lAB 5: DETERMINATION OF ANTIMICROBIAL EFFECTS OF MICROBIAL EXTRACTS

INTRODUCTION

 Certain group bacteria can produce antimicrobial substances with the capacity to inhibait the growth of pathogenic and spoilage micro-organism.An antibacterial is a compound or substance that kills or slows down the growth of bacteria.The term is often used synonymously with the term antibiotic(s); today, however, with increased knowledge of the causative agents of various infectious diseases, antibiotic(s) has come to denote a broader range of antimicrobial compounds, including anti-fungal and other compounds. While antibiotic is A substance that is capable of stopping the growth of, or destroying, bacteria and other microorganisms which cause bacterial disease. Many antibiotics are themselves produced by microorganisms (bacteria and molds). 

Antibiotics are germicides that are safe enough to be swallowed or injected into the body. With advances in medicinal chemistry, most of today's antibacterials chemically are semisynthetic modifications of various natural compounds. These include, for example, the beta-lactam antibacterials, which include the penicillins (produced by fungi in the genus 'Penicillium'), the cephalosporins, and the carbapenems. Compounds that are still isolated from living organisms are the aminoglycosides, whereas other antibacterials—for example, the sulfonamides, the quinolones, and the oxazolidinones—are produced solely by chemical synthesis. Accordingly, many antibacterial compounds are classified on the basis of chemical/biosynthetic origin into natural, semisynthetic, and synthetic. Another classification system is based on biological activity; in this classification antibacterials are divided into two broad groups according to their biological effect on microorganisms: bactericidal agents kill bacteria, and bacteriostatic agents slow down or stall bacterial growth.

RESULT: 

 part 1

Strains of LAB	Strains of spoilage/pathogenic bacteria	Inhibition zone (cm)
LAB Species	Staphylococcus aureus	(1.4+1.3)/2 =1.35 cm
	Escherichia coli	(0.8+1.0)/2 =0.9 cm

part 2

E COLI

A	B
0.173	0.172
0.270	0.225
0.346	0.362
0.392	0.389

Figure 1: graph  shows serial dilution of extracellular extract using E.coli

 S.aureus

A	B
0.557	0.659
0.770	0.670
0.820	0.960
0.846	0.981

Figure 2 : Graph shows serial dilution of extracellular extract using S.aureus

DISCUSSION

Part  1. Determination of bacterion activity via agar diffusion test
 

The concentration of the compound will be highest next to the disk, and will decrease as distance from the disk increases. If the compound is effective against bacteria at a certain concentration, no colonies will grow where the concentration in the agar is greater than or equal to the effective concentration. This is the zone of inhibition. Thus, the size of the zone of inhibition is a measure of the compound's effectiveness: the larger the clear area around the filter disk, the more effective the compound.

Part  2. Determination of bacterion activity via optical density

There is one step that we need to prepare a negative-control for ‘auto-zero’ via spectrophotometer.

 

 Spectrophotometry involves the use of a spectrophotometer. A spectrophotometer is a photometer (a device for measuring light intensity) that can measure intensity as a function of the light source wavelength. Important features of spectrophotometers are spectral bandwidth and linear range of absorption or reflectance measurement.

Spectrophotometers are commonly used for the measurement of transmittance or reflectance of solutions, transparent or opaque solids, such as polished glass, or gases. However they can also be designed to measure the diffusivity on any of the listed light ranges that usually cover around 200nm - 2500nm using different controls and calibrations. Within these ranges of light, calibrations are needed on the machine using standards that vary in type depending on the wavelength of the photometric determination.

An example of an experiment in which spectrophotometry is used is the determination of the equilibrium constant of a solution. A certain chemical reaction within a solution may occur in a forward and reverse direction where reactants form products and products break down into reactants. At some point, this chemical reaction will reach a point of balance called an equilibrium point. In order to determine the respective concentrations of reactants and products at this point, the light transmittance of the solution can be tested using spectrophotometry. The amount of light that passes through the solution is indicative of the concentration of certain chemicals that do not allow light to pass through.

The use of spectrophotometers spans various scientific fields, such as physics, materials science, chemistry, biochemistry, and molecular biology. They are widely used in many industries including semiconductors, laser and optical manufacturing, printing and forensic examination, as well in laboratories for the study of chemical substances. Ultimately, a spectrophotometer is able to determine, depending on the control or calibration, what substances are present in a target and exactly how much through calculations of observed wavelengths.

CONCLUSION 

 In conclusion, the results show that lactic acid bacteria(LAB) may act as a bio preservatives. From the experiment, the LAB succesfully shows its bio preservatives properties on both gram-positive and gram-negative bacteria which are Escherichia coli and Staphylococcus aureus. Antimicrobial compounds produced by LAB have provided these organisms with a competitive advantage over other microorganisms.