Anticholinesterase constituents from the leaves of Spondias mombin L. (Anacardiaceae)

Figures & data

Figure 1 Spray analysis of the TLC plates of precipitate and supernatant with phytochemical reagents.

Notes: Phytochemical analysis of precipitate and supernatant is shown. Samples: a = precipitate, b = supernatant; solvent system: N-hexane:chloroform (3:7); the different spray reagents: (A) vanillin/H₂SO₄, (B) Dragendorff’s reagent, (C) antimony trichloride and (D) anisaldehyde spray.
Abbreviation: TLC, thin-layer chromatography.

Figure 2 TLC bioautographic assay of precipitate and supernatant.

Notes: (A) AChE; (B) BUChE; samples: a = precipitate, b = supernatant; solvent system: N-hexane:chloroform (3:7).
Abbreviations: AChE, acetyl cholinesterase; BUChE, butyryl cholinesterase; TLC, thin layer chromatography.

Figure 3 Bulked samples from the VLC of Spondias mombin supernatant.

Notes: TLC bioautographic assay of VLC fractions is shown. Solvent system: chloroform 100%; (A) vanillin/H₂SO₄ spray; (B) enzyme, ATChI and DTNB spray.
Abbreviations: ATChI, acetylthiocholine iodide; DTNB, 5,5′-dithio-bis- (2-nitrobenzoic acid); VLC, vacuum liquid chromatography.

Table 1 Percentage cholinesterase inhibitory activity of Spondias mombin samples

Fraction	% inhibition of AChE	% inhibition of BUChE
Hexane fraction	36.16±1.11	24.86±1.09
Ethyl acetate fraction	58.10±1.08	52.66±1.34
Aqueous fraction	23.00±1.63	18.84±1.96
Precipitate	71.52±2.01	ND
Supernatant	87.33±1.98	ND

Note: Values shown as mean ± standard deviation.

Abbreviations: AChE, acetyl cholinesterase; BUChE, butyryl cholinesterase; ND, not determined.

Table 2 TLC profiles of the isolated compounds

Compounds	Detecting agent	Solvent system	Color	Rf
Compound 1	Vanillin/H2SO4	Chloroform (100%)	Purple	0.35
	Anisaldehyde	Hexane:chloroform (3:7)	Purple	0.46
Compound 2	Vanillin/H2SO4	Hexane:chloroform (2:8)	Purple	0.20
	Anisaldehyde	Chloroform (100%)	Purple	0.27
Compound 3	Vanillin/H2SO4	Hexane:chloroform (1:1)	Purple	0.64
	Anisaldehyde	Chloroform (100%)	Pink	0.51

Abbreviations: R_f, retardation factor; TLC, thin-layer chromatography.

Table 3 Analysis of isolated compounds

Properties	Compound 1	Compound 2	Compound 3
Solubility in water	Not determined	Soluble	Insoluble
Boiling point	–	–	202°C–204°C
Melting point	256°C–258°C	156°C–158°C	–
IC50(BuChE)	4.67 μg/mL	4.08 μg/mL	23.89 μg/mL
IC50(AChE)	0.88 μg/mL	1.89 μg/mL	12.51 μg/mL

Abbreviations: AChE, acetyl cholinesterase; BUChE, butyryl cholinesterase.

Figure 4 Compound 1: betulin.

Figure 5 Compound 2: campesterol.

Figure 6 Compound 3: 3,7,11,15-tetramethyl-2-hexadecen-1-ol.