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Title:Evaluation of hops essential oil cytotoxicity and anti-cryptosporidium activity
Author(s):Aycart Mite, Danielle Francesca
Advisor(s):Andrade, Juan E
Contributor(s):Engeseth, Nicki; Witola, William
Department / Program:Food Science & Human Nutrition
Discipline:Food Science & Human Nutrition
Degree Granting Institution:University of Illinois at Urbana-Champaign
Abstract:Background. Hops essential oil (HEO) is a by-product of the brewing industry with proven antibacterial and antifungal activities. Its antiparasitic activity, however, has not been explored extensively. We sought to evaluate the ability of HEO and myrcene, a monoterpene predominantly present in HEO, to reduce Cryptosporidium parvum infection in vitro and the potential for their microencapsulation for human or animal applications. Methods. HEO and myrcene in vitro cytotoxicity (IC50) were determined in an HCT-8 intestinal cell model using flow cytometry and staining for non-viable cells with propidium iodide after treatment with controls and different concentrations of bioactives of interest (i.e. HEO and myrcene) for 24 and 48 h (37℃ and 5% CO2). To determine the anti-Cryptosporidium effect of HEO, varying concentrations of HEO or myrcene alone below their IC50 values in HCT-8 cells at 48 h were used. The evaluation was performed using two modalities: (1) confluent HCT-8 cells were infected with C. parvum sporozoites (1x104 sporozoites/mL) and immediately treated with bioactives dissolved in DMSO followed by incubation for 48 h (37℃ and 5% CO2); and (2) confluent HCT-8 cells were infected with C. parvum sporozoites, and 2 h later were treated with fresh medium containing bioactives following incubation for 46 h (37℃ and 5% CO2). In both cases, control cultures were treated with DMSO amounts equivalent to those used to deliver the bioactives. The growth of the parasites in the cell cultures was analyzed by an immunofluorescence assay using a fluorescence-conjugated anti-Cryptosporidium antibody (Sporo-GloTM). For encapsulation, HEO and myrcene inclusion complexes were prepared separately with β-cyclodextrin (β-CD) using a slurry complexation technique, which combined the bioactives and β-CD in a 2:1 molar ratio with respect to the content of myrcene present. After encapsulation, the slurry was dried by freeze-drying and characterized for particle size. Results. HEO showed low cytotoxicity to HCT-8 cells (IC50= 237.5 µg/mL). Myrcene showed higher cytotoxicity in comparison to HEO (IC50=176.5 µg/mL; P<0.05). Under modality 1 and 2, HEO elicited a dose-dependent reduction in C. parvum growth with IC50 of 60.38 and 57.96 µg/mL, respectively. In modality 1, myrcene alone dose-dependently reduced C. parvum growth showing an IC50= 44.06 and 43.35 µg/mL for food grade and analytic standard, respectively. Overall, HEO and myrcene showed high anticryptosporidial activity compared to other drugs used such as paromomycin (IC50= 277 µg/mL) and nitazoxanide (IC50= 10 µg/mL) as shown in the literature. The encapsulation efficiency of HEO was 78.1%. The particle sizes of HEO and myrcene in β-CD complexes were lower than uncomplexed β-CD. Significance. HEO and myrcene showed in vitro anti-Cryptosporidium efficacy and both area amenable for encapsulation with β-CD. Due to its relatively low anti-Cryptosporidium IC50 values, further studies will explore the effect of encapsulated HEO against cryptosporidiosis in vivo.
Issue Date:2019-12-13
Rights Information:© 2019 Danielle Francesca Aycart Mite
Date Available in IDEALS:2020-08-27
Date Deposited:2020-05

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