Microbial Metabolites of 3- n-butylphthalide as Monoamine Oxidase A Inhibitors

Abstract

Novel compounds with antidepressant activity via monoamine oxidase inhibition are being sought. Among these, derivatives of 3-n-butylphthalide, a neuroprotective lactone from Apiaceae plants, may be prominent candidates. This study aimed to obtain the oxidation products of 3-n-butylphthalide and screen them regarding their activity against the monoamine oxidase A (MAO-A) isoform. Such activity of these compounds has not been previously tested. To obtain the metabolites, we used fungi as biocatalysts because of their high oxidative capacity. Overall, 37 strains were used, among which Penicillium and Botrytis spp. were the most efficient, leading to the obtaining of three main products: 3-n-butyl-10-hydroxyphthalide, 3-n-butylphthalide-11-oic acid, and 3-n-butyl-11-hydroxyphthalide, with a total yield of 0.38-0.82 g per g of the substrate, depending on the biocatalyst used. The precursor-3-n-butylphthalide and abovementioned metabolites inhibited the MAO-A enzyme; the most active was the carboxylic acid derivative of the lactone with inhibitory constant (K_i) < 0.001 µmol/L. The in silico prediction of the drug-likeness of the metabolites matches the assumptions of Lipinski, Ghose, Veber, Egan, and Muegge. All the compounds are within the optimal range for the lipophilicity value, which is connected to adequate permeability and solubility.

Keywords: 3-n-butylphthalide; biotransformation; fungal strains; monoamine oxidase A inhibitor; serotonin.

Figure 1

Synthesis of 3-n-butylphthalide (1) by the hydrolysis of 3-n-butylidenephthalide to ketoacid (i-KOH in MeOH), its reduction by borohydride, and acidification (ii-THF, NaBH₄, HCl).

Figure 2

Product content of 3-n-butylphthalide (1) biotransformations catalyzed by (A) Penicillium dierckxii AM32, (B) Penicillium sp. AM91, (C) Botrytis cinerea AM235, and (D) Botrytis sp. KKP3292, determined by an HPLC-DAD at a wavelength of 274 nm.

Figure 3

LC/ESI–MS/MS chromatograms of (A) 3-n-butylphthalide (1)—positive ion mode, (B) 3-n-butyl-10-hydroxy-phthalide (2)—positive ion mode, (C) 3-n-butyl-10-hydroxy-phthalide (2)—negative ion mode, (D) 3-n-butyl-11-hydroxy-phthalide (3)—positive ion mode, and (E) 3-n-butylphthalide-11-oic acid (4)—negative ion mode.

Figure 4

Pivotal couplings for compound 2—correlation spectroscopy (orange) and nuclear multiple bond coherence (blue arrows).

Figure 5

The structures of the main products formed during fungal-mediated biotransformations of 3-n-butylphthalide (1)—3-n-butyl-10-hydroxyphthalide (2), 3-n-butyl-11-hydroxyphthalide (3) and 3-n-butylphthalide-11-oic acid (4).