International Journal of Biotechnology and Bioengineering Research

Volume 1 Number 2 (2010) pp. 213–218

© Research India Publications
http://www.ripublication.com/ijbbr.htm

Banana Peel Waste as Substrate for Ethanol

Production

Latika Bhatia1 and Shirish Paliwal2

1,2
Department of Applied Microbiology and Biotechnology
Dr. Hari Singh Gour Central University, Sagar, Madhya Pradesh, India.
1
E-mail: latikabhatia1@yahoo.co.uk

Abstract

In recent years, potential efforts have been directed towards the utilization of
cheap renewable agricultural resources, such as banana peel waste as
alternative substrate for ethanol production. In this extend we conducted study
on potential of alcohol production of two types of yeast obtained from MTCC,
Chandigarh. Various parameters were studied which included the effect of
nutrients, inoculum level and temperature on ethanol production capacity of
both the yeasts. It was found that inorganic nitrogen sources gave good yield
of ethanol, at inoculum level of 1% wt/vol, and at incubation temperature of
30oC with Saccharomyces cerevisiae.

Keywords: Banana Peel, Acid hydrolysis, Saccharomyces cerevisiae, MTCC,

Inoculum, Pachysolen tannophilus.

Introduction
Recently the demand of ethanol has increased considerably because of the use of
gasohol in addition to their applications in industries which needs production of
alcohol on large scale [1]. Ethanol is gaining momentum as a viable fuel source due to
recent fluctuations in market of conventional fossil fuels. In addition to its common
pharmaceutical and beverage uses, ethanol has been used as a fuel additive, gasoline
enhancer and even as an alternative fuel source [2, 3].
Ethanol production is usually accomplished by chemical synthesis of
petrochemical substrates and microbial conversion of carbohydrates present in
agricultural products. Owing to the depleting reserves and competing industrial needs
of petrochemical feed stocks, there is a global emphasis in ethanol production by
fermentation process [4]. Ethanol production through fermentation may provide an
economically competitive source of energy by its incorporation into gasoline.
214 Latika Bhatia and Shirish Paliwal

Large amount of renewable biomass are available for conversion to liquid fuel,
ethanol. Ethyl alcohol can be made of three main, abundant renewable feedstock
sources, saccharines, starch materials and cellulosic materials. Low value agricultural
residues which can easily be converted to fermentable sugars, agricultural by-products
of starch industry such as potato pulp and from raw starch hydrolysate are attractive
resources for economical production of ethanol [5].
Banana (Musa sapientum) is the second largest produced fruit after citrus,
contributing about 16% of the world’s total fruit production. 27% of world’s banana
production is contributed by India alone. Banana peel is a rich source of starch (3%),
crude protein (6-9%), crude fat (3.8-11%), total dietary fibre (43.2-49.7%), and
polyunsaturated fatty acids, particularly linoleic acid and α- linolenic acid, pectin,
essential amino acids (leucine, valine, phenylalanine and threonine) and
micronutrients (K, P, Ca, Mg). Banana peels are also a good source of lignin (6-12%),
pectin (10-21%), cellulose (7.6-9.6%), hemicelluloses (6.4-9.4%) and galactouroninc
acid [6]. Banana peel, an agro waste can be used as a substrate for ethanol production
owing to its rich carbohydrate, crude proteins and reducing sugars. Moreover, banana
peels are affordable and renewable low cost raw material which makes it potential
feedstock for ethanol production.
In the present work, ethanol production was studied, using banana peel as a
substrate and employing Saccharomyces cerevisiae MTCC 178 and Pachysolen
tannophilus MTCC 1077.

Material and Methods

Pretreatment of Banana Peels
Fresh banana peels were cut into small pieces of sizes between 4cm to 5 cm. The cut
pieces were next washed with water to remove undesirable particles followed by
drying operation in hot air oven for a period of 24 h at 65oC. The peels were next
grounded using mortar-pastle to powdered form. In a test tube, 2g of ground sample
was taken and 10 ml distill water was added to it and the solution was then boiled for
10 min. Next, a 5ml sample of this liquid was taken and the glucose content was
estimated based on standard techniques using a UV spectrophotometer. Sample
contained maximum initial glucose concentration of 0.32 wt/vol % or 3.2g of initial
sugar per 100g of peel. This sample of banana peel was then used for the acid
hydrolysis experiment.

Acid Hydrolysis
Since banana peels contain both starch and sugar, in order to increase the yield of
ethanol, the starch was hydrolyzed to convert it to sugar and afterwards, the total
fermentable sugar was converted to ethanol. 5 g of ground peels was taken in a 150
ml conical flask and 50 ml 4 (N) HCl was added to it. The solution was heated in a
water bath for 10min, 20min, 30min, 60min, 75min and 90min. Table 1 shows the
amount of glucose liberated in different interval of time, revealing the comparative
increase in glucose level after acid hydrolysis. Temperature maintained was 80ºC to
100ºC. The hydrolyzed sample was cooled to room temperature. The cooled sample
was filtered using a filter paper.
Banana Peel Waste as Substrate for Ethanol Production 215

Table 1: Glucose concentration after hydrolysis.

S .No. Time (min.) Glucose (Wt/v %)

1 10 min 0.35
2 20 min 0.72
3 30 min 0.78
4 60 min 0.80
5 75 min 0.81
6 90 min 0.81

Production of alcohol from banana peel

The hydrolysed sample was cooled to room temperature. The cooled sample was
filtered using a filter paper and the pH of the clear sample was adjusted to 4-5 by
adding requisite quantity of 10 (N) NaOH and 4 (N) HCl. The total volume was made
up to 100 ml.
Various parameters were studied by us. We used different nutrients, inoculum
levels (% w/v) and temperatures (oC). Two different yeasts were employed as the
inoculant. The fermentation was carried out in 250 ml conical flasks containing
100ml hydrolysates supplemented with nitrogen (0.3% ammonium sulphate or urea),
phosphorus (0.15% potassium di-hydrogen phosphate) and growth factors (0.5% yeast
extract) or yeast nitrogen base (0.67%) or yeast extract (0.5%) and peptone (0.5%).
The solution was then sterilized in an autoclave and kept in cool place after cooling.
Saccharomyces cerevisiae MTCC 178 and Pachysolen tannophilus MTCC 1077
were procured from Microbial Type Culture Collection (MTCC), IMTECH,
Chandigarh. Characteristics of both these organisms are given in Table- 2

Table 2: Characteristics of MTCC organisms.

Characteristics Saccharomyces Pachysolen tannophilus

cerevisiae
MTCC NO 178 1077
Growth 5 (YPED) 6 (Malt Yeast Agar)
medium
Temperature 30 25
Incubation 48 hrs 48 hrs
Time
Subculture 60 days 30 days
Growth Aerobic Aerobic
condition
Special feature Distillery yeast Production of ethanol from D-xylose, D-
(Daurala strain), galactose and glycerol. Converts wheat
production of ethanol straw, cellulose/hemicellulose to ethanol
216 Latika Bhatia and Shirish Paliwal

The yeast cultures were maintained on growth medium 5 and 6 (MTCC)

respectively, which had the following composition-

Composition YPED Composition Malt Yeast Agar

Yeast extract - 3.0 g Yeast extract - 3.0 g
Peptone - 10.0g Peptone - 5.0g
Dextrose - 20.0g Glucose - 10.0g
Agar - 15.0g Malt extract - 3.0g
Distilled water - 1.0L Agar - 20.0g
Distilled water - 1.0L

Colonies of revived culture so obtained can be seen in Figure 1. The biomass of

yeast after growth for 18 h at respective 30oC and 25oC for Saccharomyces cerevisiae
MTCC 178 and Pachysolen tannophilus MTCC 1077 in a medium containing 60.0g
sucrose, 5.0g yeast extract and 5.0g peptone L-1 was centrifuged at 5000 rpm for 15
min and inoculated into the hydrolysate at a concentration of 0.5-1.5% (w/v) and
flasks were incubated at temperatures(oC) of 30, 37 and 40 under stationary
conditions and samples were analyzed for ethanol content colorimetrically [7].

Results and Discussion

The acid hydrolysate is rich in sugars but yeast nutrients are required for efficient
fermentation to take place. The recovery of ethanol was highest with yeast nitrogen
base followed by ammonium sulphate, di-hydrogen phosphate and yeast extract
(Table 3). The inorganic nitrogen source proved better among organic and inorganic
nitrogen source supplemented to the hydrolysate.
Banana Peel Waste as Substrate for Ethanol Production 217

Table 3: Effect of nutrients, inoculum size and temperature on ethanol production

from acid hydrolyzed banana peel.

Parameters Ethanol produced (gL-1)

Saccharomyces cerevisiae Pachysolen tannophilus
MTCC 178 MTCC 1077
Nutrients
Yeast nitrogen base 29.2 28.7
Peptone+ Yeast Extract 24.6 21.5
Ammonium sulphate+ 28.5 22.6
Potassium di hydrogen
Phosphate+yeast extract
Urea+ Potassium di hydrogen 24.0 20.6
Phosphate+yeast extract

Inoculum level (% w/v)

0.5 25.3 24.2
1.0 30.1 28.3
1.5 30.0 29.2

Temperature (0C)
30 32.3 29.2
36 22.2 22.6
40 12.4 15.2

Effect of variable inoculum levels of both the yeast in the fermentation were
studied. One percent inoculum of Saccharomyces cerevisiae MTCC 178 and 1.5%
inoculum of Pachysolen tannophilus MTCC 1077 was found optimum for
fermentation. This shows that the cell number added was sufficient to produce highest
amount of alcohol in 48h at 30±2oC. The variation in fermentation temperature (30-
40oC) was checked for alcohol production under lab conditions. The optimum
temperature for ethanol fermentation of hydrolysate by yeast was 30±2oC. The
alcohol production declined about 31% at 37oC. This showed that increase in
temperature had an adverse effect on alcohol production. Saccharomyces cerevisiae
MTCC 178 is a better producer of ethanol in various conditions provided. However
efficiency could be increased by various strain improvement methods.

Conclusion
Banana is a major cash crop of this country generating vast agricultural waste after
harvest. Banana peel is a cheap source of alcohol because it is a fruit/agro waste. It
can be used as a base material for alcohol production. Shredding of banana peels is
218 Latika Bhatia and Shirish Paliwal

easy and drying temperature is not high. Strain improvements can help in better yield
of alcohol, making the large scale production more economic and more feasible.

Acknowledgement
We would like to express our thanks to Head of the department, Department of
Applied Microbiology and Biotechnology, Dr. Hari Singh Gour Central University,
Sagar, for giving an opportunity to perform this set of work in the laboratory of this
department.

References
[1] Wheals, A.E., L.C. Barso, D.M.G. Alves, H.V. Amorim, 1999. Fuel ethanol
after 29 years. Trends Biotechnology, 17: 482-487.
[2] Ogbonna J.C., H Mashina, H. Tanaka, 2001. Scale up of fuel ethanol
production from sugar beet juice using loofa sponge immobilized bioreactor.
Bioresour Technol, 76: 1-5.
[3] Davidson B.H., Scot C.D. 1988. Operability and feasibility of ethanol
production by an immobilized Zymomonas mobilis in a fluidized bed
bioreactor. Appl Biochem Biotechnol, 18: 19-34.
[4] Brooks A.A. 2008. Ethanol production potential of local yeast strains isolated
from ripe banana peels. African journal of Biotechnology, 7(20): 3749-3752.
[5] Satish Babu R., S. Rentala, M.L. Narsu, Y. Prameeladevi, D.G. Rao, 2010.
Studies on Ethanol production from spoiled Starch rich vegetables by
sequential Batch fermentation. International Journal of Biotechnology and
Biochemistry, 6(3): 351-357.
[6] Mohapatra D., S. Mishra, N. Sutar, 2010. Banana and its by-product
utilization: an overview, Journal of Scientific & Industrial Research, 69: 323-
329.
[7] Caputi P., J.M. Vede, T. Brown, 1968. Septrophotometric determination of
chromic complex formed during oxidation of ethanol. Am J Enol Vitic 19:
1601-1665.