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Comparing antiseptics

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Analysis Comparing antiseptics An experiment was carried out measure the effectiveness of the antiseptics, tea tree oil and chlorhexidine on both Escherichia coli(gram negative) and Bacillus subtillis (gram positive) Once the plate had been incubated for one day (24 hours) at 30?C, the plates were analysed optically and the zones of inhibitions were measured using a metric ruler (mm), so that the average could be achieved and a justifiable conclusion to be made. E. coli and Bacilllis subtilis are Mesophiles which tolerate temperatures between 20 -40?C , temperatures above this would denature the protein structure of the microorganisms. The disc dipped in sterile water is shown as the control variable. This should not produce a zone of inhibition because the properties of water donot cause cell death to the bacterial cell wall. T-Test for the gram-negative bacteria (E.coli) Plate Zones of inhibition Diameter mm Chlorhexidine (y) Zones of inhibition Diameter mm Tea tree oil (x) Zones of inhibition Area mm2 Chlorhexidine Zones of inhibition Area mm2 Tea tree oil 1 15 20 225 400 2 15 24 225 576 3 16 20 256 400 4 14 16 196 256 5 16 15 256 225 6 16 17 256 289 7 15 20 255 400 8 15 17 255 289 Total 122 149 1924 2835 Mean 1 15.250 18.620 240.500 354.375 Mean 2 15.0 19.0 241 354 The effectiveness of the two antiseptics on E.coli (gram negative) was measured by carrying out a statistical T test. This was done to analyse, which active ingredient in the antiseptic was better in causing lysis (cell death), in the bacterium E. coli. The experiment was repeated eight times and the diameter of the zones of inhibition for tea tree oil and chlorhexidine were measured accordingly. Statistical T- test T = (x) - (y) (?x2/nx) - (x)2 + (?y/ny) - (y)2 nx - 1 ny-1 Null hypothesis - There is no significant difference between the two sets of results, due to chance. ...read more.


?x = 18 + 20 + 17 + 14 + 21 + 24 + 15 + 18 = 147 ?x2 = 324 + 400 + 289 + 196 + 441 + 576 + 225 + 324 = 2775 nx= 8 number of samples (x) = Mean of x is 147/8 = 18.375 18.0 (3.sf.) (x)2= 182 which is 324 (3.s.f) Calculations for the Chlorhexidine y = Chlorhexidine (mean of diameter) ?y = 24 + 23 + 15 + 20 + 23 + 26 + 21 + 22 = 174 ?y2 = 576 + 529 + 225 + 400 + 529 + 676 + 441 + 484 = 3860 ny = 8 samples (y) = Mean of y, therefore 174/8 = 21.75 22.0 (3.s.f) (y)2 = 222 which is 484 (3.s.f) Inserting the values of the Tea tree oil and the chlorhexidine into the statistical T- test equation T = (x) - (y) (?x2/nx) - (x)2 + (?y/ny) - (y)2 nx - 1 ny-1 Inserting the values for the Tea tree oil Step 1 2775/ 8 324 347 (?x2/nx) - (x)2 nx - 1 7 ?x2= 2775/8 =346.875 347 (3.s.f) nx = no. of samples which is 8 (x) = 18.0 (x)2 = 324 Inserting the values for the Chlorhexidine Step 2 3860 / 8 484 482 (?y2/ny) - (y)2 ny - 1 7 ?y2= 1924 ny = no. of samples which is 8 Therefore 3860/8 =482.5 (y) = 22.0 (y)2 = 484 It is now possible to add the values of the tea tree oil and the chlorhexidine. (?x2/nx) - (x)2 + (?y2/ny) - (y)2 nx -1 ny - 1 347 - 324 + 482 - 484 7 7 347 - 324 = 23 482 - 484 = -2 23 + -2 = 21 7 7 Step 3 It is now possible to add both the results for values for Tea tree oil and chlorhexidine into the statistical T -test equation. ...read more.


Chlorhexidine on the other hand seems to work better on the gram-positive bacteria rather than on the gram-negative bacteria. " Treatment with Chlorhexidine on gram positive bacteria (Bacillus substilis) had a mean diameter of 22.0mm, whereas on the gram negative bacteria (E.coli) the mean diameter value of 15.0mm" This resistance arises because E.coli are able to produce specific enzymes for destroying the active properties of chlorhxidine glucanoate and due to the plasmids that may be carried. These plasmids are extra chromosomal pieces of DNA, which code for the resistance to a number of antibiotics. Resistance can be developed within the bacterium's chromosome. This happens because gram-negative bacteria are generally more resistant to the antiseptic properties of chlorhexidine. The main active property in chlorhexidine, 'chlorhexidine glucanoate' is slow acting antiseptic and its chemistry is not as complex as terpine-4-ol, resulting in the development of resistant strains of an organism in otherwords enables development of resistant enzymes in E.coli. The plasmid DNA carries the resistance and can developed within the bacterium's chromosome. The antimicrobial agents such as the chlorhexidine glucanoate naturally act selectively on specific types of organisms or on specific strain of organism. This action of selectivity favours the survival of organisms that either escape full-strength assault, or those capable of resisting the assault. It seems from the practical investigation I have carried that the E.coil (gram negative) was able to resist full assault of the chlorhexidine, while the Bacillus subtillus (gram positive) endured the full effect of the active ingredient in chlorhexidine and produced less resistance to the antiseptic. Using scientific knowledge, It could be that the outer membrane of gram-negative bacteria (E.coli) however acted as a barrier (layer of peptidoglycan is protected by a complex membrane including lipopolysaccarides, lipids and phospholipids, which limits the entry of many chemically unrelated types of antibacterial agents such as the chlorhexidine gluconate. This could also be a partial reason why the chlorhexidine worked better in gram-positive bacteria, than that of the gram negative. Donald Blankson-Hemans A2 Biology coursework ...read more.

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