Utilization of the field experiment results of University of Debrecen in the development of maize-based bioethanol production

Maize is currently the single raw material of bio-ethanol production in Hungary. The aim of our examinations is the observation of yield and nutritional characteristics of commercial maize hybrids in Hungary from the aspect of efficient bio-ethanol production. We set up a randomized block field trial. We determined the starch content and starch yield (t ha-1) of the 51 maize hybrids involved in the field trial. In laboratory conditions, we examined the amylose and amylopectin ratio and the amount of resistant starch of the selected 20 maize hybrids. According to our results, there is a significant difference between the starch yield the amylose component of the starch content and resistant starch of the examined maize hybrids. Our studies reveal that maize as a raw material must be selected based on the cultivation objectives. If the objective is bio-ethanol production, detailed knowledge of starch content is necessary. There is a significant difference among commercial maize hybrids in Hungary in terms of characteristics determining the producible amount of bio-ethanol.


Utilization of the field experiment results of University of Debrecen in the development of maize-based bio-ethanol production
Rátonyi Tamás -Nagy János -Harsányi Endre

INTRODUCTION
Maize is currently the single starting material of bio-ethanol production in Hungary.The cultivation conditions of Hungary are perfect for maize production, maize is grown on the quarter of cultivable land (1.2 million ha).Most of the current research is aimed towards improving the efficiency of maize-based bioethanol production.Conversion efficiency to produce ethanol by fermentation is usually 90-95%.The reasons for the imperfect conversion efficiency may include incomplete hydrolyses of starches (Wu et al., 2006).The most important measure of value of maize-based bio-ethanol production is starch content and starch yield of the raw material.Amylose and amylopectin represent 98 to 99% starch dry weight in maize kernel (Sharma et al., 2010).The ratio of amylose/amylopectin is essentially important, because it determines the gelificiation characteristics within the process of saccharification.In the course of production, alphaamylase and amyloglucosidase enzymes are used in most cases; therefore it is important to know the content of starch resistant to alpha-amylase.

MATERIAL AND METHODS
The objective of this study was to characterise maize starch content for bio-ethanol production purpose.We set up random block layout maize field experiment and starch yield (t ha -1 ) of maize hybrids were investigated in 2011.Maize grain samples were collected before harvest.
The laboratory examinations of maize grain samples were done at the Institute of Land Utilisation, Technology and Regional Development, Centre for Agricultural and Applies Economic Sciences, Debrecen University.After sample preparation starch content of maize hybrids were evaluated by Foss InfratecTM 1241 Grain Analyzer.The measurements are based on the fact that the main components in the grain (maize), such as protein, moisture, starch, fat, absorb electromagnetic radiation in the near-infrared region of the spectrum (570-1100 nm).The Near Infrared Transmission (NIT) instrument measures the transmission absorption, the main components of whole maize grain samples are determined by calibration curves, the accuracy of prediction could be comparable to chemical methods (Wei et al., 2005).
Amylose/amylopectin ration and resistant starch (%) content of 20 maize hybrids were determined in the laboratory.Amylose/amylopectin ratio and resistance starch of the maize starch content were determined according to the protocol of Megazyme International Ireland Ltd.Maize starch was solubilized in water by dimethyl-sulfoxide (DMSO).Concanavalin A precipitation of amylopectin was carried out.
Then the amylose determined by glucose-oxidase enzyme after treatment with -amylase and amyloglucosidase.The absorbance of created glucose from amylose was detected at 510 nm.To determine of resistant starch firstly we solubilized and hydrolized the non-resistant starch with enzymes (-amylase and amyloglucosidase).Then the starch resistant to -amylase exist as pellet.This pellet were solubilized by 2 mol potassium-hydroxide on ice, the next steps were the neutralization and the hydrolization.The resulting glucose was defined by glucose-oxidase enzyme at 510 nm.Statistical analysis package of SPSS 14.0 for Windows statistical software was used.

Starch content of the maize hybrids
The kernel of maize (Zea mays L.) is mainly composed of starch.The starch content is genetically determined parameter varied between very narrow ranges.Researchers were concluded that ethanol production of maize is not exclusively dependent on starch content and that the chemical structure of starch (Dien et al., 2002;Sing et al., 2005).Starch content of the kernel alone cannot predict the ethanol production of maize.

Amylose ratio in starch of the maize kernels
Starch in maize kernels can be separated into two fractions: amylose and amylopectin.Maize hybrids differ for amylose to amylopectin ratios.Amylose content has a significant effect on ethanol fermentation efficiency.Amylose content negatively correlated to bioethanol concentrations from starch, conversion efficiency decrease as the amylose content increased, however authors concluded that there was no significant effects on fermentation efficiency when amylose content was less than 30% (Wu et al., 2006;Sharma et al., 2010).
Amylose making up 12.2-30.71%,the component, amylopectin makes up 69.29-87.8% of the starch content of the investigated maize hybrids (Figure 3).The difference amongst the amylose content of the maize hybrids was significant; we found twofold differences between the maize hybrids.starch resulted in lower conversion of starch into sugars and lower final ethanol production (Sharma et al., 2010).The amount of resistant starch is depends on the genotypes of raw material.Significant difference can be found between the resistant starch content of the examined maize hybrids.The average resistant starch content of the maize kernels of the maize hybrid in the experiment was 4.06%.We measured a 5.93% difference between the maize hybrids with the highest (8.03 %) and the lowest (2.1%) resistant starch content (Figure 4).Differences amongst resistant starch of maize kernels reach fourfold value.

Correlation analysis
Relationship between starch content and the amylose content and between starch content and resistant starch content in the kernels of the investigated maize hybrids could not be found.The amount of starch content did not influence the amount of amylose and resistant starch.Total starch content had a weak correlation (R 2 = 0.11 and 0.08) with amylose content and resistant starch.
The amylose content and resistant starch content had a moderate negative correlation (R 2 =0.51).Resistant starch contents decrease as the amylose content in starch increases (Figure 5).

CONCLUSIONS
Our studies reveal that maize as a raw material of bio-ethanol production shall be selected based on the cultivation objectives.If the objective is bio-ethanol production detailed knowledge of starch content, starch yield, amylose/amylopectin ratio and resistant starch content are necessary.There is a significant difference among commercial maize hybrids in Hungary in terms of characteristics determining the producible amount of bio-ethanol.Significant difference can be found between the starch content, starch yield, amylose ratio and resistant starch of the examined maize hybrids.Starch content of the maize had a weak correlation with amylose content and resistant starch.Moderate negative correlation can be found between amylose content and resistant starch, as the amylose content in the starch increase in the maize kernels the resistant starch content decrease.

Figure 5 :
Figure 5: Correlation between amylose and resistant starch of the investigated maize hybrids