Estudo espectroscópico da solubilização do 5,7-dimetoxicumarina em solventes hidrofóbicos, hidrofílicos e micelares usando a espectroscopia de emissão a 77 K e cálculos TD-DFT

Abstract

The 5,7-dimethoxycoumarin (DMC) is a compound derived from the coumarin family, which is present in plants and in food such as lemon of the citrus species and papaya of the carica papaya species. Coumarins stand out for their biological properties that include antimutagenic action against aromatic heterocyclic amines (AHA), mutagenic compounds produced during the cooking of animal protein, such as 2-amino-3-methylimidazo[4,5-f]quinoline (IQ). In order to understand the pharmacological action and chemical behavior of DMC in bio membranes, a spectroscopic study of the solubilization of DMC in hydrophobic, hydrophilic and micellar solvents at 77K was performed using 77K emission spectroscopy, UV/Visible absorption and theoretical calculations at the TD-DFT level and semi empirics for the determination of their species in the following solvents: methylcyclohexane (MCH), ethanol and 2,2,2-trifluoroethanol (TFE) and buffer solution with pH under physiological conditions and controlled ionic strength. The species found were: (i) in MCH, molecular aggregate via intramolecular hydrogen bonding (AM-H) and the monomer (M); (ii) in ethanol: and the monomer of the complex linked by hydrogen bonding between DMC and ethanol (M-H); (iii) in TFE: the monomer of the strong hydrogen bonding complex (strong M-H) between DMC and TFE; (iv) buffer solution: strong hydrophobic molecular aggregate (AHF). Each species showed distinct photophysical properties and can be used as direct fluorescent probes to investigate which species is solubilized inside the neutral, anionic and cationic micelles, prepared under physiological conditions. For instance, the active species in the MCH solvent (hydrophobic medium), is the DMC monomer, it emits fluorescence through the excited electronic state of lower singlet energy S1 formed via internal conversion and marked as (π, π*), the maximum of band is at 388 nm. Phosphorescence is emitted by the excited electronic state of the lowest triplet energy T1 (π, π *), formed through spin-orbital coupling of the type: S1 (π, π *) ↔ T2 (n, π *) ↔ T1 (π, π *), it presents a spectrum with the 0-0 band located at 470nm, originating from the double bond C3 = C4 of the ring of the pyrone. For the neutral micelle, the active species inside the micelle is DMC-H, which is anchored in the aqueous region with the double ethylene bond of the pyrone ring and forms a hydrogen bond with water through the pyrone carbonyl. The DMC's di-methoxy-benzene group is in the hydrophobic region. The species determined inside the micelles were used to simulate the inhibition of IQ mutagenesis and some of its more potent derivatives, initially using PM3 semi-empirical calculations. The results of formation heat calculations (ΔHf) indicated that the energy of the mutagenic IQ decreased 131% when forming a hydrogen bond with the antimutagenic DMC, indicating that the inhibition of the mutagenic IQ by DMC may be via interaction between the amine group of the IQ and DMC carbonyl through intermolecular hydrogen bonding. These results can help to understand the transport mechanism through membranes and demonstrate that 77K emission spectroscopy is a very sensitive technique, capable of probing directly inside the micelle, using fluorescent probes and contributing to the understanding of the mechanism of inhibition of mutagenesis of IQ and derivatives by DMC species.