by Size Exclusion Chromatography (SEC) coupled to Multi Angle Laser Light Scattering (MALLS), Refractive Index (RI) or Conductivity Detectors; High Performance Anion Exchange Chromatography (HPAEC) coupled to an Evaporative Light Scattering Detector (ELSD) or Pulsed Amperometric Detector (PAD) and enzymatic/chemical methods. Composition of the polysaccharides present in peel wash after steam explosion will be determined by enzymatic hydrolysis and liquid chromatography with electrochemical detection. Use of polysaccharide specific enzymes (arabinase, arabinofuranosidase, etc.) will allow for determination of the contribution of individual polysaccharides. Pectin populations will be examined via interaction with antibodies that bind to specific structural epitopes on individual pectin molecules. Pectin populations will be produced by enzymatic and/or chemical methods that contain various sizes of ionically-charged or neutral, methyl-protected domains. Elucidation of the modes of anti-inflammatory actions of the health promoting compounds in citrus byproducts will be accomplished by characterizing their metabolites and pharmacokinetics, and elucidating their biochemical actions at the cellular level using in vitro assay microplate technologies. These biochemical actions subsequently will be investigated in animal trials conducted through collaborations with other research laboratories or through commercial contract research laboratories. The research will first require the isolation and chemical characterization of mammalian metabolites of the test citrus byproduct compounds, and these isolations will be achieved through established chromatographic and HPLC-MS techniques.

Progress Report
Operating parameters of temperature and time-at-temperature were tested on a variety of citrus and non-citrus biomasses using a static, batch apparatus to determine the appropriate conditions on each type of biomass for the recovery of multiple types of value-added co-products. The biomasses studied included fresh, juice-extracted red and white grapefruit peel, banana residues and olive leaves. Value-added co-products recovered included sugars, pectic hydrocolloids and phenolics. Response surface methodology was utilized to predict the optimum conditions for recovery of pectic hydrocolloids from stabilized citrus pectin peel. Additionally, citrus fruit from disease (HLB) resistant cultivars that are selected for rootstock properties, not juice quality, were processed by continuous steam explosion to determine if the high-value co-products could be recovered. The exploration of steam explosion as a means of solubilizing citrus peel pectins, sugars, flavonoids, hydroxycinnamates and limonoids and promoting their recovery using a simple water wash, hence avoiding organic solvent or harsh mineral acid extraction, was expanded by exploring its use for the production of soluble dietary fiber. Results from these experiments indicate that soluble dietary fiber can be produced using steam explosion. These results advance the achievement of Objective 1 by expanding the types of biomass and high-valued co-products that can be recovered and the optimal conditions for doing so. Modifications of citrus byproduct flavonoids by bacterial and fungal fermentations that advanced Objective 2A were carried out collaboratively. Many metabolites of the polymethoxylated flavones as well as flavanone glycosides were generated during these fermentations. Analyses of these newly formed flavonoid metabolites showed that the majority of these compounds are the same as those that occur in humans after ingestion of the polymethoxylated flavones. The microbial production of the polymethoxylated flavone metabolites typically found in humans will allow us to isolate these compounds and thus, will enable us to conduct further small animal (rodent) testing. Results of collaborative studies in mice showed that dosing the animals with eriocitrin had significant beneficial effects when the mice were fed a high fat diet. At the end of the study mice supplemented with eriocitrin showed improved blood serum levels of glucose, triglycerides, insulin, HOMA-IR, resistin, total-cholesterol, lipid peroxidation and liver triglycerides, and helped to normalize glucose and lipid metabolism in an ongoing obesogenic environment. In another collaborative study, beneficial effects were also observed with the ingestion of the polymethoxylated flavones, tangeretin and heptamethoxyflavone by rats fed a high fat diet. Significant impacts were observed on the production of key signaling proteins produced during inflammation, i.e. increased production of an anti-inflammatory cytokine IL-10, and decreased production of the pro-inflammatory cytokine, TNF-alpha. Both effects lower the health damaging effects of high fat diets in mammals. Optimal spray drying parameters for encapsulating an anti-microbial essential oil (carvacrol) in a shell of citrus pectin and alginate for use as a shelf-life extender for packaged foods were determined. This advances Objective 2B by extending our baseline studies on the use of pectic hydrocolloids for encapsulating bioactive ingredients for food formulation. A subordinate project on the effect of fruit maturity on the recovery of value-added co-products from citrus processing biomass has completed its first season for fresh peel and peel has been dried for comparison of the recovery of pectin, sugar and flavonoids from both.


Review Publications
Sun, X.N., Cameron, R.G., Bai, J. 2019. Microencapsulation and antimicrobial activity of carvacrol in a pectin-alginate matrix. Food Hydrocolloids. 92:69-73.
Cameron, R.G. 2018. Pectin in foods. In: Encyclopedia of Food Chemistry. Elsevier. pp. 208-213.
Dorado, C., Cameron, R.G., Manthey, J.A. 2019. Study of static steam explosion of Citrus sinensis juice processing waste for the isolation of sugars, pectic hydrocolloids, flavonoids and peel oil. Food and Bioprocess Technology. 12(8):1293-1303.
Kim, Y., Cameron, R.G., Williams, M.A., Lee, C. 2019. Charged functional domains introduced into a modified pectic homogalacturonan by a cocktail of pectin methylesterases isozymes from sweet orange (Citrus sinensis L. Osbeck var. Pineapple). Food Hydrocolloids. 96:589-595.