Hydroxylamine chemically engineered extracts as source of antimycobacterial compounds

PARES, V; FURLÁN, R; RAMALLO, IA; GRAMAJO, H; GAGO, G.

Salta

Congreso; XIV PABMB Congress and LV Reunión Anual de la Sociedad Argentina de Investigación Bioquímica y Biología Molecular (SAIB).; 2019

Tuberculosis (TB) is one of the top ten causes of death and the leading cause from a single infectious agent, exceeding both malaria and HIV. The emergence of multidrug-resistant TB (MDR-TB) presents an increasingly difficult therapeutic challenge; so much that MDR-TB is now the main cause of death due to antimicrobial resistance. Unfortunately, the lethality of TB combined with its multidrug-resistant capacity has now transformed this long-neglected disease into a global health priority. Therefore, there is an urgent need for the development of new drugs that could reduce the duration and complexity of current therapies. Biological selection makes natural products (NPs) an excellent source of bioactive substructures. The chemical diversification of natural extracts uses natural substructures as scaffolds to produce new bioactive semi-synthetic molecules. This approach involves the chemical transformation of reactive fragments commonly found in NPs (like carbonyl groups) to introduce functionalities that are relevant for bioactivity (like N-O fragment). We report here the chemical diversification of a set of 18 essential oils (EOs) with NH2OH.HCl in refluxing ethanol for 7 h to produce chemically engineered EOs with different chemical composition, according to the GC-MS/NMR analysis. The chemically engineered EOs were used to study Mycobacterium smegmatis growth inhibition using a microplate assay at 200 μg/mL final concentration. Of the complete set, the chemically engineered Cuminum cyminum L. essential oil (CCY-M) was the only mixture that showed inhibitory properties against this microorganism. Quantitative biological analysis of CCY-M and the starting essential oil C. cyminum (CCY) was performed giving a MIC= 50 μg/mL and a MIC> 200 μg/mL, respectively. A M. smegmatis TLC-bioautography assay was developed ad hoc, and applied to the CCY-M showing an intense inhibition halo that was absent in CCY. With this evidence, the reaction was scaled up and, the bioassay-guided fractionation of CCY-M led to the purification of one compound that was identified by NMR and HRMS as (Z)-4-isopropyl benzaldehyde oxime (1). This isolated structure contains the N-OH moiety confirming that it is a product of the reaction. This compound was probably generated by the reaction of cuminaldhyde (major constituent of CCY) with hydroxylamine. Interestingly, the oxime 1 MIC was 9.06 μM, two and a half time better than isoniazid (first-line antibiotic in the treatment of TB, MIC= 36 μM). The potency of the oxime 1 against M. tuberculosis H37Ra was tested, but the results showed a MIC= 72.45μM, much higher than the expected according to the previous M. smegmatis results. Future chemical modifications of oxime 1 could be introduced to increase its activity against M. tuberculosis H37Ra. However, the most relevant point reported here is the chemical diversification of natural mixtures itself: it is a simple and low-cost strategy to produce new antimycobacterial molecules.