Transcript File6

Fractionation of Starch
References:
1.
Chapter 14, “Commercial separation of amylose and amylopectin
from starch”, Starch Production Technology, edited by J.A. Radley.
2.
Chapter 8, “Fractionation of starch”, Austin H. Young, Starch:
Chemistry and Technology, edited by R.Y Whistler.
Amylose VS Amylopectin
• Molecular size
• Molecular arrangement
• Functional groups
Methods of Fractionation

Aqueous leaching of gelatinized granules
 Dispersion of the granule and fractionation
with complexing agents (selective precipitation)
 Fractionation by retrogradation
 Fractional precipitation (salting out)
Raw Materials
• Cereal starches have a great disadvantage as a raw
materials because they have a high content of fatty
compounds.
• Potato starch is very suitable for fractionation process, for it
contains no fatty substances. The phosphate groups, bound
only to the amylopectin molecules, increase the difference
between both starch components.
• Waxy starches contain practically no amylose, so that it is
not necessary to fractionate them.
• Amylomaize can be subjected to fractionation.
Aqueous leaching of gelatinized granules
 Leaching occurs from swelling starch granules in
water at temperatures of 57 – 100 °C. Mobile
amylose molecules diffuse out of the swallen granule
while the granule is intact.
 Most of the amylopectin remains H-bonded or
crystallized in the granule residue.
25 g starch + 25 ml cold water
Add slowly with stirring 150 ml of water at 80 °C
Maintain at 70 °C for 5 mins
Pour into 1200 ml of water at 60 °C
Stir very slowly at this temperature for 4 h
(avoid rupture of the swollen granule)
Cool, centrifuge
Ref:
Supernatant
amylose
Gelatineous deposit
amylopectin
Supernatant
Gelatineous deposit
Add methanol into supernatant
(to get 20% alcohol solution)
precipitate
Grind in mortar with
95% ethanol
centrifuge
Filter and dry
Grind precipitates in mortar
with 95% ethanol
in vacuum oven
Filter and dry in vacuum oven
Amylopectin
Amylose
 defatting starch geanules by heating in aqueous
methanol enhances leaching of the amylose. (useful for
starch which strongly resists swelling and gelatinization.
 removal of oxygen from the water reduced
degradation of starch.
 low temperature pretreatment of granules induces
further crystallization, resulting in higher yield of
extraction without granule rupture.
(for example; froze at -78 °C for 1 min)
Dispersion of the granule and fractionation
with complexing agents (selective precipitation)
• Defattation
• Dispersion
• Inclusion complex formation
• Separation of complex
Ref: Schoch T.J., 1941
Defatting procedure
Reflux the starches for several hours with 85%
methanol. After five extractions the lipids are
sufficiently removed.
Dispersion
Disperse 1-3% of starch in water at 105 – 109 °C for 2-3 h
Dispersion by chemicals
 1 N NaOH/KOH at 25 °C
 Dimethyl solfoxide at 25 °C
DMSO is a powerful H-bond acceptor, thereby breaking
association H-bond in polysaccharide and in water.
CH3
Starch-OH ----- -O – S+
CH3
Inclusion complex formation
“precipitating the amylose as a complex with
complexing agents (organic compounds)”
Starch + Complexing agent
K =
Complex
[complex]
[starch] [complexing agent]
K = complex formation constant
Kam
>>
Kamp
Complexing agents
Aliphatic Alcohols; isopropyl, n-butyl, isoamyl, methyl
Lower aliphatic ketone
Lower aliphatic fatty acids
Benzenoid derivatives having aldehyde groups
Alkyl halides
Cyclic alcohols
Phenol
Esters
FRACTIONATION OF AMYLOSE AND AMYLOPECTIN
STARCH+DMSO
BOILED, STIRRING ( UNDER N2)
ADD ETOH
MIXTURE STORED 00 C
CENTRIFUGATION
REDISSLOVED IN DMSO
CENTRIFUGE
SUPERNATANT 2 (AP )
PRECIPITATE DISPERSE IN WATER
1 PEAK
ULTRACENTRIFUGATION
SUPERNATANT + WTER +1-butanol
ADD 1-butanol
water
3-methyl-1-butanol
REFLUX 3 hr.
Cool, keep overnight at RT.
And 8 0C 24 hr.
CENTRIFUGATION
Supernatant 1 ( AP)
*1RECRYTALLISATION*
CENTRIFUGE
PRECIPITATE DISPERSE IN 10% 1-butanol 1 liter
REFLUX 10 min.
FILTERED THROUGH GLASS FILTER G-5
PRECIPITATE SUSPEND IN 10% 1-butanol
REFLUX 10 min.
REFLUX 1h.
Cool, keep overnight at RT.
And 8 0C 24 hr.
CENTRIFUGE
GROUND WITH ETOH
Reflux 1h.
Cool, keep overnight at RT. and 8 0C 24 hr.
PRECIPITATE DISPERSE IN WATER
CHECK PURITY OF AMYLOSE
GPC TOYOPEARL
HW-75 F
*2 RECRYTALLISATION*
Cool, keep overnight at RT. and 8 0C 24 hr.
WASH WITH Diethyl Ether
DRY IN VACUUM AT RT.
OVER CaCl2
AMYLOSE
Fractionation by retrogradation
A 7-9% slurry of potato starch is gelatinized at 85
°C and homogenized in a blender. Sufficient energy
is given the system to cause starch dissolution.
The mixture is then centrifuged and cooled so
that the amylose separates in the form of globules
from a liquid containing the amylopectin.
Fractional precipitation (salting out)
“industrially acceptable method”
A commercial process salted out the amylose from
potato starch with magnesium sulfate.
This method was based upon the fractional
crystallization of amylose from a 10% by weight
aqueous solution of potato starch in the presence of
10-13% magnesium sulfate.
In 1960, the Avebe Company in Holland produced 5.4
ton/day of amylose and 15.4 tons/day of amylopectin.
10% starch in 10-13% MgSO4 solution is solubilized by
heating to 160 °C
Cool down the solution
(a phase separation occurs )
The amylose separates in the form of small droplets and
the amylopectin remains in the solution
By cooling down to lower temperatures the droplets of amylose
solution retrograde rapidly, forming small particles of amylose gel.
Amylose gel is separated by centrifugation at 1000 g for 5 min
Wash out the salt by cold water
AMYLOSE
Amylopectin solution
Treat with more MgSO4 to precipitate all starch
(including amylose remained)
Ripen for 8 h at RT, the flocculent precipitate
become insoluble
Wash the precipitate with water
AMYLOPECTIN
Amylopectin fraction is less pure than the amylose fraction