Investigación de biocompuestos polifenólicos con potencial antimicrobiano para el diseño y desarrollo de productos de aplicación en biomedicina

Abstract

This Doctoral Thesis entitled "INVESTIGACIÓN DE BIOCOMPUESTOS POLIFENÓLICOS CON POTENCIAL ANTIMICROBIANO PARA EL DISEÑO Y DESARROLLO DE PRODUCTOS DE APLICACIÓN EN BIOMEDICINA" digs into the antimicrobial capacity of propolis and rosemary extract (carnosic acid) against different bacteria and fungi. The investigation ranges from the elementary research to the industrial application and the main objectives are listed below: o Evaluation of the antifungal effect of propolis (P3) and carnosic acid (CA) against different bacteria and fungi (Candida y Cryptococcus) for comparative purposes related to their efficacy, in order to know the breadth of its action spectrum. o Isolate and identify the main compounds present in propolis responsible for its antifungal properties. o Propose possible mechanisms involved in the antifungal capacity of both natural extracts. o Study the potential use of these compounds at an industrial level, formulating a commercial product of wide use such as a dental gel. Moreover, using this dental gel to carry out the relevant in vitro tests and accomplish a biomedical intervention trial in humans to study its efficacy in vivo. The experimentation performed has consisted of different complementary methods, such as determination of cell viability in liquid medium, formation of macroscopic colonies in solid YPD medium or MICs calculation (minimum antifungal concentration necessary to inhibit the proliferation of an inoculum standardized). Additionally, studies on cell morphology have been carried out. In later stages, HPLC and LC / MS analysis have been necessary to determine and isolate the active P3 biocompounds, a key aspect to formulate hypotheses of the action mechanisms. Finally, and due to the industrial nature of this Doctoral Thesis, a dental gel has been manufactured as a prototype for the actual application of the active ingredients (CA and P3) that has undergone in vitro tests to verify its efficacy and stability. Afterwards, a biomedical intervention trial in humans was conducted. The main results obtained can be summarized in the following conclusions: " Propolis has a limited antifungal activity against different fungi, being in general, that of higher carnosic acid. " The combination of certain propolis with a chromatographic profile similar to that of P3 with CA is synergistic, and has antifungal activity against different strains of different Candida albicans, Candida glabrata, as well as Cryptococcus neoformans. " The identification of the P3 active compounds begins with the determination of its most antifungal fraction and ends with Z as the potential active compound. " The results obtained exhibit the existence of a synergistic fungicidal action induced by the combination of CA and P3. Furthermore, this breakthrough supports the continuous search for more natural bioactive molecules, for the purpose of their application in clinical therapies against C. albicans and possibly against other pathogenic fungi. This strategy would avoid the undesirable harmful side effects caused by some conventional antifungals. " Different hypotheses are formulated on the mechanisms of action of CA + P3 but none are conclusive, so future research is necessary. Nevertheless, one of the possible mechanisms based on the pro-oxidant capacity of some polyphenols, which can cause toxic effects, seems plausible. " Conveying CA + P3 in a widely used product such as dental gel, is totally satisfactory, both for its in vitro efficacy and for its stability. This dental gel was undergoing an in vivo trial on patients subject to antitumor treatments and with evidence of oral candidiasis. The results was quite successful considering that 85% of the patients experienced some clinical improvement. At the molecular level 60% of them had a positive response. Furthermore, 82% of participants stated that they would use the gel outside of the study