INTRODUCTION Madagascar and the Mascarene Islands. Anthocleista vogelii is

INTRODUCTION

Anthocleista vogelii Planch is a tree- and shrub-like plant of the genus Anthocleista in the Gentian family (Gentianaceae) and native mainly to tropical Africa, Madagascar and the Mascarene Islands. Anthocleista vogelii is commonly called “cabbage tree” and is a medicinal plant widely used in West Africa. The plant is traditionally used to treat various diseases such as diabetes mellitus, constipation, hernia, malaria, hypertension, hemorrhoidsO1 , syphilis, stomach aches and diabetes (Okorie, 1976; Olubomehin, Abo, & Ajaiyeoba, 2013).  

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Anthocleista vogelii is a medicinal plant with pharmacological activities such as laxative, analgesic, antiulcerogenic, antiplasmodial, antimicrobial, hypoglycaemic, antiobesity, anti-inflammatory, antitrypanosomal and spasmogenic activities (Anyanwu, Ur-Rehman, Onyeneke, & Rauf, 2015). A number of research on medicinal plants stops at the pharmacological activities of the extracts or fractions. This study sought to isolate and characterize compounds from Anthocleista vogelii root bark and O2 to investigate their pharmacological activities as it relates to their ethno- botanical O3 use.

RESULTS AND DISCUSSION

The n-butanol and chloroform fractions were concentrate were subjected to column chromatographic separation and purification on a PTLC to give Compounds 1 (7.2g) and 2 (3.9 g) respectively.

 

1

2

Figure 1: Structure of compound 1and 2 isolated from the root bark of A. vogelii Planch

Compound 1: The FT-IR data ?max : 2927.7, 2958.6, 2860.2 are C-H stretching vibrations, 1728.1 C=O of ester, 1276.8 and 1072.3 C-O of ester, 1461.9 of C=C of aromatic, 1579.6 C=N and 1124.4 C-N of aromatic amine, monosubstituted aromatic ring at 742.5,702.0, 651.9  cm?1 (Figure S1, Supporting Information).The 1H NMR spectrum of compound 1 (Table 1) exhibited two methyl groups at ?H 0.87 and 0.89 ppm, five methine protons at ?H  1.67, 7.52, 7.52, 7.68, and 7.69 ppm, eight methylene groups at ?H 1.18, 1.38, 1.29, 1.40, 1.41, 1.59, 1.91, 4.21 ppm (Figure S2, Supporting Information).

Consistent with the observations on 13C NMR spectrum with 17 carbon signals (Figure S3, Supporting Information), and DEPT-90 and -135 sub-spectra with two methyl, eight methylene, five methane, and four pyridine carbons (Figure S4 and S5, Supporting Information).

Further analysis of the HSQC data reveals a direct correlation of H-2, H-3´ and H-6´ (?H 7.52 and 7.68) to C2, C-3´ and C-6´ (?C 68.1, 124.1, and141.7) (Figure S6, Supporting Information). COSY data showed that there is a correlation  between H-2 at ?H 4.66 and H-9 at ?H 1.67 indicating that the oxy-quaternary carbon is  two bonds away from the methylene carbon at C-4 (?C 26.2) and the absence of further correlation is due that the underlying fact that the chemical environment of the carbon is bordered  by oxo group and quaternary carbons (Figure S7, Supporting Information). In the key cross peak in the HMBC spectrum (Figure 2) (Figure S8, Supporting Information), the acetate unit is located at C-1´ of the isonicotinic acid methyl ester unit was verified by correlations from H-4, H-3´ and H-7´ (?H 4.21, 7.52 and 7.52) to C-1´  (?C 166.9).

Figure 2: Key HMBC (red curved arrows) and 1H?1H COSY (blue curved arrow) correlations of compounds 1 and 2.

In the NOESY spectrum (Figure 3) (Figure S9, Supporting Information), spatial correlations between H-2 at ?H 4.66 to H-3 at ?H 1.35, H-4 at ?H 1.35 and H-15 at  ?H 1.35 indicated that H-11 and -15 are ?-oriented. The pyridine moiety of compound 1 (Isonicotinic acid 8-methyl-decyl ester) was confirmed by after acid hydrolysis. The structure of compound 1 is shown in Figure 1.

Figure 3: Key NOESY correlations of compounds 1

Sweroside, (4aS,5R,6S)-1-Oxo-5-vinyl-4,4a,5,6-tetrahydro-1H,3H-pyrano3,4-cpyran-6-yl ?-D-glucopyranoside (2): The FT-IR data ?max; 3417.6 O-H of  alcohol and 1026.1 and 1001.0 C–O of the ?-D-glucopyranoside, C–H of alkane at 2914.2, 2061.8,C = O of ester absorption moved to lower wavenumbers due to conjugation at 1693.4, C = C of alkene at 1616.2, cm?1 (Figure S10, Supporting Information). 1H-NMR (DMSO-d) spectrum of compound 2 (Table 1) exhibited  ten methane protons at ?H  2.69, 3.27, 3.27, 3.31, 3.44, 3.65, 3.76, 4.05, 4.67, 5.26, 5.31, 5.53, 5.54, and 7.58 ppm and four methylene protons at ?H 1.72 , 3.68, 4.35, 5.24 ppm (Figure S11, Supporting Information).

The 13C-NMR spectrum of compound 2 showed 16 signals; 10 carbon signals attributed to the aglycone part and 6 signals to the sugar moiety (Figure S12, Supporting Information). The DEPT-90 and -135 sub-spectra confirm the proton signals with four methylene and fourteen methine carbons (Figure S13 and S14, Supporting Information). The 1H-NMR spectrum showed a downfield doublet signal at ? 7.58 d, J = 7.4 Hz, indicating an oxyolefinic proton of the secoiridoids (Duke, 1985; Lim, 2014). Proton signal at ? 4.35,d, J = 6.5 was assigned to the methylene of C-12 by comparison with the data  of Aberham, Pieri, Croom, Ellmerer, & Stuppner (2011) and  Dwarika Prasad & Sati (2012). Also the two doublets at ? 5.50, d, J = 8.7, 9.4 Hz and ? 5.24, d, J = 9.4, 1.9 Hz that were assigned to two protons of a methylene group. The HSQC data indicated a direct correlation between 7.46 ,d, J = 2.4, 5.50 ,d, J = 8.7 and 9.4 to  C-7 (151.4 ppm), C-11 (132.3 ppm) respectively(Figure S15, Supporting Information), while thepostion of the olefinic bond is confirmed by COSY data showing a correlation at H-7 (?H 7.46) and H-1 (?H 3.13) while the position of olefinic methylene hydrogen is verified by correction at H-12 (?H 4.35 ppm) and H-1 (?H 3.13 ppm) (Figure S16, Supporting Information). The HMBC data shows that the ?-D-glucopyranoside anomeric carbon is located at C-6  by the correlation between C-6(104.8 ppm) and H-2′ (?H 4.94 ppm) (Figure S17, Supporting Information) In addition, the sugar moiety includes resonances of an anomeric proton signal at ? 4.94, d, J = 8.0 Hz, together with five proton signals for the remaining ?-D-glucopyranoside protons. The NOESY spectral data are strongly in agreement with those reported for sweroside (Figure S18, Supporting Information) (De Oliveira et al., 2013; Devi Prasad et al., 2000).

 

 

 

 

 

 

 

 

 

 

 

Table 1. 1H and 13C (600 MHz) NMR Spectroscopic Data for Compounds 1 (Chloroform-d) and 2 (DMSO-d) (? in ppm, J in Hz)

                                1

                          2

Position

?H (J in Hz)

?C, type

?H (J in Hz)

?C, type

1

 

 

3.13,ddd,(7.7,6.5)

26.8,CH

2

4.23,t,( 11.5,6.1)

68.1, CH2

1.72, dt,(6.1,4.9)

24.3,CH2

3

1.67,dd,(13.8,6.8)

28.9, CH2

5.24,d,(9.4,1.9)

67.7,CH2

4

1.41,dd, (6.8,13.0)

23.0, CH2

 

 

5

1.40,dd, (6.8,13.0)

29.7, CH2

 

164.7,C

6

1.29,dd,(6.8,13.0)

23.7, CH2

 

104.8,C

7

1.31,dd,( 6.6, 4.4)

30.1, CH2

7.46 ,d,(2.4)

151.4,CH

8

1.18,m,( 12.72,6.08)

38.6, CH2

 

 

9

1.41,m,( 6.1,7.0)

36.6, CH

5.50 ,d,(8.7,9.4)

98.1,CH

10

1.64,m,( 13.1,7.4)

30.3, CH2

2.49,ddd,(7.7,6.1)

41.5,CH

11

0.84,d,( 7.38)

10.9, CH3

2.49,ddd,(7.7,6.1)

132.3,CH

12

0.91,t,( 7.24)

14.1, CH3

4.35,d,(6.5)

120.3,CH2

 

167.8,C

 

 

 

130.9, C

4.94, d,(8.0)

95.6, CH

7.51,d,( 1.5,5.64)

128.7, CH

4.48,dt,(2.0, 6.9)

73.1, CH

7.69,d,( 0.40,5.64)

132.3, CH

3.66,m,(12.0,5.0)

76.4, CH

7.69,d,( 0.40,5.64)

132.3, CH

3.40, m,(6.9)

70.1, CH

7.51,d,( 1.5,5.64)

128.7, CH

3.15 ,m,(6.9)

77.3, CH

 

 

3.68 , dd,(8.8)

61.1,CH2

The results of screening C1 and C2 against lipase, ?-amylase and ?-glucosidase is shown in Table 2. Compound 1 was not active against PLE and ?-amylase, O4 but was moderately active against ?-glucosidase (IC50 = 40.28 ± 0.063 ?g/mL). Compound 2 had inhibitory activities against PLE and ?-glucosidase with IC50 values of 24.43 ± 0.096 and 10.28 ± 0.015 ?g/mL respectively, O5 but showed no activity against ?-amylase.

Table 2: Calculated IC50 values of lipase, ?-amylase and ?-glucosidase inhibitory activities

Compounds

PLE
IC50 (?g/mL)

alpha amylaseO6 
IC50 (?g/mL)

alpha-glucosidase
IC50 (?g/mL)

1

40.28 ± 0.063

2

24.43 ± 0.096

10.28 ± 0.015

Orlistat

0.068 ± 0.001

NA

NA

Acarbose

NA

41.10 ± 0.031

2.59 ± 0.001

Values are mean ± SD, n = 3. NA- not applicable; (-) compounds did not inhibit 50 % of enzymes

In figure 4, the compound 1 significantly (p