Research Article
Antibiotics Produced by Bacteria of the Genus Bacillus
Rami Hammod*
Corresponding Author: Rami Hammod, High Institute of Marine Research, Tishreen University, Syria.
Received: December 18, 2024; Revised: December 27, 2024; Accepted: December 30, 2024 Available Online: Janauary 24, 2025
Citation: Hammod R. (2025) Antibiotics Produced by Bacteria of the Genus Bacillus. J Infect Dis Res, 8(1): 398-401.
Copyrights: ©2025 Hammod R. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
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The results showed that the total concentration of the antibiotic polymyxin present in the organic extract of each of the four species of the genus Bacillus (B. cereus, B. subtilis, B. circulans, B. polymyxa), was 2.3 mg/l for the extract of B. subtilis, it was 4.8mg/l for B. polymyxa extract and 1.7 mg/l for B. circulans extract. While no concentration was recorded for B. cereus extract.

Keywords: Bacillus, Antibiotics, Density liquid layer
INTRODUCTION

The Bacillus species are resistant to unfavorable environmental conditions by producing spores [1-3]. In addition, bacteria of this genus are distinguished from other bacteria by producing primary and secondary metabolic compounds (such as antibiotics, enzymes and organic acids) in large quantities. The most important characteristic of antibiotics of this genus is that they are peptide antibiotics. They may be linear, but most of them are ring-shaped [1,4]. Bacillus bacteria produce many peptide antibiotics, which are effective against a broad spectrum of pathogenic bacteria.

Identification of antibiotics and their proportions in extracts of species of the genus Bacillus:

Four antibiotics (Polymyxin, Tyrothrici, Bacitracins, Gramsidin) and their ratios were identified in organic extracts of Bacillus species. The optimal conditions for the separation and purification of these antibiotics were determined at the lowest economic cost, given the importance of these antibiotics in the pharmaceutical industry and their effective effect on pathogenic microbes and cancer cells. This is because the manufacture of some antibiotics requires secondary compounds that contain different chemical groups, hence the importance of obtaining these compounds from their natural sources [1,3].

Chemical analyzes were conducted for the methanolic extracts of four species of the genus Bacillus isolated from the Apamea region in summer, because they showed higher activity than the dechloromethane extracts of the species isolated from other sites.

A new method was devised to isolate and purify these antibiotics and obtain them naturally with high purity, and their concentration was studied using a spectrophotometer.

The method depends on the following steps:

Bacillus species extracts were placed in a centrifuge (eppendorf 5810r, Germany) at a centrifugation speed of 4000 rpm, at different temperatures (-5, 0, 5)°C.

During different periods of time (10,20,30) min, the lower density liquid layer was separated from the higher density layer.

In order to find out what is the absorption wavelength of each antibiotic, the spectrophotometer was filtered using the organic solution methanol (as a control).

The appropriate wavelength was determined experimentally by scanning the wavelength within the range (190-800) nm for each of the four titer antibiotics (polymyxin, Tyrothrici, Bacitracins, and Gramsidin) to find out the appropriate wavelength at which the antibiotic has the highest absorbency using spectrophotometric technology.

A titer series for each of the four antibiotics with different concentrations (0.01, 0.02, 0.05, 0.1, 1) mg/ml was used to create the standard curve and obtain the equation to calculate 

the antibiotic concentrations and their ratios based on the absorbance values of these different concentrations.

The concentration of each antibiotic in the organic extracts was determined in each of the layers forming the liquid layer with low density and the bottom layer with high density by spectrophotometer for each antibiotic.

The percentage of each antibody in each isolate (the percentage of purification) was calculated according to the relationship: (the concentration of the antibody in the separated layer ÷ the concentration of the antibody in the original sample) x 100 (Table 1).

RESULTS AND DISCUSSION

The Isolation and purification of antibiotics (proteins) from extracts of the genus Bacillus:

The proportion of antibiotics in extracts of the genus Bacillus isolated from the Apamea region during the summer season was determined because it was more effective against pathogenic bacteria.

It had a wide spectrum of biological activity against pathogenic bacteria and various biologically active chemical compounds compared to other sites.

It was found through the results after the process of separating the liquid layer of low density (upper layer) from the liquid layer of high density (lower layer) and measuring the absorbance through a spectrophotometer.

For each antibiotic, it was found that the optimal conditions for the purification and isolation of antibiotics were at a centrifugation speed of 4000 rpm, a centrifugation time of 30 min, and a temperature of -5°C. The antibiotic concentration was determined in each layer, and we noticed that the separation rate of some antibiotics exceeded 95%.

The concentration of the polymyxin antibiotic

The results showed that the total concentration of the antibiotic polymyxin present in the organic extract of each of the four species of the genus Bacillus (B. cereus, B. subtilis, B. circulans, B. polymyxa), was 2.3 mg/l for the extract of B. subtilis. It was 4.8mg/l for B. polymyxa extract and 1.7 mg/l for B. circulans extract. While no concentration was recorded for B. cereus extract (Table 2).

As for the study of each phase (liquid with low density, liquid with high density) of the methanolic extract when centrifugation at different temperatures (0,5,5) during different centrifugation periods (10,20,30) min. The results showed that:

When centrifuged at 0°C for 10 min for the low-density liquid layer, the concentration of polymyxin present in the bacterial extract of Bacillus species was 0.16 mg/l with respect to the extract of B. subtilis. It was 3.7 mg/l for B. polymyxa extract and 0.8 mg/l for B. circulans extract.

As for the liquid layer with a high density, it was observed that the concentration of polymyxin present in the extracts of species of the genus Bacillus was 0.9 mg/l for B. subtilis extract, 2.4 mg/l for B. polymyxa extract and 0.3 mg/l for B. circulans extract (Table 2).

The percentage of the isolate upon centrifugation at 5°C for 10 min was 39.13% for B. subtilis extract, 45.28% for B. polymyxa extract, and 17.64% for B. circulans extract (Table 2).

As for centrifugation at 5°C for 10 min for the low-density liquid layer, the polymyxin concentration was 1.2 mg/l for the B. subtilis extract, 2.9 mg/l for the B. polymyxa extract, and 0.4 mg/l for the B. polymyxa extract of B. circulans extract.

As for the liquid layer with a high density, it was observed that the concentration of polymyxin present in the bacterial extracts of the species of the genus Bacillus was 1.4 mg/l for the B. subtilis extract, it was 3.1 mg/l for the B. polymyxa extract and it was 0.9 mg/l for the B. subtilis extract circulans.

The percentage of the isolate upon centrifugation at 0°C for 10 min for B. subtilis extract was 56.52%, for B. polymyxa extract was 58.49% and for B. circulans extract was 52.94% (Table 2).

As for centrifugation at a temperature of -5°C for 10 min for the low-density liquid layer, the polymyxin concentration was 1.5 mg/l for the B. subtilis extract, 2.6 mg/l for the B. polymyxa extract and 0.1 mg/l. For B. circulans extract.

As for the liquid layer with high density, the results showed that the polymyxin concentration was 1.7 mg/l for B. subtilis extract, 3.5 mg/l for B. polymyxa extract and 1.1 mg/l for B. circulans extract (Table 2).

The results showed that the percentage of polymyxin antibiotic upon centrifugation at -5°C for 10 min was 73.91% for B. subtilis extract, 66.03% for B. polymyxa extract and 64.70% for B. circulans extract (Table 2).

To find out the effect of centrifugation on the concentration of antibiotics in organic extracts, centrifugation was carried out at 20 and 30 min at a temperature of -5°C, and the results were as follows:

When centrifuging at a temperature of -5 °C for 20 min for the low-density liquid layer, it was observed that the polymyxin concentration was 1.6 mg/l for B. subtilis extract, 2.8 mg/l for B. polymyxa extract and 0.4 mg/l for B. polymyxa extract for B. circulans extract.

As for the liquid layer with a high density, the results showed that the concentration of polymyxin present in the bacterial extracts of species of the genus Bacillus was 1.9 mg/l for the B. subtilis extract, 4.1 mg/l for the B. polymyxa extract and 1.3 mg/l for the B. circulans (Table 2).

The results showed that the percentage of polymyxin upon centrifugation at -5°C for 20 min was 82.60% for B. subtilis extract, 77.35% for B. polymyxa extract and 78.47% for B. circulans extract (Table 2).

When centrifuging at a temperature of -5°C for 30 min for the low-density liquid layer, it was observed that the polymyxin concentration was 1.8mg/l for the B. subtilis extract, it was 2.3 mg/l for the B. polymyxa extract, and it was 0.1mg/l for the B. polymyxa extract for B. circulans extract. As for the liquid layer with high density, the results showed that the polymyxin concentration was 2.1 mg/l for B. subtilis extract, 4.7 mg/l for B. polymyxa extract and 1.5 mg/l for B. circulans extract (Table 2).

The results showed that the percentage of polymyxin antibiotic upon centrifugation at -5°C for 30 min was 91.30% for B. subtilis extract, 97.91% for B. polymyxa extract and 88.23% for B. circulans extract.

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