Effects of Fertilizers on Biomass, Sugar Content and Ethanol Production of Sweet Sorghum Effects of Fertilizers on Biomass, Sugar Content and Ethanol Production of Sweet Sorghum

Sweet sorghum ( Sorghum bicolor ) is a promising alternative crop for bioethanol production in developing countries. However, to extend the cultivative area of this crop, it needs to develop an appropriate growing protocol for farmers. This chapter describes the examination of different doses of fertilizers combined with manure and micronutrients, in various applied times, on biomass, sugar content and ethanol production of sweet sor- ghum. It was observed that the application of 90 N + 90 P 2 O 5 + 60 K 2 O provided maximum stem yield and optimum contents of sugar and ethanol yield, however nontreatment of any among P, P 2 O 5 and K 2 O caused significant reduction of biomass and ethanol produc- tion. Higher fertilization >90 N may provide greater productivity of this crop but it may cause lodging and economic deficit for farmers in developing countries. It was also found that the applied times of fertilization should be at 3–4 to 7–8 leaf stage. In contrast, when the fertilization was as close to the flowering stage caused remarkable reduction of stem yield and ethanol production. The supplementation of (NH4) 2 MO 7 O 2 .4H 2 O at 5 kg/ha provided an increase of 10–12 tons/ha of stem yield and a remarkable enrichment of ethanol production. Findings of this study are useful for farmers and agricultural extension- ists to promote biomass and ethanol productivity of this crop for bioethanol production. This research also highlights a greater possibility of exploiting sweet sorghum cultivation in infertile and hilly, abandoned areas for ethanol production.


Introduction
It has been estimated that there are 700 million light duty vehicles, automobiles, light trucks, SUVs and minivans on roadways around the world. These numbers may increase to 1.3 billion by 2030 and to over 2 billion vehicles by 2050, with most come from developing countries [1]. This growth will affect the stability of ecosystems and global climate as well as global oil reserves [2]. The use of biofuels can contribute to the mitigation of greenhouse gas emissions, provide a clean and sustainable energy source and may promote the agricultural income for rural poor in developing countries. Currently, biofuels are predominantly produced from biomass resources, because they are the renewable resource that could be sustainably developed, resulting in no net release of carbon dioxide and very low sulfur content and economic potential as fossil fuel prices can be increased in the future [2]. The principal sources of biofuels are liquid or gaseous fuels made from plant matter and residues, including agricultural crops, municipal wastes and agricultural and forestry by-products.
Bioethanol has a high octane number (108), broader flammability limits, higher flame speeds and higher heats of vaporization. It helps for a higher compression ratio and shorter burn time, which lead to efficiency advantage over gasoline in an IC engine [3]. Bioethanol can be directly used or it can be blended with gasoline in different ratio from 3 to 25%, depending on countries. The most common that has been widely used is E10 (10% bioethanol blended with 90% gasoline), however in Brazil, bioethanol 100% (E100) or bioethanol 25% (E25) are used [4].
Recently, the production of bioethanol has focused on crops, such as maize, sugarcane and soybean. It includes three major groups: sucrose-containing feedstocks (sugarcane, sugar beet, sweet sorghum and fruits), starchy materials (corn, milo, wheat, rice, potatoes, cassava, sweet potatoes and barley) and lignocellulosic biomass (wood, straw and grasses) [5]. Major countries produced bioethanol include Brazil (sugarcane 100%), the USA (corn 98%, sorghum 2%), China (corn 70%, wheat 30%), EU (wheat 48%, sugar beet 29%) and Canada (corn 70%, wheat 30%) [2]. Although the production of bioethanol can provide environmental and economic benefits, but food security in developing countries has been a key factor to the instability, whereas a large amount of bioethanol is produced from crops in developed countries, that has been the crucial social controversy for bioethanol production.
Sweet sorghum (Sorghum bicolor) is a C 4 crop and is considered as one of the most important food and fodder crops in arid and semiarid regions of the world [6]. This crop occupies approximately 45 million ha in Africa and India, accounts for >80% of the global cultivated area. Similar to grain sorghum, sweet sorghum has rapid growth and wide adaptability [7], good drought resistance [8], salinity tolerance [9] and high yield of biomass [10]. Sweet sorghum has played an important role in promoting the development of agricultural production, livestock husbandry [11], biofuel [12,13], refining sugar and paper making [10]. Carbohydrates in sweet sorghum can be nonstructural as sugars and starch, or structural such as cellulose, hemicellulose and pectic substances [14]. The major sugars in sweet sorghum are the monosaccharides, glucose and fructose; the disaccharides, sucrose and maltose; and the trisaccharide, raffinose. Depending on the kind of sugar in the stalk, there are saccharin and syrup types of sweet sorghum [14]. Sugars in sweet sorghum stalk are mostly sucrose and invert sugars are glucose, fructose, maltose and xylose [15]. It was reported that other sugars such as mannose, galactose and arabinose were not found in juice of the crop; therefore, the production of bioethanol from sweet sorghum is appropriate, as the carbohydrates present are easily converted to bioethanol. The bagasse and grain of the crop are also possible to produce bioethanol, but they are not economically efficacy [16].
Although sweet sorghum has been suggested as a potential crop for bioethanol production in developing countries [6,17,18], but it is still acknowledged as a new crop in many countries. To earn permission for cultivation in a large area for bioethanol production, the cultivation protocol for this crop should be critically examined. In our previous report, we have selected a cultivar 4a from 66 sweet sorghum cultivars through 3 years of cultivation in Vietnam and evaluated the effects of sowing times, densities and soils to biomass and bioethanol production [19]. In this chapter, we describe the application of different fertilizers, including the combinations, doses and time of fertilization and examined their effects on the productivity of biomass, sugar content and ethanol production of sweet sorghum.

Materials and methods
The cultivar 4a (India origin) was selected from a previous experiment as the most potential of high biomass and sugar content [19]. Seeds of this cultivars were tested and their germination rate was >95%.
The experiments were conducted in three different locations in northern Vietnam including Phu Tho, Hoa Binh and Bac Giang provinces, in continuous cropping seasons from 2010 to 2013. In Vietnam, all major ethanol factories are currently using cassava as a material for production, with exception of Phu Tho factory where both cassava and sugarcane can be produced for ethanol production. Therefore, Phu Tho province is the first priority to be chosen as a place of sweet sorghum production. Bac Giang and Hoa Binh are selected because their close distance to the ethanol factory in Phu Tho province (less than 100 km). In addition, the three provinces have a large area of hilly and abandoned areas which can be exploited for cultivation of sweet sorghum.

Applied doses
Different ratios of N, P 2 O 5 and K 2 O from 30, 60, 90 kg/ha with 2 tons/ha of manure were combined. The commercial fertilizers were purchased from Ha Bac nitrogenous fertilizer & chemicals company limited (Bac Giang city, Vietnam). The details of fertilizer ratios are presented in Table 1. Each treatment was conducted in thrice in a plot of 30 m 2 and repeated in three provinces Hoa Binh, Phu Tho and Bac Giang provinces, Vietnam. Effects of different-treated doses on stem yield, sugar content and ethanol production of the cultivar 4a were measured.

Chemical composition of soils in locations selected for the experiment
The criteria for selecting locations in each Hoa Binh, Phu Tho and Bac Giang provinces are where there are abandoned and difficult for cultivating food crops because of their low nutrient soils, to examine the possibility of exploiting the cultivation of sweet sorghum for ethanol production. There were nine villages were selected (total 27 villages) and soils in the selected area in each villages were taken and analyzed for pH, contents of percentages of N, P, K, OC (organic carbon), CEC (cation exchanged capacity) (cmol (+) /kg), Zn (mg/kg), Fe (mg/kg) and Al (mg/kg) ( Table 2). The analysis of these criteria was conducted by conventional methods.

Applied time of fertilization
This experiment was conducted in a plot of 30 m 2 in thrice, in the three locations in Hoa Binh, Phu Tho and Bac Giang provinces in summer-autumn cropping season 2010 and repeated in 2011. The dose of fertilizers per ha included 90 N +60 P 2 O 5 +60 K 2 O + 2 tons manure. Applied times of fertilization are presented in Table 3.

Improvement of biomass and ethanol production by micronutrient fertilizers
This experiment was conducted to examine the role of micronutrient fertilizers by an additional application of (NH4) 2

Determination of biomass, sugar content (Brix %) and ethanol production
Sweet sorghum at flowering stages in different trials was harvested in random. The biomass weight, sugar content (Brix %) and ethanol production were determined by a method described in Ref. [19].

Statistical analysis
All trials of the study were conducted in a completely randomized design with three replications. Means and differences between the treatments were determined by using a two-way analysis of variance (ANOVA) with values of least significant difference (LSD) at P ≤ 5% level from IRRISTAT program version 5.0.

Effects of different applied doses of fertilizers on stem yield, sugar content and ethanol production of sweet sorghum
There were 10 different applied doses were used in the designated locations in Hoa Binh, Phu Tho and Bac Giang provinces. The effects of these different fertilization doses on stem yield, sugar content and ethanol production of the cultivar 4a are presented in Table 4.
It is observed that the stem yield of sweet sorghum varied between 44.7 and 77.3 tons/ ha, depending on applied doses of fertilizers and locations (

Effects of applied times of fertilizers on stem yield, sugar content and ethanol production of sweet sorghum
There were three trials of fertilizer application in different times and fertilizer doses ( Table 4), applied in the three locations in Hoa Binh, Phu Tho and Bac Giang provinces ( Table 5). It indicated that times of fertilizer application at 3-4 to 7-8 leaf stages provided the highest stem yield and ethanol yield in every location and their values were markedly different as compared among the 1st, 2nd and 3rd applied times ( Table 5). However, when the amount of fertilizers were divided into three times of application, of them, the third fertilization was either after 20 d of the second application (trial 2) and before flowering, the yields of stems and ethanol production were significantly reduced, as compared with other treatments ( Table 5). The stem yield was markedly decreased >20 tons/ha, whereas the amount of ethanol production was significantly reduced > tons/ha. For location, Phu Tho province obtained higher yields of stems and ethanol production than Hoa Binh and Bac Giang provinces. For the sugar content (Brix %), different times of fertilizer application did not influence on the sugar content of sweet sorghum, as no significant difference among treatments were observed ( Table 5).

Effects of micronutrient fertilizers on stem yield, sugar content and ethanol production of sweet sorghum
In this experiment, the control was the based fertilization (BF) that included 90 N + 60 P Note: Trials 1-3 are different applied times as indicated in Table 3. It was found that by the application of based fertilization, the stem yield of sweet sorghum was 62.4-66.0 tons/ha. However, the supplemented application of Mo significantly increased the stem yield 10-12 tons/ha and it was higher than either the supplementation of P 2 O 5 or K 2 O ( Table 6). For the sugar content, there was no remarkable difference among treatments (14.8-15.5 Brix %). Following the promoted amount of stem yield, the ethanol productivity was the maximum with Mo application, followed by P 2 O 5 and K 2 O supplementation. They were all significantly higher than the application of unique-based fertilizers ( Table 6).

Discussion
Although sweet sorghum is well adaptable to many soils and climates and has a strong resistance to pests and diseases, an effective fertilization protocol is crucial to farmers and agricultural extensionists to promote the biomass and ethanol productivity of this crop. Trials of this study were replicated in the three locations in three different provinces, where are the most suitable for ethanol production in the North of Vietnam [19]. Among different doses of fertilizers, this study found that the application of 90 N + 90 P 2 O 5 + 60 K 2 O was the best for maximum biomass and ethanol productivity of the crop. Their values were significantly higher than those of other treatments ( Table 4). Nontreatment of N, P 2 O 5 and K 2 O was examined and the lack of fertilization of any among these nutrients caused severe markedly reduction of stem yield, sugar content and ethanol production; however, N was the most important for biomass and ethanol productivity ( Table 4).
It has been described that sweet sorghum can become a profitable crop for farmers in developing countries, however its agronomic practices should be optimized to improve the productivity. Among them, nitrogen is the most important nutrient element as this crop responds well to N fertilization [20]. Turgut et al. [21] investigated the optimum N input for maximizing sweet sorghum yield in a clay loam soil. Reddy et al. [22] also studied the effects of N input on sweet sorghum growth. Uchino et al. [20] reported that the amount of N from 90 to 120 kg/ha was the optimum for ethanol productivity, but the  Table 6. Effects of micronutrient fertilizers on stem yield, sugar content and ethanol production of sweet sorghum. amount of N >150 kg/ha may cause lodging. In this study, the application of N >90 kg/ha was not studied as it may cause economic troublesome for farmers in developing countries such as Vietnam.
This research also verified the application of K 2 O and P 2 O 5 in combination with nitrogen and manure ( Table 4) to reduce the dependency on nitrogen application. The use of manure, K 2 O and P 2 O 5 with reduced quantity of nitrogen fertilizers are efficacy for soil health and maintain the biomass and ethanol productivity of sweet sorghum. The time of application was also examined to optimize the time of application and doses of fertilizers. The best time for fertilization was found to be on 3-4 to 7-8 leaf stage ( Table 5). Findings of this study indicate that fertilization should not be conducted after 7-8 leaf stage, as the application of fertilizers at 20 d after the second application (7-8 leaf stage) to flowering stage caused significant reduction of stem yield and ethanol production, although the sugar content (Brix %) was not influenced ( Table 5). This also helps to save the labor costs as the too much fertilization may attribute to heavy fieldwork and economic deficit in sweet sorghum cultivation.
In this study, for micronutrient fertilization, 5 kg/ha of (NH4) 2 MO 7 O 2 .4H 2 O (Mo) was used as supplement to the base fertilizer application of 90 N + 60 P 2 O 5 + 60 K 2 O + 2 tons/ha manure. The supplements of 60 kg/ha of P 2 O 5 and K 2 O were also conducted to compare with that of the Mo supplement. Interestingly, the use of Mo was the most effective in enhancing the biomass and ethanol production and it was higher than that of either P 2 O 5 or K 2 O supplements ( Table 6). The use of Mo was found to be more effective than the adding of P 2 O 5 and K 2 O. In addition, the application of Mo at 5 kg/ha provided less heavy fieldwork than the supplementation of 60 kg of either P 2 O 5 or K 2 O. The application of micronutrients such as ZnSO 4 and sodium borate was effective on the stalk yield of the sweet sorghum ICSV 93046 variety, which is widely cultivated in India and Philippines [23]. The supplements of micronutrients of Zn and Fe significantly promote the stalk yield of sweet sorghum [24]. It is therefore proposed that the application of micronutrient fertilizers is effective to improve the productivity and profitability of sweet sorghum than the supplement of unique either P 2 O 5 or K 2 O.
The influences of fertilization including times of application and applied doses on energy crops have been extensively studied. It has been reported that the application of N fertilizer significantly improves the production of bioenergy crops, such as switchgrass (Panicum virgatum L.) and corn (Zea mays L.) [25]. Individual applications of N, P, or K fertilizers markedly improved cassava (Manihot esculenta Crantz) yield. The best yield performance in this research to increase the yield of cassava (62.8 t/ha) was the use of 400, 200 and 400 kg/ ha of N, P, K fertilizers, respectively, although the use of P fertilizer incorporation with high levels of N and K fertilizers was not effective to improve cassava yields [26]. In addition, Olugbemi and Ababyomi [27] examined different treatment of nitrogen fertilizer on the growth and ethanol yield of four sweet sorghum varieties and found that the application of 120 kg N/ha was the best for ethanol production. There are also several research studies on optimizing the nitrogen requirements of sweet sorghum for various environments and soil types [20][21]. The applied doses on sugarcane were important on the yield and ethanol production in Brazil [28]. Appropriate doses of fertilizers are useful to increase the yield and ethanol production of biofuel crops, but the overuse of fertilizers may cause critical environmental pollution [29]; therefore, the examination of applied doses and times of application as conducted in this research is indispensable for the cultivation of biofuel crops.

Conclusions
Sweet sorghum is a promising alternative crop for bioethanol production in developing countries to reduce the social criticisms on the use of food crops for biofuel production. Findings of this study, together with a previous work, examined the sowing times, densities and soil chemicals [19] to determine the cultivar 4a which was continuously examined in this research for different doses of fertilizers and times of application. Achievements of the two research studies are useful to establish a protocol of sweet sorghum cultivation for farmers and agricultural extensionists in developing countries, as the fertilization of different doses and times was examined in different locations with low nutrient contents that were difficult to cultivate other crops. This study also highlighted the higher possibility of using sweet sorghum in infertile soil in hilly areas than other bioenergy crops. The combination of manure and micronutrients with a reduced amount of inorganic fertilizers is crucial to establish a sustainable agriculture in sweet sorghum production.