BIOSURFACTANTS

SUBATHRA.S

BIOSURFACTANTS

• Biosurfactants are proteins which present potential application in theenvironment protection, organic substances treatments and oil recoveries.

• Microbial surface active molecules able to reduce superficial andinterfacial tensions either in water solutions or in mixtures of hydrocarbons.

• Low toxicity, • high biodegradability,• environment compatibility• and specific activity in extreme conditions. • when comparing with chemical surfactants.

Screening Organisms

• Chroococcus minor BDU 20321• Synechococcus elongatus BDHKU 33801• Synechococcus elongatus BDU 171741• Oscillatoria laetevirens BDU 10143• Oscillatoria formosa BDU40261• Oscillatoria willei BDU 130481• Phormidium valderianum BDU 43131• Phormidium valderianum BDU140842• Phormidium tenue BDU 46241• Nostoc calcicola BDU 40302

CTAB TEST-Nostoc calcicola BDU 40302

SCRENNING METHODS

• 1 EMULSIFICATION INDEX(%)• 1ml culture supernatant+4mlwater and 6ml of oil.• Mixed well• Leave it 24 hrs• Height of emulsion formed(cm)Х100

Total height(cm)

Organisms Petrol Castor oil Crude oil Gingelly oil Neem oil Diesel Sunflower oil

Synechococcus elongatus BDHKU 33801

11.4 16 6.9 9 7.1 5.7 4.1

Synechococcus elongatus BDU 141741

28 18 10 23.9 5.7 8.5 16

Nostoc calcicola BDU 40302

51.2 38 28.5 12 12.6 9.5 8.9

Emulsification Index(%)

Emulsification assay

• 3ml culture supernatant+1ml oil

• Mixed well.

• Leave it for 1hr

• OD was measured at 400nm.

EMULSIFICATION ASSAY

Emulsification assay

• 1ml of Nostoc calcicola BDU 40302 culture supernatant .

• 7.5 ml of Tris- Mg and 1ml of oil was added.

• Optical density was measured at 540nm.

Emulsification assay for Nostoc calcicola BDU40302

OIL SPREAD METHOD

• Petri plate filled with water and covered with oil.• Then supernatant was added.• Clear zone was formed on the oil.• Nostoc calcicola BDU 40302 was displaced the diesel.

Concentration of Biosurfactant

• Extraction with Diethyl ether in the ratio of 1:3 volume.

• The organic layer was taken and dried in the rotary vacuum.

• PURIFICATON OF BIOSURFACTANT

• Culture supernatant was taken and made up to pH 2 by using Conc. HCL and incubate for over night, precipitate was formed.

• The precipitate was taken and carbohydrate test was done.

Nostoc calcicola Supplemented with Petrol

Control 1% 5% 10% 25%

Nostoc calcicola Supplemented with Petrol

Control 50% 75% 100%

Emulsification Index(%) for petrol supplemented Nostoc culture

Concentration of oil Emulsification index(%)

Control 0.0

1% 4

5% 6.8

10% 11

25% 21

50% 45

75% 33

100% 0.6

Emulsification Index for Nostoc calcicola with petrol

Control 1% 5% 10% 25%

Emulsification Index for Nostoc calcicola with petrolControl 50% 75% 100%

Nostoc calcicola supplemented with Petrol For seven days

Nostoc calcicola BDU 40302 Supplemented with Petrol for seven days

Growth Curve For Nostoc calcicola BDU 40302

Growth Curve For Nostoc calcicola BDU 40302

Nostoc calcicola Supplemented with Crude oil For seven days

Control 1% 5% 10%

Nostoc calcicola Supplemented with Crude oil For seven days

Control 25% 50% 75% 100%

Nostoc calcicola BDU 40302 supplemented with crude oil

Nostoc calcicola BDU 40302 supplemented with crude oil

• Thin Layer Chromatography• To find –Carbohydrates, Lipids.• To Isolate DNA • Polymerase Chain reaction and • Sequencing.