Ultrasound assisted extraction is very efficient extraction procedure. Sonication induces cavitation, the process in which bubbles with a negative pressure are formed, grown, oscillated, and may split and implode. By this process different chemical compounds and particles can be removed from the matrix surface by the shock waves generated when the cavitation bubbles collapse. The implosion of the cavities creates microenvironments with high temperatures and pressures. Schock waves and powerful liquid micro jets generated by collapsing cavitation bubbles near or at the surface of the sample accelerate the extraction (R. Kellner et al., 2004). Ultrasonic assisted extraction has many advantages since it can be used for both liquid and solid samples, and for the extraction of either inorganic or organic compounds (S. L. Harper et al., 1983). If extracted from solid samples, different problems can occur: there is a possibility of the decomposition of the analyte which could be trapped inside of the collapsing cavitational bubbles. The ultrasound extraction system can be also applied as a dynamic system in which the analytes are removed as soon as they are transferred from the solid matrix to the solvent. In this process, furthermore, the sample is continuously exposed to the solvent (I. Rezic’ et al., 2008).

This is a modified maceration method where the extraction is facilitated by the use of ultrasound (high-frequency pulses, 20 kHz). Ultrasound is used to induce a mechanical stress on the cells of biomass solid samples through the production of cavitations in the sample. The cellular breakdown increases the solubilization of metabolites in the solvent and improves extraction yields. The efficiency of the extraction depends on the instrument frequency, and length and temperature of sonication. Ultrasonification is rarely applied to large-scale extraction; it is mostly used for the initial extraction of a small amount of material. It is commonly applied to facilitate the extraction of intracellular metabolites from plant cell cultures (Kaufmann, 2002; Sarker, 2006).

Anaerobic treatment processes require the presence of a diverse closely dependent group of bacteria to bring about the complete conversion of complex mixtures of substrates to methane gas. It is puzzling that single species of bacteria have not evolved to convert at least simple substrates such as carbohydrates, amino acids, or fatty acids all the way to methane [26].

Conventional phase and high-rate two-phase anaerobic digestion processes have frequently been employed in order to treat both soluble and solid types of domestic and industrial wastes. The most significant outcome of anaerobic digestion processes is that they generate energy in the form of biogas namely, methane and hydrogen. Therefore, due to current imperative environmental issues such as global warming, ozone depletion, and formation of acid rain, substitution of renewable energy sources produced from biomass, such as methane and hydrogen, produced through anaerobic digestion processes will definitely affect the demand and consumption of fossil-fuel derived energy [27].

The anaerobic treatment is a biological process widely used in treating wastewater. When these have a high organic load, presents itself as the only alternative would be an expensive aerobic treatment, due to the oxygen supply. The anaerobic treatment is characterized by the production of "biogas" consisting mainly of methane (60-80%) and carbon dioxide (40-20%) and capable of being used as fuel for generating thermal energy and / or electric. Furthermore, only a small fraction of COD treated (5-10%) is used to form new bacteria, compared to 50-70% of an aerobic process. However, the slow anaerobic process requires working with high residence times, so it is necessary to design reactors or digesters with a high concentration of microorganisms [28, 29]. Actually is a complex process involving several groups of bacteria, both strictly anaerobic and facultative, which, through a series of stages and in the absence of oxygen, flows mainly in the formation of methane and carbon dioxide. Each stage of the process, described below, is carried out by different groups of bacteria, which must be in perfect balanced. Figure 1 shows a schematic representation of the main conversion processes in anaerobic digestion, suggested by Gujer [30].

(1) Hydrolysis. In this process complex particulate matter is converted into dissolved compounds with a lower molecular weight. The process requires the mediation of exo­enzymes that are excreted by fermentative bacteria. Proteins are degraded via (poly) peptides to amino acids, carbohydrates are transformed into soluble sugars (mono and disaccharides) and lipids are converted to long chain fatty acids and glycerine. In practice, the hydrolysis rate can be limiting for the overall rate of anaerobic digestion. In particular the conversion rate of lipids becomes very low below 18°C.

(2) Acidogenesis. Dissolved compounds, generated in the hydrolysing step, are taken up in the cells of fermentative bacteria and after acidogenesis excreted as simple organic compounds like volatile fatty acids (VFA), alcohols and mineral compounds like CO2, H2, NH3, H2S, etc. Acidogenic fermentation is carried out by a diverse group of bacteria, most of which are obligate anaerobe. However, some are facultative and can also metabolize organic matter via the oxidative pathway. This is important in anaerobic wastewater treatment, since dissolved oxygen (DO) otherwise might become toxic for obligate anaerobic organisms, such as methanogens [31].

(3) Acetogenesis. The products of acidogenesis are converted into the final precursors for methane generation: acetate, hydrogen and carbon dioxide. As indicated in Figure 1, a fraction of approximately 70% of the COD originally present in the influent is converted into acetic acid and the remainder of the electron donor capacity is concentrated in the formed hydrogen. Naturally the generation of highly reduced material like hydrogen must be accompanied by production of oxidized material like CO2.

(4) Methanogenesis. Methanogenesis may be the rate liminting step in the overall digestion process, especially at high temperatures (> 18°C) and when the organic material in the influent is mainly soluble and little hydrolysis is required. Methane is produced from acetate or from the reduction of carbon dioxide by hydrogen using acetotrophic and hydrogenotrophic bacteria, respectively:

Acetotrophic methanogenesis: CH3COOH ^ CH4 + CO2 (3)

Hydrogenotrophic methanogenesis: 4H2 + CO2 ^ CH4 + 2H2O (4)

Different from aerobic treatment where the bacterial mass was modeled as a single bacterial suspension, anaerobic treatment of complex wastewaters, with particulate matter in the influent, is only feasible by the action of a consortium of the four mentioned groups of bacteria that each have their own kinetics and yield coefficients. The bacteria that produce methane from hydrogen and carbon dioxide grow faster than those utilizing acetate that the acetotrophic methanogens usually are rate limiting for the transformation of acidified wastewaters to biogas [32].

The different groups of bacteria involved in the conversion of influent organic matter all exert anabolic and catabolic activity. Hence, parallel to the release of the different fermentation products, new biomass is formed associated with the four conversion processes described above. For convenience, the first three processes often are lumped together and denominated acid fermentation, while the fourth step is referred to as methanogenic fermentation.

The removal of organic matter-COD during the acid fermentation is limited to the release of hydrogen only 30% of the organic matter is converted into methane via the hydrogenotrophic pathway. Hence, a necessary condition for efficient organic matter removal in an anaerobic treatment system is that a sufficient mass of acetotrophic methanogens develops.

Acid fermentation tends to cause a decrease in the pH because of the production of VFA and other intermediates that dissociate and produce protons. As methanogenesis will only develop well at a neutral pH values, instability may arise, if for some reason the rate of acid removal by methane production falls behind the acid production rate: the net production of acid will tend to cause a decrease in pH, and thus may reduce the methanogenic activity further. In practice, this so called "souring" of the anaerobic reactor contents is the most common cause for operational failure of anaerobic treatment systems. The danger of souring can be avoided, by maintaining the proper balance between acid and methanogenic fermentation which in fact means that both the methanogenic digestion capacity and buffer capacity of the system should be sufficiently high [29, 33].

Weed control is necessary when establishing willows from cuttings, because its takes a relatively long time for willow cuttings to develop attributes which make them competitive against weeds. Competition is an interaction between plants which require the same limited resources like nutrients, water and light. Harper [56] defines competition as ‘An interaction between individuals brought about by a shared requirement for a resource in limited supply and leading to a reduction in the survivorship, growth and/or reproduction of the individuals concerned’, and thereby points to the effects of competition. The aim of weed control is to ensure that as much resources as possible are accessible for the crop and not for the weeds, and to reduce or delay growth and development of the weed flora [57].

Willows under establishment from cuttings have a relatively low competitive power against weeds because it takes a while for willows to develop roots needed for the uptake of nutrients and water. Consequently, perennial weeds, which have a developed root system prior to the onset of leaves, have to be removed completely before planting willows. This commonly is done by means of one or two applications of a glyphosate-based herbicide, applied at the appropriate rate, during the summer/autumn prior to spring planting. If the area has not been used for agricultural purposes for a number of years before planting, it is recommended to grow cereals there for at least one season to ensure an adequate weed control [42]. The relative competitive ability is also affected by seed rate (plant density), which is low for willow (between 1 and 2 cuttings m-2), in comparison to the amount of germinating annual weeds triggered by seed bed preparation. Such weeds may germinate only a few days after seed bed preparation, while it may take a week or more for willow cuttings to exhibit a first bud burst after planting. This implies that the time between the last seed bed preparation and willow planting should be minimized. To counteract the effects of the inherent differences in relative emergence time between willow and weeds, soil cultivation by different types of cultivators, rototillers or harrows are recommended as a weed control measure during willow establishment [46, 58]. There are also different soil — applied herbicides that are permitted to be used at planting or shortly thereafter. Given the low planting density of willow cuttings, a full canopy closure, which for willow implies a leaf area index > 6 m2*m-2 [4, 38] is hardly ever reached during the establishment year, which means that if weeds are not kept back during the establishment year, they may establish and compete with willows for light. The use of mechanical weeding may therefore proceed even after bud burst and early shoot formation in willows. As the cuttings are well fixed in the soil and young willow shoots are flexible, they will not become damaged by this treatment. The current recommendation is to perform these control measures at least three times during the first year [46].

Weed control might also be necessary to perform the year after planting depending on weed management success the first year, clones and site conditions. As the willow plants will be better established by then, it is usually enough to perform mechanical weeding two times early in the season [46]. Another possibility is to spray a soil-applied herbicide well before bud burst [42] or to use a selective herbicide during spring or early summer [46]. Weed control the second year usually requires that the first year shoots are cut back. This practice has been questioned [59] and is no longer recommended in Sweden [42]. If weed control has been efficient during the establishment phase, no additional measures are required to control the weeds the following years. If early plant mortality has led to gaps in the stand, weeds may establish and maintain themselves below canopy gaps (Figure 7). In case weeds survived below such gaps, weeds may be controlled directly after each harvest.

If the weeds are not controlled during the establishment phase, willow growth might be dramatically reduced. Field experiments conducted in Southern Sweden by Albertsson in 2010-2012, with 10 modern willow varieties, grown both with — and without weeds, have shown that weeds can increase plant mortality, and reduce growth the first year by more than 95% [42], see Figure 8. Several other studies have also shown that willow, in the establishment phase, is very sensitive to competition from other plants [60, 61, 62]. Preliminary data from the Swedish study also suggest that there is an interaction between voles and weediness, since plots with weeds show more damage by voles than plots without weeds, thereby making weed control even more important.

Weeds in willow short rotation coppice might, in the future, be controlled with other measures than the above mentioned. Studies are ongoing to investigate if willow clones differ in their ability to compete with weeds. Fast initial growth, early bud burst, fast canopy closure and the ability to tolerate or release allelophatic substances might be favorable weed competing traits. If differences exist, it might be possible to breed for these traits or to use competitive willow varieties that combine well with a specific weed control measure.

Different cover crops such as rye (Secale cereale L.), dutch white clover (Trifolium repens L.), buckwheat (Fagopyrum esculentum Moench) and caragana (Caragana arborescens Lam.) have been studied as a way of controlling weeds and improve nutritional management in willow [63, 64, 65]. However, there is still more research to be conducted in this area before a suitable willow cover crop system is ready for commercial use. Mechanical weeding techniques are under constant development and recent results indicate that automatic intra­row weeding is possible [66]. Hence, these techniques may be further developed to be used in willow since weeds within the rows are hard to control mechanically with conventional equipment.

The textile industry consumes the largest portion of the colorant available in the world market. Due to the high customer demand, more than 100,000 commercial dyes exist in the market causing more than 700,000 tonnes of dyes to be produced annually (McMullan et al., 2001; Pearce et al., 2003). The result is a very high production of colored wastewater. The characteristics of textile wastewater (either quantitatively or qualitatively) vary greatly depending on the type of raw materials, chemicals, techniques or specific process operations at the mill, the equipment used and the production design of the textile processes (Bisschops and Spanjers, 2003; Dos Santos et al., 2006).

The textile industry consumes huge amounts of water in its wet processes. The average wastewater generation from a dyeing facility is estimated between 3800 and 7600 million m3 per day. Desizing, scouring, bleaching, mercerizing and dyeing are the common wet textile process operations. Among these, the mercerizing and dyeing processes consume the biggest specific volumes of water with a water usage of 230-310 L/kg and 8-300 L/kg of textile processed, respectively (Dos Santos et al. 2007).

Due to inefficiency of the textile processing activities, only 10% of the chemicals in the pre­treatment and dyes in dyeing operations remain on the fabric. In other words, about 90% of chemical substances will be discharged as textile effluent (IPPC, 2003). Others have reported that between 50 and 95% of the dyes are fixed on the fiber while the remainder is discarded in the subsequent textile-washing operations (EPA, 1997; Trovaslet et al., 2007). The amount of dye lost into the wastewater depends upon the type of dyestuff used, as well as the methods and application routes of the textile processing operation. Additionally, it depends on the intended color intensity that is required for each particular design (Willmott et al., 1998).

Textile wastewater is characterized with high chemical and biochemical oxygen demand, suspended solids, high values of conductivity and turbidity and intense color. This is caused by the presence of dye residues or intermediates and auxiliary chemicals added in the many stages in textile processing (Mohan et al., 2007a; Miranda et al., 2009). Textile processes with natural fibers generate higher pollution load as compared to synthetic fibers mainly due to the use of pesticides for preservation of the natural fibers (Correia et al., 1994).

Textile dyeing wastewater is also characterized by high salt content, which also imposes potential environmental problems. Typical cotton batch dyeing operations use quantities of salt that range from 20 to 80% of the weight of goods dyed, with common concentrations between 2,000 mg/L and 3,000 mg/L. Sodium chloride and sodium sulfate constitute the majority of the total salts used. Magnesium chloride and potassium chloride are used as raw materials in lower concentrations (EPA, 1997).

Common characteristics of textile wastewater from cotton textile wet processing for different processing categories are shown in Table 1. The highest concentration of organic pollutants (in terms of COD) is generated from bleaching while the highest concentration of total solids comes from the desizing process. The highest concentration of color, ranging from 1450-4750 ADMI, is generated from the dyeing process (Bisschops and Spanjers, 2003;

Dos Santos et al., 2007). Metals such as copper, cadmium, chromium, nickel and zinc are also found in textile effluents, as they are the functional groups that form the integral part of the dye molecule (IPPC, 2003).

Pyrolysis is an important process in energy recovery from biomass and also as a previous stage to other processes such as gasification. Valuable gases, such as H2 and CO, can also be generated by pyrolysis. These gases can be useful, among other applications, in chemical synthesis and high efficiency combustion systems such as fuel cells.

The hydrogen-rich product gas from biomass pyrolysis is believed to become a valuable energy source with natural carbon dioxide. However, biomass has low energy density, so the enhancement of the product gas quality from biomass gasification is necessary. Beside a particular problem which has not been completely solved so far is tar formation. Catalytic processes are considered as the most promising method with the highest potential to contribute a solution to this problem. Temperature is an important variable in thermal decomposition processes of biomass, significantly influencing the product distribution. Pyrolysis is an endothermic process, and the use of low temperatures in this process decreases the input energy for a system that is very positive from an energetic point of view and the system operation is easier than high temperature systems.

For both catalytic activity and economic reason, a nickel catalyst is the most suitable choice and the most widely used in the industry among metals like Co, Pt, Ru and Rh, which were investigated by many authors [10,17-23]. Moreover, nickel based catalysts are reported to be quite effective not only for tar reduction, but also for decreasing the amount of nitrogenous compounds such as ammonia [24]. In most reports, conventional nickel catalysts, which had been developed for steam tar reforming, were tested [6,9,13,15,25,26]. However, the investigations are still limited because of coking [19,22] or the use of expensive materials as catalyst supports (CeO2, AhO3, Al, SiO2, TiO2, ZrO2, MgO or WO3). Among the above investigations, the most interesting one concerning the current study is brown coal gasification by the addition of a nickel catalyst in fluidized bed gasification at low temperature [21-23]. However, nickel catalyst has only been used for coal gasification itself.

To satisfy both high quality of product gas and use of a cheaper catalyst support (brown coal char), in this study, nickel loaded brown coal char catalyst has been developed for the new purpose of decomposing tarry material from woody biomass pyrolysis. A suitable temperature for catalytic tar decomposition was investigated in a two-stage fixed bed reactor. The effect of temperature on gas yield and carbon conversion have been discussed in detail and compared in the case of non catalyst and Ni/BCC catalyst. The better temperature is a reference result, being used in fluidized bed gasification which is available for continuous tests to assess durability. Catalytic activity was tested, evaluated by woody biomass pyrolysis in a fluidized bed gasifier for both of Ni/BCC and reference catalyst

Ni/АЬОз. In a lab scale fluidized bed gasifier (FBG) Experiments, inside of FBG reactor is constructed by two beds, the primary one is a fluidized bed with sand where biomass was fed to produce tar, and the other is a catalyst bed that is used to evaluate and to compare catalytic activity between the new catalyst and a conventional Ni/AhO3 catalyst. The Ni/BCC catalyst is prepared by ion exchange method, dried at 380 K in nitrogen for 24 h, and is then calcined at 923 K in nitrogen for 90 min. Sample for characterization of catalyst was prepared on the fixed-bed reactor under various conditions such as nickel loaded brown coal particle size range of 0.5 to 2 mm, pyrolysis temperature range of 823 to 1023 K that are needed to investigate the effect both of catalyst particle size and pyrolysis temperature on crystallite size of Ni/BCC. The temperature as a function of gas yield and stable activity of catalyst absence of steam are investigated in this chapter.

The state of art scenario presented here makes use of the conversion rates from the experiment data (Table 2). Approximately, 200 L of bioethanol yields per dry tones of lignocellulosic biomass and anticipated prices of RM 250 per dry tones of lignocellulosic biomass and RM 264 per tones of 60% concentrated sulfuric acid. The feedstock cost for one litre of bioethanol produced using either from oil palm trunk or wood wastes is estimated at about RM 1.25/litre. The production cost for one litre bioethanol from lignocellulosic biomass is estimated at RM 0.26 with the hydrolysis cost contributed RM 0.20 based on the sulfuric acid is added at a ratio of 5:1 (acid: dry weight of biomass) with acid lost in the sugar stream is not more than 3% during recovery (97% recoverable). Fermentation cost contributed RM 0.06 with the yeast would be grown at the site without cultivation process [38]. Therefore, the total cost per litre of bioethanol produced is RM 1.51 excluding capital and fixed variable costs. However, without the recovery of sulfuric acid during hydrolysis, the cost of bioethanol from lignocellulosic biomass would be rose up to RM7.85, excluding capital and fixed variable costs. With ethanol prices now at RM 2.10 per litre, it is possible for the Malaysia to produce the bioethanol from oil palm trunk and wood wastes, yet it would be not profitable to produce ethanol from lignocellulosic biomass without using the recovery system for sulfuric acid during hydrolysis.

The table below shows different scenario on the biomass feedstock and bioethanol yield that might affect the cost of bioethanol in Malaysia. The scenarios were based on 97% sulfuric acid recovered during hydrolysis and no change on the cost of fermentation production.

Scenario Analysis :

The economic feasibility of bioethanol production in Malaysia from lignocellulosic biomass is highly dependent on the feedstock cost and recovery yield. The cost of feedstock contributed approximately 80% (excluding capital and fixed variable costs) to the total bioethanol cost when the feedstock price estimated at RM 250 per dry weight ton. As the feedstock price increase 5% to 15% per dry ton, the cost of bioethanol increased from as low as 4% up to almost 13%. Higher recovery yield from the bioethanol process will surely reduce the cost of bioethanol produced per litre when the cost of feedstock remains the same. However, as the conversion yield of bioethanol decrease from 200 L per dry weight ton of biomass, the cost of biothenol per litre increase from 5% up to 17%.

Like corn in the United States and sugarcane in Brazil, the relatively low feedstock cost will only makes this process economically competitive. The cost of producing ethanol from sugarcane in Brazil is estimated at about RM 0.60 per litre, excluding capital costs. U. S. ethanol conversion rates utilizing corn as the feedstock are estimated at approximately 2.65

gallons of ethanol per bushel for a wet mill process and 2.75 gallons per bushel for a dry mill process. Net feedstock costs for a wet mill plant are estimated at about RM 0.30 per litre with total ethanol production costs estimated at RM 0.76 per litre. Net feedstock costs for a dry mill plant are estimated at RM 0.38 per litre with total ethanol production costs at RM 0.76 per litre. Molasses, from either sugarcane or sugar beets, was found to be the most cost competitive feedstock beside the lignocellulosic biomass. Estimated ethanol production costs using molasses were approximately RM 0.92 per litre with a RM 0.66 per litre feedstock cost [39].

Table 3. Cost of bioethanol per litre with different scenario on cost of raw materials and conversion yield

8. Conclusion

The studied lignocellulosic biomass has a higher bioethanol yield per tonne feedstock (L/t) than most of the commercialized bioethanol feedstock. However, improvement had to be made on the conversion efficiency to obtained higher ethanol yield to make it more comparable with the sugar containing and starchy material. The composition of substance that can be converted to glucose played a big influence on the ethanol yield per tonne feedstock. With the large amount of glucose convertible material and abundant availability, these lignocellulosic biomasses are potential feedstock for bioethanol production.

Author details

K. L. Chin1 and P. S. H’ng

Faculty of Forestry, Universiti Putra Malaysia, UPM Serdang, Selangor, Malaysia

Today, only a small numbers of chemicals are produced from lignocellulosic feedstocks via fermentation. Much less attention has been given to biomass as a feedstock for organic chemicals, while there has been a strong political and technical focus on using biomass to produce transportation fuels. However, replacement of petroleum-derived chemicals with those from biomass will play a key role in sustaining the growth of the chemical industry [88]. One way to replace petroleum is through biological conversion of lignocellulosic resources into products now derived from petroleum. The current developments especially in fermentation technologies, membrane technologies and genetic manipulation open new possibilities for the biotechnological production of market relevant chemicals from renewable resources [5].

In lignocellulosic feedstocks biorefinery processes, the sugars or some of the fermentation products can be chemically converted into a variety of chemicals, which could be used to form biological materials, such as protein polymers, xantham gum, and polyhydroxybutyrate. The lignin as remaining fraction from lignocellulosic feedstocks, could be converted through hydrogenation processes into materials, such as phenols, aromatics, and olefins, or simply burned as a boiler fuel for cost efficiency of the overall process. Currently, conventional chemicals include acetaldehyde, acetic acid, acetone, n — butanol, ethylene, and isopropanol can simply be derived from LCF. Appropriate organisms could then convert the sugars into the desirable products and co-products for this process. The advantage to such products is that the market is already established, and minimal effort is required to integrate these products into existing markets. However, co-product markets might be limited, and caution must be taken in considering their impact on overall economics, especially for large-scale implementation. A sequence of processes comparable to those employed for cellulosic ethanol production would be used to pre-treat the lignocellulosic biomass to open its structure for the weight of the feedstock. Therefore, lignocellulosic biomass might be expected as the low cost of raw materials could be converted to a variety of commodity chemicals and materials [20].

3.1.1. Physical characteristics of samples

Bio-oil used in the single separation process was produced by the pyrolysis of Mongolian pine sawdust (Wang et al., 2008). Wang et al. (Guo et al., 2009b; Wang et al., 2009) carried out experimental research on molecular separation of the bio-oil, which was pre-treated by centrifugation and filtration to remove solid particles. Molecular distillation of the bio-oil at 50, 70, 100, and 130 °C, respectively, was investigated under a fixed pressure of 60 Pa. Under all of the tested conditions, the light fraction collected by the second condenser placed before vacuum pump was designated as LF, the middle fraction condensed by the internal condenser as MF, and the heavy fraction as HF.

The color of the distilled fractions becomes lighter while the residual fractions become darker. Under the four conditions, water was concentrated in the LFs, which had water contents of about 70 wt%. The LFs could not be burned because of their high water contents. The pH values of the LFs were in the range 2.13-2.17 as a result of their carboxylic acid contents. On the other hand, the HFs had the highest heating values and the lowest water contents, resulting in good ignitability but inferior fluidity. At a distillation temperature of 70 °C, the water content of the MF was as low as 2 wt%. The total mass of the bio-oil distillation fractions amounted to more than 97% of the bio-oil feed. With increasing temperature, the yield of the LF increased without any coking or polymerization problem. Water and volatile carboxylic acids were evaporated from the feedstock in the temperature range 50-130 °C under low pressure, and more carboxylic acids escaped from the liquid at higher temperature. However, on further increasing the temperature, this phenomenon was not so pronounced, due to more and more molecules of higher boiling point also being distilled. The yield of the distilled fraction increased with increasing distillation temperature. However, too high temperature may lead to decomposition of some chemical compounds in the crude bio-oil. Hence, there must be an optimum temperature to realize reasonable separation.

The relatively large surface area of microorganisms exposed to the environment where they live enables them to take up and assimilate nutrients readily and a to multiply at impressive rates. An important parameter is the specific growth rate, wich is often expressed as the mean doubling time, defined as the time required by the microbial population to douyble its cellular protein content. The doubling time varies widely depending on the microbial species, the nature of the substrate and the degree of adaption to the substrate.

The growth of a bacterial culture can be determined by measuring the increase of turbidity in the medium as optical density or OD. The most common way to do this is to compare the absorbance of the culture to inoculated medium by shining light with a wavelength of 600 nm through the culture. The more growth occurs, the more turbid the culture will become and more light will be absorbed. Using optical density gives indirect measurement of bacterial growth. It doesn’t tell anything about how many living cells are in the culture. This becomes especially important in stationary phase. Dead cells still absorb light. To determine the actual number of live bacteria the broth is diluted and plated on appropriate media and incubated. In theory, colonies arising on a plate originate from one single bacterium and give therefore an accurate number of the live cells in the culture at that time point.

Procedure:

The kinetics of selected bacterial growth, it is used 10 mL of 24 h culture of the strains and inoculated into 60 mL of nutrient broth, the following conditions: 35 °C and 100 rpm agitation. Samples were read every 30 minutes in a Spectronic 20D+ visible spectrophotometer at 600 nm, substituting readings transmittance (% T) in the following equation:

A = 2 — logM(%T)

Where A, is the absorbance.

The different phases of growth kinetics can be observed by plotting the log (% T) versus time [1].

An experiment was designed to determine different classes of metabolizable energy (AME, AMEn, TME, TMEn) in artemia meals [13] .For determination of metabolizable energy of artemia meal and comparison with fish meal, samples gathered from 3 regions include : Urmia Lake Artemia Meal(ULAM), Earth Ponds Artemia Meal (beside urmia lake)(EPAM) and Ghom Salt Lake Artemia Meal(GSLAM). Then samples dried, milled and used in a biological experiment with fish meal. 20 Rhode Island Red cockerels with approximately same live weight used in Sibbald assay with completely randomized design with 5 treatments and 4 repetitions for determination of AME, AMEn, TME and TMEn.

Results showed there were significant differences between treatments from standpoints of metabolizable energy (P<0.05). ULAM and FM had highest ME and EPAM and GLAM had lowest ME. The highest TME belong to FM and the lowest TME pertained to EPAM. Except to EPAM that had the lowest TMEn, other treatments didn’t have any differences between them.