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7/23/2019 Project Proposal - No Ref

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David Curtis Mintah 10420660

Assessment of the minimum inhibitory concentration of essential oils from

Ghanaian spices on Escherichia coli, Listeria spp, and Klebsiella spp.

INTRODUCTION:

It has been estimated that as many as 0! of people in industriali"ed

countries su#er from a foodborne disease each year and in 2000 at least t$omillion people died from diarrheal disease $orld$ide% &here is therefore still

a need for ne$ methods of reducin' or eliminatin' foodborne patho'ens(

possibly in combination $ith e)istin' methods *the hurdle principle+% At the

same time( ,estern society appears to be e)periencin' a trend of -'reen-

consumerism( desirin' fe$er synthetic food additives and products $ith a

smaller impact on the environment% .urthermore( the ,orld /ealth

r'ani"ation has recently called for a $orld$ide reduction in the

consumption of salt in order to reduce the incidence of cardiovascular

disease% If the level of salt in processed foods is reduced( it is possible that

other additives $ill be needed to maintain the safety of foods% &here istherefore scope for methods of main' food safe $hich have a natural or

-'reen- ima'e% ne such possibility is the use of essential oils as antibacterial

additives%

3ssential oils *3s+ *also called volatile or ethereal oils+ are aromatic oily

liuids obtained from plant material *5o$ers( buds( seeds( leaves( t$i's(

bar( herbs( $ood( fruits and roots+% &hey can be obtained by e)pression(

fermentation( en5eura'e or e)traction but the method of steam distillation is

most commonly used for commercial production of 3s% An estimated 000

3s are no$n( of $hich about 00 are commercially important destined

chie5y for the 5avours and fra'rances maret%7esides antibacterialproperties( 3s or their components have been sho$n to e)hibit antiviral(

antimycotic( antito)i'enic( antiparasitic( and insecticidal properties% &hese

characteristics are possibly related to the function of these compounds in

plants% &he 'reatest use of 3s in the 3uropean 8nion *38+ is in food *as

5avourin's+( perfumes *fra'rances and aftershaves+ and pharmaceuticals

*for their functional properties+% &he $ellno$n use of 3 in aromatherapy

constitutes little more than 2! of the total maret% Individual components of

3s are also used as 5avourin's( either e)tracted from plant material or

synthetically manufactured%

9team distillation is the most commonly used method for producin' 3s on a

commercial basis% 3)traction by means of liuid carbon dio)ide under lo$

temperature and hi'h pressure produces a more natural or'anoleptic pro:le

but is much more e)pensive% &he di#erence in or'anoleptic pro:le indicates

a di#erence in the composition of oils obtained by solvent e)traction as

opposed to distillation and this may also in5uence antimicrobial properties%

&his $ould appear to be con:rmed by the fact that herb 3s e)tracted by


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he)ane have been sho$n to e)hibit 'reater antimicrobial activity than the

correspondin' steam distilled 3s% 3s are volatile and therefore need to be

stored in airti'ht containers in the dar in order to prevent compositional

chan'es%

3s can comprise more than si)ty individual components% Ma;or componentscan constitute up to


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environment and on foodprocessin' euipment liely also re5ects its ability

to survive in the natural environment for e)tended periods%

Klebsiella spp% are Gramne'ative nonmotile bacilli that belon' to the family

3nterobacteriaceae% &he 'enus lebsiella consists of a number of species(

includin' K. pneumoniae, K. oxytoca, K. planticolaand K. terrigena% &heoutermost layer of Klebsiella spp% consists of a lar'e polysaccharide capsule

that distin'uishes the or'anisms from other members of the family%

Appro)imately 60F


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9pectrophotometer pti"en 2120 8 K pti"en III *9hin et al%( 200+%

Dilutions( to 'et the :nal concentration ran'in' from 0 to 1000

micro'ramsHm of essential oil and e)tracts of he)ane( chloroform( ethyl

acetate and methanol in yso'eny broth medium $ere prepared in a L6$ell

microplates% .inally( 20 microliter inoculum of each bacteria strain *10

C.8Hm+ $as inoculated on to the microplates and the tests $ere performedin a volume of 200 microliter% &he plates $ere incubated at de'rees

Celsius for 24 h% &he lo$est concentration of the test samples( $hich did not

sho$ any visual 'ro$th of tested or'anisms after macroscopic evaluation(

$as determined as MIC( $hich $as e)pressed in micro'ramsHm%

Method 2B

As per the procedure follo$ed by Moreira et al%( *200=+ a'ar di#usion assay

$as used to determine minimum inhibitory concentration of the essential oils

of spices% In to 100 ml of sterile &ryptic soy a'ar $hich $as maintainin' at

4=EC( 1 ml of the prepared bacterial culture $as added to 'ive a :nalconcentration of 10 cells H ml in the medium% &he culture $as thorou'hly

mi)ed in the appropriate arti:cial medium and the plates $ere poured(

allo$ed to solidify and placed in a refri'erator for 10 minutes% /oles $ere

then punched into the a'ar to create $ells( into $hich measured volume of

100 microliter of the essential oil of spices $ere placed after sealin' the

bottom of the $ell $ith sterile a'ar of 1!% &his ensured that radial di#usion

from the $ell 'ave a clear and easily measured "one of inhibition% &he plates

$ere then incubated at EC for 24 hours% &he inhibition "ones $ere

measured after the incubation period% &his assay $as carried out in duplicate

$ith t$o replicates on each occasion% &he lo$est concentration of the testessential oil samples( $hich did not sho$ any visual 'ro$th of tested

or'anisms( $as determined as MIC( $hich $as e)pressed as 1B dilution

factor%

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