The antimicrobial ability of Xylo-oligosaccharide Essay

Chapter 4: Consequence AND DISCUSSION

Chapter 4

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RESULT AND DISCUSSION

Antimicrobial Activity

Prebiotic affect the intestine microflora ( probiotic bacterium ) , by conveying the much desired effects such as diminishing the enteric pH, production of SCFA and vitamins and immune activation ( Schley and Field, 2002 ) . It is besides been considered that prebiotic/probiotics can modulate growth/activity of infective bacteriums by virtuousness of their antimicrobic activity.

In the present survey, the repressive consequence of different infusions ( viz. Water and Methanol: Water ( 1:1 ) ) of Xylo-oligosaccharides and its purified constituents ( impersonal oligosaccharide and Acidic oligosaccharides ) for bacterial and fungous strains. The antimicrobic activity was evaluated by utilizing agar good diffusion method and micro dilution method summarized in Table 1-2. The activity was quantitatively estimated on the footing of suppression zone and their activity index was besides calculated along with minimal repressive concentration ( MIC ) .

Measurement of antimicrobic activity utilizing Agar good diffusion Method

The antimicrobic ability of Xylo-oligosaccharide and its purified constituents was determined harmonizing to their zone of suppression against assorted pathogens and the consequences ( zone of suppression ) were compared with the activity of the criterions, viz. , Ampicillin ( 1.0 mg/disc ) , and Cyclohexamide ( ) . The consequences proclaimed that all the infusions are powerful disinfectants against all the micro-organisms studied. Among the different dissolvers infusions studied mixture of xylo oligosaccharide, H2O extractible and methyl alcohol: H2O extractible along with standard sugars ( arabinose, galactronic acid, glucuronic acid and wood sugar ) showed high to intercede grade of suppression. For all the tested microorganisms’ mixture of xylo-oligosaccharide, methyl alcohol: H2O and criterion sugar galactronic and glucuronis acid showed maximal antibacterial activity inEleucina coracana.Table no. represents the antimicrobic activity of the samples and standard sugar.In samples tested with the highest concentration used maximal suppression zone diameter was obtained inM.luteuswith diameter 25±5 millimeter 13±16mm, and 17±3 for samples severally and for criterions used 2±0.5mm, 1±0.5mm, 16±2mm and17±1.5 millimeter severally, is represented in figure no.

Sample

Concentrat-ion

Organism

M.luteus

B.subtlius

S.aureus

P.aeruginosa

E.coli

Ampere

100µg/mL

50µg/mL

10µ/mL

+++++

++++

++++

+++++

++++

++++

+++++

++++

++++

+++++

++++

++++

+++++

++++

++++

XOS

12.6mg/mL

13.5mg/mL

15mg/mL

++

++

+++

±

±

+

++

++

+++

±

±

±

+

+

++

No

3.2mg/mL

7.4.mg/mL

8.5mg/mL

+

±

±

±

±

±

±

±

AOS

0.7mg/mL

1.5mg/mL

5.5mg/mL

+

++

+++

±

+

±

+

+

±

±

+

±

±

+

Ara

1g/mL

±

±

±

±

±

Gallon

1g/mL

++++

++

++

++

++

GUL

1g/mL

++++

++

++

++

++

XYL

1g/mL

±

±

±

±

±

Table: Antimicrobial activity of Xylo-oligosaccharides and its purified constituents agar diffusion method consequences.

Amp – Ampicillin ( positive control )

XOS – mixture of xylo-oligosaccharide ( Sample control )

NOS – Neutral Oligosaccharide

AOS – Acidic Oligosaccharide

ARA – Arabinose ( Standard sugar )

GAL – Galactronic acid ( Standard sugar )

GUL – Glucuronic acid ( Standard sugar )

XYL – Xylose ( Standard sugar )

Determination of MIC values

Minimum Inhibitory Concentration ( MIC ) is defined as the highest dilution or least concentration of the sample used that inhibit growing of beings. Determination of the MIC is of import in diagnostic research labs because it assists in corroborating opposition of microorganism to an antimicrobic agent and it guides the activity of new antimicrobic agents. MIC was determined of the sample utilizing ELISA home base in microtiter Wellss. Ampicillin was used as the positive control to find the MIC against five different beings used. Sample was tested with four different dilutions and the reading was read at 600nm.

Organism

Space

Control

Concentration g/mL

0.5

1

1.5

2

2.5

M.luteus

0.057

0.215

0.168

0.060

0.059

0.058

0.057

B.subtilis

0.056

0.267

0.105

0.099

0.082

0.068

0.058

S.aureus

0.056

0.462

0.064

0.061

0.060

0.059

0.058

P.aeruginosa

0.056

0.448

0.392

0.247

0.079

0.076

0.068

E.coli

0.056

0.316

0.304

0.299

0.249

0.214

0.146

Table: MIC reading of positive control Ampicillin

Space:merely civilization media.

Control:Test being along with civilization media but without antibiotic.

Reading:All the O.D. readings were read at 600nm and subtracted from the clean reading.

Standard graphs of MIC of Ampicillin

Figure:M.luteusMIC Ampicillin graph

Figure:B.subtilius MICAmpicillin graph

Figure:S.aureus MICAmpicillin graph

Figure:Pseudomonas aeruginosaMIC Ampicillin graph

Figure:E.coliMIC Ampicillin graph

  • Sample MIC reading

Oraganism

/sample

Space

Control

Dilutions

25

50

75

100

M.luteus XOS

0.047

0.214

0.098

0.093

0.079

0.056

M.luteus NOS

0.047

0.174

0.151

0.151

0.133

0.116

M.luteus

AOS

0.047

0.179

0.176

0.136

0.094

0.099

B.subtilius XOS

0.047

0.344

0.324

0.188

0.121

0.121

B.subtilius NOS

0.047

0.267

0.457

0.358

0.359

0.104

B.subtilius AOS

0.047

0.307

0.227

0.169

0.151

0.071

S.aureus

XOS

0.090

0.757

0.405

0.207

0.173

0.168

S.aureus

No

0.089

0.723

0.469

0.456

0.332

0.293

S.aureus

No

0.089

0.782

0.496

0.476

0.465

0.438

P.aeruginos XOS

0.086

0.818

0.494

0.365

0.291

0.284

P.aeruginos NOS

0.086

0.820

0.548

0.487

0.420

0.340

P.aeruginos AOS

0.088

0.817

0.543

0.525

0.520

0.361

E.coli

XOS

0.047

0.407

0.299

0.103

0.092

0.064

E.coli

No

0.048

0.386

0.337

0.137

0.124

0.085

E.coli

AOS

0.047

0.398

0.271

0.166

0.137

0.091

Table: MIC reading of the sample used.

XOS – mixture of xylo-oligosaccharide ( Sample control ) , NOS – Neutral Oligosaccharide and AOS – Acidic Oligosaccharide.

Space: Culture media

Control: Organism along with civilization media but without sample.

All the readings were subtracted from clean reading.

  • Concentration of the sample used for the experiment.

SAMPLE / VOLUME

25µL

50µL

75µL

100µL

XOS>CONC.

370µg

740µg

1120µg

1500µg

NOS>CONC.

175µg

350µg

525µg

700µg

AOS>CONC.

125µg

250µg

375µg

500µg

Table: concentration of sample used for MIC.

For experiment carried out with three different samples different concentrations of the sample was used to look into the MIC of it against five different trial being. Due to presence of different degree of concentration of the sample there was difference amongst each concentration usage. Mixture of xylo-oligosaccharide being the high in sugar concentration, high concentration was easy to utilize, were as impersonal and acidic oligosaccharides being the purified constituents of xylo-oligosaccharide its concentration varies consequently. The ability of each of the sample used depends upon the phenolic compound nowadays in the sample. Phenolic are been reported to hold more antimicrobic activity than any other compound nowadays in the oligosaccharide composing. Phenolic acid derived functions holding long alkyl concatenation shows more antimicrobic activity. Butyl esters of the phenolic acids efficaciously inhibits the gm positive micro-organism.

  • Graph of MIC consequences of the experimental sample.
  • M.luteus

Figures: Xylo-oligosaccharide MIC graph

Figure: Impersonal oligosaccharide MIC graph

Figure: Acidic oligosaccharide MIC graph

Figures: comparing of MIC reading of sample.

  • B.subtilius

Figure:Xylo-oligosaccharide MIC graph

Figure: Impersonal oligosaccharide MIC graph

Figure: Acidic oligosaccharide MIC graph

Figures: comparing of MIC reading of sample

  • S.auerus

Figure:Xylo-oligosaccharide MIC graph

Figure: Impersonal oligosaccharide MIC graph

Figure: Acidic oligosaccharide MIC graph

Figures: comparing of MIC reading of sample

  • P.aeruginosa

Figure:Xylo-oligosaccharide MIC graph

Figure: Impersonal oligosaccharide MIC graph

Figure: Acidic oligosaccharide MIC graph

Figures: comparing of MIC reading of sample

  • E.coli

Figure:Xylo-oligosaccharide MIC graph

Figure: Impersonal oligosaccharide MIC graph

Figure: Acidic oligosaccharide MIC graph

Figures: comparing of MIC reading of sample

Sample

Organism

M.luteus

B.subtilius

S.aureus

P.aeruginosa

E.coli

XOS

1500µg

+1500µg

1500µg

+1500µg

1500µg

No

+700µg

+700µg

+700µg

+700µg

700µg

AOS

500µg

+500µg

+500µg

+500µg

+500µg

Table: Minimal repressive concentration of the sample used against the trial being

The consequence of the three sample on the trial pathogen were summarized in the tabular array. Average of the three experiment were carried out and it was antimicrobic activity of these samples was reported in the present survey. The consequence showed that sample fromElucina coracanahad antimicrobic activity against trial microbic pathogen. In conformity with surveies carried out on xylo-oligosaccharide ad its purified constituents show high activity against gm positive beings. M.luteus was found to be more sensitive amongst all were as other were relatively immune against the sample, sensitiveness of these infective bacteriums would increase with addition in the concentration of the sample as noticeable activity was reported.

It was observed that the antimicrobic activity of different sample varies from one sample to other in different research carried all about. This may be due to assorted factors like, consequence of clime, soli composing, age and flora rhythm phase, sample beginning, quality, measure and composing of the extracted merchandise and besides on the different trial pathogens used. ( Masotti, V. et.al. 2003 ) . Furthermore, different surveies found that the type of dissolver has an of import function in the procedure of pull outing.

In the present survey it was reported that gm positive bacteriums are more immune particularB.subtiliusas spores from it more immune to environmental status than any other tried bacteriums.E.coliandS.aureuswhich are already known to be multi-resistant pathogens showed low sensitiveness. The activity difference between gm positive and gram negative bacterium is due to lipopolysaccharide bed of gram naegative bacteriums in the outer membrane have a high hydrophobicity which acts as a strong permeableness barrier against hydrophobic molecules ( Smith Palmer et. al. , 1998 ) . Hydrophobic molecules can apss through cell of the gm positive bacteriums easier than the gram negative bacteriums because cell wall of the gm positive bacterium contains merely peptidoglycan ( Nikaido H. et.al. , 1985 ) .

The consequences revealed variableness in the repressive concentration of each infusion against trial pathogen.

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