Caracterização química de extratos de folhas de bacupari (Garcinia gardineriana), síntese de compostos análogos a produtos naturais e avaliação de atividades fitossanitárias

Abstract

As bis(piridilmetileno)ciclohexanonas (14C-15C), a 125 µmol L -1, inibiram totalmente a germinação de H. vastatrix e apresentaram IC 50 estimados em 1,3 e 2,6 vezes menores que o obtido para o fungicida comercial oxicloreto de cobre. A bis(2-piridilmetileno)ciclohexanona 13C foi ativa contra Rhizoctonia solani com IC 50 = 155,5 nmol por plugue. Esses compostos mais ativos foram submetidos a teste de citotoxicidade contra a linhagem de células Vero e apresentaram melhor perfil toxicológico quando comparado ao oxicloreto de cobre. Além disso, foram realizados cálculos in silico de parâmetros físico-químicos para bis(ilideno)ciclohexanonas, e os valores obtidos foram considerados adequados para agroquímicos. As 2-amino-4H-pirano-3-carbonitrilas (1D-14D) foram sintetizadas com bons rendimentos (29-94%). A maioria destes compostos, quando testados na concentração de 200 nmol por plugue, inibiu em pelo menos 50% o crescimento micelial de Botrytis cinerea. No entanto, foi observado que as soluções estoque preparadas para testes biológicos apresentavam alteração de cor quando mantidas na bancada do laboratório, iluminadas por lâmpadas de LED comuns. Isso nos levou a investigar a possível degradação fotoinduzida destes compostos. Experimentos de FT-IR-ATR e HPLC confirmaram a fotodegradação. O produto da reação fotoquímica do composto 7D, que apresentou o maior percentual de conversão fotoquímica, foi isolado e identificado como (E)-7-ciano-5-(4-nitrobenzilideno)-8-(4- nitrofenil)biciclo[4.2.0]oct-1(6)-eno-7- carboxamida (7D*). Este composto foi testado contra B. cinerea, mas nenhum efeito foi observado. Enfim, as bis(ilideno) cicloalcanonas e 2-amino- 4H-pirano-3-carbonitrilas foram avaliadas quanto ao seu efeito fitotóxico sobre folhas de plantas daninhas e efeito inibidor da AChE. Os compostos não mostraram efeito fitotóxico apreciável. A 2-amino-4H-pirano-3-carbonitrila (12D) produziu a maior inibição enzimática AChE (52,3%) a 50 µmol L -1 . Com o objetivo de se obterem compostos com maiores atividades anti-AChE, foram sintetizados análogos ao composto 12D. Entretanto, nenhum dos compostos produziu aumento da atividade inibitória. Desta forma, a principal atividade observada para os compostos foi a ação contra fungos fitopatogênicos, o que indica que tais compostos apresentam potencial para o desenvolvimento de novos fungicidas de interesse agrícola. Palavras- chave: 2-amino-4H-pirano-3-carbonitrilas. Bis arilidenos. Botrytis cinerea. Ferrugem da folha do café. Garcinia gardineriana. Hemileia vastatrix. Ilideno ciclohexanonas. Reações fotoquímicas.

In the present work, extracts from leaves of the tree Bacupari (Garcinia gardneriana) and a series of compounds derived from triterpene (lupeol), compounds analogues to curcumin and chalcones (bis(ylidene) cyclohexanones) and 4H-pyrans (2-amino-4H-pyran-3-carbonitriles) were produced and evaluated for their antifungal, phytotoxic, and anti-acetylcholinesterase (AChE). The extracts of G. gardneriana leaves were obtained using the solvents hexane, ethyl acetate and ethanol, successively. The hexane extract inhibited the germination of Hemileia vastatrix urediniospores by 98% with an IC 50 value of 211.3 µg mL -1 . The hexane extract showed low phytotoxic effect on cucumber and sorghum seed germination, and low anti-AChE activity at the concentrations evaluated. GC-MS analysis revealed lupeol as the main constituent of the hexane extract. Lupeol was isolated and tested on H. vastatrix and presented low antifungal activity. Thus, the effect produced by the extract was attributed to the synergistic action involving lupeol and minor components of the extract. Lupeol was used to synthesize a series of esters (1B-8B) and to perform the evaluation of their biological activities. However, no appreciable effects were observed in the tests performed. The bis(ylidene) cyclohexanones (1C-20C) were synthesized with good yields (28-97%). bis(pyridylmethylene) cyclohexanones (14C-15C), at 125 µmol L -1 , totally inhibited the germination of H. vastatrix, and presented IC 50 values estimated as 1.3 and 2.6 times lower than that obtained for the commercial fungicide copper oxychloride. Bis(2-pyridylmethylene) cyclohexanone 13C was active against Rhizoctonia solani with IC 50 = 155.5 nmol per plug. These more active compounds were submitted to cytotoxicity test against a Vero cell line and presented a better toxicological profile when compared to copper oxychloride. In addition, in silico calculations of the physicochemical parameters for bis(ylidene) cyclohexanones were performed, and the values were considered suitable for agrochemicals. The-amino-4H-pyran-3-carbonitriles (1D-14D) were synthesized in good yields (29-94%). Almost all compounds, when tested at 200 nmol per plug, inhibited at least 50% of the mycelial growth of Botrytis cinerea. However, we observed that stock solutions prepared for biological tests sunderwent color changing when kept on the laboratory bench, illuminated by common LED daylight tubes. This prompted us to investigate the possible photoinduced degradation of these compounds. FT-IR-ATR and HPLC experiments were carried out and confirmed the photodegradation. The product of the photochemical reaction of compound 7D, which showed the highest percentage of photochemical conversion, was isolated and identified as (E)-7-cyano-5-(4-nitrobenzylidene)-8-(4-nitrophenyl)bicyclo[4.2.0]oct-1(6)- ene-7-carboxamide (7D*). This compound was tested for antifungal activity against B. cinerea, but no effect was observed. Finally, as bis(ylidene) cycloalkanones and 2-amino-4H-pyran-3- carbonitriles were evaluated for their phytotoxic effect on leaves of weeds and for their AChE inhibitory activity. None of the produced phytotoxic activity. 2-amino-4H-pyran-3-carbonitrile (12D) produced the highest AChE enzymatic inhibition (52.3%) at 50 µmol L -1 . In order to obtain compounds with greater anti-AChE activities, analogs to compound 12D were synthesized. However, none of the compounds produced an increase in inhibitory activity. Thus, the main activity observed for the compounds was the action against phytopathogenic fungi, which indicates that such compounds present potential for the development of new fungicides of agricultural interest. Keywords: 2-amino-4H-pyran-3-carbonitriles. Bis arylidenes. Botrytis cinerea. Coffee leaf rust. Garcinia gardineriana. Hemileia vastatrix. Ylidene cyclohexanones. Photochemical reactions.