Ethanol_17novx - aos-hci-2012-research-program

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22 Φεβ 2013 (πριν από 4 χρόνια και 6 μήνες)

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Done by:

Aman

Mangalmurti


Kara Newman

Leong Qi Dong

Soh Han Wei


Rationale

Depletion of non
-
renewable fossil fuels
due to excessive
consumption as a
source of energy

Conversion of
renewable sources,
e.g. organic wastes, to
fuel ensures continual
energy supply

Potential for
producing ethanol
from fruit peel wastes
through fermentation
by microorganisms

Ethanol as a
renewable, alternative
energy source

Rationale

Heavy metal water
contamination of
water is rampant in
many countries.

Heavy metal ions
accumulate inside
organisms and cause
adverse health effects

Biosorption

in
removal of heavy
metal ions by fruit
peel wastes

Literature Review


Demand for renewable energy resources has increased
due to increased prices for oil and concerns about
global warming
(Wilkins , Widmer & Grohmann,
2007)


Production of ethanol by
Saccharomyces cerevisiae

from


Mango fruit processing solid and liquid wastes (Reddy,
Reddy & Wee, 2011)


Pineapple waste (Hossain & Fazliny, 2010)

Literature Review


Industries such as electroplating, mining and paint
contribute to heavy metal pollution in the ambient
environment


Heavy metal ions that pollute water include antimony,
copper, lead, mercury, arsenic and cadmium

(
US
Environmental
Protection
Agency, 2011)


Methods of removal of ions include chemical
precipitation and solvent extraction


Expensive and low efficiency at low metal ion
concentrations



Objectives

To prepare extracts of fruit peel for
ethanol fermentation

To determine which fruit peel gives
highest ethanol yield from the
fermentation of fruit peel extract

To determine which fruit peel waste
gives rise to maximal adsorption of
heavy metal ions of Cu
2+
,
Cu
3+

ions

Hypothesis


Ethanol yield from fermentation of the banana peel
would be higher than that of the mango peel


Zymomonas

mobilis

produces more ethanol during
fermentation as compared to
Saccharomyces

cerevisiae


The mango peel would adsorb heavy metal ions better
as compared to banana peels

Experimental outline

Preparation of fruit peel extract

First ethanol fermentation

Heavy

metal ion adsorption for
Copper(II), Copper (III) ions

Second ethanol fermentation after
treatment of peel residue with
cellulase

Variables

Constant


Temperature of
growth of organisms
(30

C)


Initial concentration
of heavy metal ions
(50 ppm)

Independent


Fruit peels used
(AOS: banana, HCI:
mango)


Organism used (
S.
cerevisiae
,
Z.
mobilis
)


Heavy metal ions
(Cu
2+
,Cu
4+
)

Dependent


Initial
concentration of
reducing sugars in
fruit peel extracts


Ratio of ethanol
yield to initial
sugar
concentration


Final ethanol yield


Final
concentration of
heavy metal ions

Apparatus & Materials

Apparatus

Materials


Blender


Sieve


Boiling water bath


Spectrophotometer cuvettes


Spectrophotometer


Centrifuge


Glass rod


Hot Plate


Incubator


Dropper


Sieve: 0.25mm (60 Mesh)


Shaking
incubator


Fractional
distillator


Test tubes


Filter funnel


Filter paper


Beaker


Volumetric Flask


Colorimeter



Quincy Lab Model 30 GC hot
-
air
oven


Measuring cylinder


Magnetic stirrer


Rotary mill



Mango Peel


Banana Peel


Deionised

water


Dinitrosalicylic

acid (DNS acid)


Zymomonas

mobilis


Saccharyomyces

cerevisiae


Glucose
-
yeast
medium (
Yeast malt extract
broth)


sodium alginate medium


calcium chloride
solution


sodium chloride solution


acidified potassium chromate
solution


Cu
2+
ion
solution


Cu
4+
ion
solution


MgSO
4
∙7H
2
O 0.1 and (magnesium sulfide
hydrate)


KH
2
PO
4

0.1 (potassium phosphate)



cellulase




Extraction of sugars from fruit peels

30 g of fruit peels are
blended in 300 ml of
deionised water
using a blender.

The liquid is passed
through a sieve to
remove the residue.

Determination of sugars in extracts

To 0.5 ml of extract,
0.5 ml of DNS
(
dinitrosalicylic

acid) is added.

The mixture is left in
a boiling water bath
for 5 minutes.

4 ml of water is then
added.

The samples are placed in
spectrophotometer cuvettes and
the absorbance is taken at 530 nm
using a spectrophotometer.

The concentration of reducing
sugars in
μmol
/ml is read from
a maltose standard curve.

Growth
of
Z.
mobilis

Z.
mobilis

cells are inoculated in 20 ml GY
medium (2% glucose, 0.5% yeast extract) and
incubated at 30
°
C for 2 days with shaking.

Immobilisation

of cells

The Z.
mobilis

preculture

and S.
cerevisiae

preculture

are

centrifuged
at 7000 rpm for 10
minutes and the cell
pellets are
resuspended

in
7.5 ml of fresh GY
medium.

The absorbance of the
cultures are taken at
600 nm.

7.5 ml of 2% sodium
alginate is added to
the cell suspension
and mixed well.

The mixture is
dropped into 0.1 mol
dm‐3 calcium chloride
solution to form Z.
mobilis alginate beads.

The beads are rinsed
with 0.85% sodium
chloride solution.

Growth of
S.
cerevisiae

S.
cerevisiae

cells are inoculated in 50 ml YM
broth medium with the pH adjusted to 5.6 and
incubated at 35
°
C for 1 days with shaking,
before being concentrated in a refrigerated
centrifuge at 10, 000 rpm.

Ethanol fermentation by immobilized
Z.
mobilis

cells

200 beads are added
to 50 ml waste
extract.

A control is
prepared in which
200 empty alginate
beads are added to
the same volume of
waste extract
instead.

All the set‐ups are
incubated with
shaking at 30
°
C for 2
days for ethanol
fermentation to
occur.

The beads are then
removed and the
extracts are distilled
to obtain ethanol.

Ethanol fermentation
by
S.
cerisiae


To be added








Back

Determination of ethanol yield with the
dichromate test

2.5 ml of acidified
potassium
dichromate solution
is added to 0.5 ml of
distillate in a ratio
of 5:1.

The samples are
placed in a boiling
water bath for 15
minutes.

The absorbance is
measured at 590 nm
using a
spectrophotometer,
and the
concentration of
ethanol is read from
an ethanol standard
curve.

Adsorption of heavy metal ions

Desiccate fruit peel
residue, (put the residue
in the hot air oven and
dry them at 60 degrees
for 23 hours)

Using a rotary mill to
grind desiccated residue

Sieve to 0.25 mm particle
size.

Add residue powder to
50ppm Cu
2+
solution
.

Allow solution to set for
20 min, preferably at
100rpm to increase
contact time

Repeat for Cu
4+

Determination of final ion
concentration

Allow solution to set for
20 min, preferably at
100rpm to increase
contact time

Remove fruit product,
by filtering the
suspension

Using a copper reagent,
the remaining
concentration of
copper ions will be
found

Treatment of residue with
cellulase

Fruit peel
particles are
added into
the beaker.

50 ml water
is added to
beaker

25ml of
cellulase is
added to the
beaker.

Beaker is left
standing for
1 hour with
continuous
stirring.

The beaker
is drained
and fruit
peel is left to
dry.

Second ethanol fermentation


Identical to above


Ethanol fermentation

Determination of final ethanol
yield

2.5 ml of acidified
potassium
dichromate solution
is added to 0.5 ml of
distillate in a ratio
of 5:1.

The samples are
placed in a boiling
water bath for 15
minutes.

The absorbance is
measured at 590 nm
using a
spectrophotometer,
and the
concentration of
ethanol is read from
an ethanol standard
curve.

Applications

Cost
-
effective
method of
producing ethanol

Reduces reliance on
non
-
renewable fossil
fuels

Recycles fruit peels

Viable method in
wastewater
treatment

Timeline

Finalizing of
project details
12
-
23 Nov

1
st

round of
experiments 7
Dec
-

Mar

2
nd

round of
experiments
Mar
-

May

Final round of
experiments
and Data
Analysis May
-

Jul

Bibliography


Anhwange
, T. J. Ugye, T.D. Nyiaatagher (2009). Chemical composition of Musa
sapientum (Banana) peels.
Electronic Journal of Environmental, Agricultural and Food
Chemistry, 8,
437
-
442


Retrieved on 29 October 2011 from:

http://ejeafche.uvigo.es/component/option,com_docman/task,doc_view/gid,495



Björklund, G. Burke, J. Foster, S. Rast, W. Vallée, D. Van der Hoek, W. (2009, February
16). Impacts of water use on water systems and the environment (United Nations World
Water Development Report 3). Retrieved June 6, 2011,

from

www.unes
co.org/water/wwap/wwdr/wwdr3/pdf/19_WWDR3_ch_8.pdf


US
Environmental
Protection Agency
(
2011) .Drinking Water Contaminants. Retrieved
June 6, 2011, From

http://water.epa.gov/drink/contaminants/index.cfm




Mark R. Wilkins , Wilbur W. Widmer, Karel Grohmann (2007).
Simultaneous
saccharification and fermentation of citrus peel waste by Saccharomyces cerevisiae to
produce ethanol.
Process Biochemistry,
42, 1614

1619.


Retrieved on 29 October 2011 from:

http://ddr.nal.usda.gov/bitstream/10113/16371/1/IND44068998.pdf




References


Hossain
, A.B.M.S. &
Fazliny
, A.R. (2010). Creation of alternative energy by bio‐ethanol production
from pineapple waste and the usage of its properties for engine. African Journal of Microbiology
Research, 4(9), 813‐819. Retrieved October 27, 2011 from
http
://www.academicjournals.org/ajmr/PDF/Pdf2010/4May/Hossain%20and%20Fazliny.pdf


Mishra, V.,
Balomajumder
, C. &
Agarwal
, V.K. (2010).
Biosorption

of Zn(II) onto the surface of
non‐living biomasses: a comparative study of adsorbent particle size and removal capacity of three
different biomasses. Water Air Soil Pollution, 211, 489‐500. Retrieved October 27, 2011 from
http://www.springerlink.com/content/2028u2q551416871/fulltext.pdf


Tanaka, K., Hilary, Z.D. &
Ishizaki
, A. (1999). Investigation of the utility of pineapple juice and
pineapple waste material as low‐cost substrate for ethanol fermentation by
Zymomonas

mobilis
.
Journal of Bioscience and Bioengineering, 87(5), 642‐646.


Ban‐
Koffi
, L. & Han, Y.W. (1990). Alcohol production from pineapple waste. World Journal of
Microbiology and Biotechnology, 6(3), 281‐284.


Reddy, L.V., Reddy, O.V.S. & Wee, Y.‐J. (2011). Production of ethanol from mango (
Mangifera

indica

L.)
peel by
Saccharomyces
cerevisiae

CFTRI101. African Journal of Biotechnology, 10(20), 4183‐4189.
Retrieved October 27, 2011 from
http
://www.academicjournals.org/AJB/PDF/pdf2011/16May/Reddy%20et%20al.pdf


Isitua
, C.C. &
Ibeh
, I.N. (2010). Novel method of wine production from banana (
Musa
acuminata
) and
pineapple (
Ananas

comosus
) wastes. African Journal of Biotechnology, 9(44), 7521‐7524.


Nigam, J.N. (2000). Continuous ethanol production from pineapple cannery waste using immobilized
yeast cells. Journal of Biotechnology, 80(2), 189‐193.

Saccharomyces
cerevisiae

ATCC 24553
immobilised in k‐carrageenan