In vitro activity, extraction, separation and structure elucidation of antibiotic produced by Streptomyces anulatus NEAE-94 active against multidrug-resistant Staphylococcus aureus

Figures & data

Figure 1. Schematic representation of the procedures for antibiotic isolation, purification, characterization and structure elucidation.

Table 1. Diameter of inhibition zone (mm) formed by multidrug-resistant Staphylococcus aureus following exposure to crude extracts from the fermentation broth of Streptomyces anulatus NEAE- 94.

Solvents	CCl4	Butanol	Petroleum ether	Chloroform	Hexane	Ethyl acetate	Diethyl ether	Benzene
pH 3	20	22	0	23	16	36	27	22
pH 7	21	0	0	22	0	33	22	0
pH 10	23	0	0	27	0	32	23	0

Table 2. Physico-chemical properties of bioactive metabolites in the ethyl acetate extract from the fermentation broth of Streptomyces anulatus NEAE-94.

Properties
Colour	Yellowish orange
Nature	Oily
	Soluble in: hexane, diethyl ether, ethyl acetate, benzene, chloroform, butanol, carbon tetrachloride
Solubility	Insoluble in: petroleum ether
Rf value	0.83
UV λmax (nm)	206, 218, 225, 237, 256, 428, 434 and 446
Elemental analysis	C, 36%; N, 54%; O, 9%
Molecular weight (EI-MS (m/z))	1255.40

Figure 2. FT-IR spectrum of the bioactive metabolites in the crude ethyl acetate extract from the fermentation broth of Streptomyces anulatus NEAE-94.

Table 3. ¹H-NMR chemical shifts of the bioactive metabolites in the ethyl acetate extract from the fermentation broth of Streptomyces anulatus NEAE-94.

Hydrogen type	ppm (multiplicity)
Solvent residual peak	0.86
RCH2R	1.23–1.27
(RCH2 CH2R)	1.6
R2C = CRCHR2	1.9
RCOCH3	2.1–2.3
COOR	2.5
ArCH2R	2.8
R2CHOR	3.2
RCH2OH	3.4
OCOR	3.6
RCOOCH3	3.84
Aromatic H	6.8–6.81
Solvent residual peak	7.2
Aromatic H	7.84–7.87

Figure 3. ¹H-NMR spectrum of the antibacterial agent produced by the Streptomyces anulatus NEAE-94.

Figure 4. Mass spectrum of the antibacterial agent produced by Streptomyces anulatus NEAE-94.

Table 4. Bioactive constituents identified in the ethyl acetate extract from the fermentation broth of S. anulatus NEAE-94 using GC-MS analysis.

Molecular weight	Molecular formula	Peak area (%)	Compound	Retention time (min)
170	C10H18O2	4.2	2-Decenoic acid	15.53
268	C19H40	0.75	Nonadecane	22.69
270	C17H34O2	3.68	Hexadecanoic acid,methyl ester	23.07
256	C16H32O2	9.30	n-Hexadecanoic acid	23.61
282	C20H42	0.74	Eicosane	24.00
294	C21H42	1.03	1-Heneicosene	25.08
296	C21H44	0.75	n-Heneicosane	25.24
294	C19H34O2	3.51	Octadecanoic acid, methyl ester	25.59
284	C18H36O2	9.83	Octadecanoic acid	26.11
310	C22H46	1.70	Docosane	26.42
324	C23H48	6.41	Tricosane	27.56
338	C24H50	3.89	Tetracosane	28.66
352	C25H52	5.62	Pentacosane	29.72
366	C26H54	5.23	Hexacosane	30.73
380	C27H56	6.22	Heptacosane	31.82
358	C21H42O4	5.89	Octadecanoic acid,2-hydroxy-1-(hydroxymethyl)ethyl ester	32.15
392	C28H56	0.67	1-Octacosene	32.55
394	C28H58	3.46	Octacosane	33.04
410	C30H50	1.74	Squalene	33.56
408	C29H60	2.47	Nonacosane	34.47
424	C28H56O2	1.34	Tetradecanoic acid, tetradecyl ester	35.64
422	C30H62	1.39	Triacontane	36.18

Figure 5. Chemical structure of bioactive components identified in the ethyl acetate extract from the fermentation broth of Streptomyces anulatus NEAE-94, using GC–MS analysis.