The 5th PSU-UNS International Conference on Engineering and

Technology (ICET-2011), Phuket, May 2-3, 2011 Prince of Songkla University, Faculty of Engineering

Hat Yai, Songkhla, Thailand 90112

Abstract: Ethanol production from the weak skin of jelly

seeds of Palmyra Palm using Loog-Pang Kao Mhark

(Rice Cake starter) was investigated in this work. The

seed skin was crushed and boiled in a pretreatment step

to study the influence of: pretreatment temperature of 75

– 90 °C, pretreatment times of 15 – 60 min. Then ethanol

fermentation of the boiled seed skin was carried out to

study the influence of: fermentation times of 1 – 9 days

and Loog-Pang amount of 2 – 5%wt. The optimum

condition for the batch production that operated in 250

ml air-locked flasks was boiled at 80 ºC for 45 min in

pretreatment step, 5%wt of Loog-Pang for 7 days with

initial pH 5.5 at a room temperature. Under this

condition, it could provide 9.6%v/v purify of ethanol

product.

Key Words: Palmyra palm / Ethanol /Loog-Pang

1. INTRODUCTION

Ethanol (or bio-fuel) is defined as the ethyl alcohol

made from biomass (agricultural products and waste).

This renewable source is as efficient as petroleum fuel. It

releases less pollution into the air than the petroleum,

especially SO2. Using fuel from the plants is definitely a

way to preserve the environment by helping to reduce the

greenhouse effect as CO2 and NOX [1]. Thailand is one

of the countries that have a large quantity of fuel imports.

The alternative energy source, especially from

agricultural residue, will both increase farming incomes

and decrease our currency loss to foreign countries.

Ethanol can be used directly as fuel or blended with

benzene and diesel, and used as additive to gasoline

instead of methyl tertiary butyl ether (MTBE) [2].

Chemical and biological technology can be used for

producing ethanol. For the biological production, raw

materials can be divided into 3 major types: (1) sugar-

sugarcane and molasses, (2) starch-potato and rice, and

(3) fiber-wood and fruit skin [3]. The research of ethanol

production is focused on reducing production costs and

increasing energy efficiency of plant process.

Loog-Pang is a microorganism source (Yeast). It is

isolated and characterized by its morphological, genetic,

physiological and fermentation properties. It is a

traditional starter culture of alcoholic production for food

and drink industries [4]. Moreover, Loog-Pang can be

used to produce ethanol. The ethanol production from

Cassava starch was studied by using yeasts isolated from

Loog-Pang. The optimum condition was 5% (w/v)

cassava starch, 3% Saccharomycopsis sp. YCY1 at 37 ºC

with 100 rpm shaking rate. It could provide 5.72 g/l

ethanol product [5]. The ethanol fermentation of Sato for

14 days gave 11.58% and 10.66% purity of ethanol

products by using the pure culture microorganism,

Saccharomyces cereviceae TISTR 5049, and yeast cake,

respectively [6]. The ethanol production from shredded

cassava was carried out with 8 g. of Chinese yeast cake

and fermentation time of 31 days. The obtained highest

ethanol was 9.32% (v/v) [7]. The studying of ethanol

production from Cassava starch by using the selected

fungi from Tan-Koji (Loog-Pang) and Saccaromycetes

cereviseae were carried out with 6% Cassava starch. The

highest ethanol product content was achieved at 14.36 g/l

after 24 h of saccharification process (SFF) [8].

The main purpose of this work is to investigate the

possibility of ethanol production from the weak skin of

edible jelly seeds of Palmyra Palm (agricultural waste)

and to find the optimum condition for the production

using Loog-Pang Kao Mhark.

2. MATERIAL AND METHOD

2.1. Materials

The weak skin of edible jelly seeds of Palmyra palm

was obtained from agriculturist group that transforms the

Palmyra Palm product in Sathing Phra, Songkhla

province. The seed skin composition is given in table 1.

Loog-Pang was bought from a local market in

Narathiwat province, Thailand.

Ethanol production from the weak skin of

jelly seeds of Palmyra Palm by Loog-Pang

Asma Mardla1*, Sininart Chongkhong

1, Pakamas Chetpattananondh

1

1Prince of Songkla University, Faculty of Engineering, Thailand

*Authors to correspondence should be addressed via email: ma_mamoko@hotmail.com

43

Table 1.The composition of the weak skin of edible jelly

seeds of Palmyra Palm.

Component Palmyra palm

Protein 1.47%

Crude Fat 0.15%

Moisture 92.78%

Ash 0.52%

Crude Fiber 0.44%

Total Carbohydrate 5.08%

energy 27.55 kcal

2.2. Pretreatment

The seed skin was crushed to be 1 mm particle size

and it was boiled for the physical pretreatments.

Operating parameters of the pretreatments were included

the boiling time in the range of 15 – 60 min and

temperatures in the range of 75 – 90°C with a water to

the seed skin ratio of 60%wt. Then it was cooled down

until it reached at room temperature.

2.3. Fermentation

The ethanol fermentation was carried out in 250 ml

air-locked flasks. The pretreated seed skin was poured

into the flasks following by Loog-Pang powder and

nitrogen gas for helping to scavenge the air or oxygen in

the flasks. The ethanol fermentation should be the

process without oxygen. The studied operating

parameters were the quantity of Loog-Pang in the range

of 2 – 5 %wt and fermentation times in the range of 1 – 9

days under a room temperature with an initial pH 5.5.

The fermented products were passed into a fabric filter to

separate a solid phase. Then a liquid phase was

centrifuged at 5000 rpm for 15 min to obtain the clear

liquid product before ethanol content analysis by a

refractometer.

2.4 Analytical Methods

Reducing sugar was measured by DNS

(Dinitrosalicylic Acid) Method using a double beam UV-

Vis Spectrophotometer by UV-Visible ChemStation

Software (Model: HP 8453) [9].

A standard curve of glucose solution (reducing sugar)

was prepared. One gram of the pure glucose was

dissolved in distilled water to obtain the 100 ml solution.

This stock solution (10.0 g/l) was used to make seven

appropriate glucose dilutions from 0.5 – 10.0 g/l that was

added DNS reagent into each tube at 1:1 volume ratio.

The blank solution was prepared similarly to stock

solution by using distilled water. The solutions in the

tubes were boiled at 85 °C for 30 min. Then they were

cooled until reach to a room temperature and diluted to

10.0 ml with distilled water. The transmittance value was

measured at 570 nm on a spectrophotometer.

Ethanol content was determined by the Refractometer

(ATAGO-Japan, model DR-A1 Digital ABBE). It can

read the ethanol content value by eye measurement. A

standard curve of ethanol in distilled water solution was

prepared with difference eight appropriate ethanol

solutions (0 – 45 %v/v).

3. RESULTS AND DISCUSSION

3.1 Standard curves

The standard curve of reducing sugar was shown in

Fig. 1.

Fig. 1. The standard curve of reducing sugar.

The standard curve of ethanol solution was shown in

Fig. 2.

Fig. 2. The standard curve of ethanol concentration.

3.2 Pretreatment

The partial components of the thermal pretreated

products as fiber and carbohydrate are transformed into

the sugars (smaller molecules). They can be assayed in a

term of glucose (reducing sugar).

3.2.1 Effect of boiling time

Fig. 3 shows the effect of boiling time on reducing

sugar content at a boiling temperature of 85°C. It can be

seen that the boiling time at 45 min and 60 min gives the

optimum reducing sugar amount. Therefore, the boiling

time for 45 min was enough to break down the starch

44

molecule into fermentable sugars for economic reason.

Moreover, it was found that a short-time of boiling was

not enough to reduce the size of molecule in the weak

skin reasonably.

Fig. 3. The effect of boiling time on reducing sugar

content.

3.2.2 Effect of boiling temperature

Fig. 4. The effect of boiling temperature on reducing

sugar content.

Fig. 4 shows the effect of boiling temperature on

reducing sugar content for a boiling time of 45 min. The

suitable boiling temperature was 80°C that molecular

structure was readily break down into fermentable

sugars.

The pretreated product was a higher viscosity

because the pretreatment process could change the

molecule structure of carbohydrate to be an active

molecule that was easier to be hydrolyzed. This process

could be called gelatinization.

The results (Fig. 3 and 4) show that the optimum

gelatinization condition was 80°C for 45 min.

3.3 Fermentation

3.3.1 Effect of Loog-Pang amount

Fig. 5 shows the effect of Loog-Pang amount on

ethanol content. For fermentation time of 7 days, the

rapid formation of ethanol was observed when Loog-

Pang amount was higher (2-5%). However, the

maximum ethanol content could be obtained by using

5%wt of Loog-Pang.

Fig. 5. The effect of Loog-Pang amount on ethanol

content.

3.3.2 Effect of fermentation time

It can be seen in Fig. 6 that for using 5%wt Loog-

pang, the ethanol conversion was raised when the

fermentation time was increased. The fermentation time

of 7 days was suitable for this process.

Fig. 6. The effect of fermentation time on ethanol

content.

4. CONCLUSION

The ethanol production from the seed skin of Palmyra

Palm by using microorganisms from Loog-Pang Kao

Mhark could provide 9.6%v/v ethanol product. The

optimum condition was 5%wt Loog-Pang amount for 7

days with pH-5.5 at a room temperature. The ethanol

products could be purified by a distillation or

evaporation.

5. ACKNOWLEDGEMENTS

The author gratefully acknowledges the financial

support from the Graduate School of Prince of Songkla

University.

45

6. REFERENCES

[1] S. Ture, D. Uzun, and I. E. Ture, “The potential use

of sweet sorghum as a non-polluting source of

energy”, Energy., 1997, Vol. 22, pp. 17-19.

[2] J. E. McCarthy, and M. Tiemann, “CRS report for

congress. MTBE in gasoline: clean air and drinking

water issues”, 1998, Available online;

http://www.epa.gov/otaq/consumer/fuels/mtbe/crs-

mtbe.pdf. [23/12/2010]

[3] S. Abedinifar, K. Karami, and M. Khanahmadi,

“Ethanol production by Mucor indicus and Rhizopus

oryzae from rice straw by separate hydrolysis and

fermentation”, Biomass and bioenergy, 2001, Vol.

33, pp. 828-833.

[4] V. Kitpreechavanich, T. Maneeboon, Y. Kayano,

and K. Sakai, “Comparative Characterization of l-

Lactic Acid- Producing Thermotolerant Ghizopus

Fungi”, Journal of Bioscience and bioengineering.,

2008, Vol. 106, 6, pp. 541-546.

[5] K. Saelim, “Ethanol Production from Cassava

Starch by Yeasts Isolated from Loog-Pang”, Prince of

Songkla University, Songkhla, 2006.

[6] W. Kheowrod, “Comparative study on the

fermentation of Sato by mixed pure culture

microorganism and yeast cake”, Mahidol university,

Bangkok, 2002.

[7] P.Dumrongmanee, “Comparison of the amounts of

Chinese yeast cake for ethanol production using

shredded cassava”, Chiang Mai University, Chiang

Mai, 2001.

[8] T. Y. Amornpitak, “Ethanol Production from Cassava

Starch by Selected Fungi from Tan-Koji and

Saccaromycetes cereviceae”, Mahasarakham

University, Mahasarakham, 2010, pp. 84-88.

[9] G. L. Miller, “Use of dinitrosalictlic acid reagent for

determination of reducing sugar”, Anal. Chem., 1959,

Vol. 31, pp. 426-428.

46