Reactive Extraction of Microalgae for Biodiesel Production ...aelsindia.com/rjcesaugust2014/2.pdfUnder these conditions, the best biodiesel yields was 98.2%, based on the oil content

RJCES Vol 2 [4] August 2014 10 | P a g e © 2014 AELS, INDIA

ORIGINAL ARTICLE

Reactive Extraction of Microalgae for Biodiesel Production; an

Optimization Study

El Shimi, H1*; Attia, N2; El Sheltawy, S1; El Diwani, G2 1Chemical Engineering Department, Faculty of Engineering, Cairo University, Egypt

2Chemical Engineering and Pilot Plant Department, National Research Center, Egypt *Corrosponding author: [email protected]

ABSTRACT

Concerns about energy security and declining of fossil fuels have led to growing worldwide interests in renewable energy sources such as biofuels. Algal biodiesel is a technically alternative and renewable diesel fuel without excessive engine modifications. This paper provides the biodiesel production from Spirulina-platensis microalgae via reactive extraction methodology "in-situ transesterification" using alcohol and acid catalyst, since the engineering factors that strongly affect the percentage yield of algal biodiesel like oil-to-alcohol molar ratio, acid-catalyst concentration, time and temperature of reaction were studied. The lipids content of Egyptian Spirulina-platensis microalgae was estimated to be 11% wt. The weight of the co-product glycerol obtained was employed to estimate the percentage yield of algal biodiesel. Experiments to look for algal biodiesel was successful; since 84.7% as a yield was achieved at 1:3714 molar ratio of oil-to-alcohol, 100% (wt./wt. oil) catalyst concentration, 8h reaction time and 65oC reaction temperature with continuous agitation at 650 rpm. Various properties of biodiesel were investigated according to EN 14214 standards. Spirulina-platensis methyl esters were closest to a practical biodiesel as seen from its properties. Keywords: Renewable energy; Biofuels; Algal biodiesel; Spirulina-platensis; Reactive extraction; In-situ transesterification; Yield; Lipids; Standards. Received 12.05.2014 Accepted 20.06.2014 © 2014 AELS, INDIA INTRODUCTION The need for a reliable renewable fuel source grows; due to the ever-increasing demand for energy and depletion of fossil fuels. There are many options in this area, but unlike solar and nuclear, biofuels such as biodiesel has the capability of providing a fuel source ideally suited to existing infrastructure within the transportation industry [1-4]. Biodiesel is a green attractive fuel that can be produced from plants oil, animal fats, used cooking oil as well as microalgae oils [1-15]. It is a renewable, non toxic and clean energy alternative to petrol diesel [5-11]. Biodiesel, when blended with petroleum diesel, can be used in unmodified diesel engines. It has the added benefit of higher lubricity and biodegradability, so it helps provide for greater longevity within diesel engines. To use pure biodiesel (B100), an engine usually needs slightly inexpensive modifications; to prevent fouling [4-21]. Microalgae are promising candidate biomass resources as potential feedstock for renewable energy production [4, 7, 22-26]. Microalgae have high oil productivity (1,000–6,500 gallons of oil per acre per year), high lipid content (usually, 30-65% by wt), low cultivation cost, and no competition with agricultural land [5-9]. Biodiesel from microalgae are renewable and carbon neutral (about 2 kg of CO2) are fixed by 1 kg of algal biomass), and can use non-arable land [25-29]. The biodiesel production from microalgae is needed to the well-known transesterification reaction that illustrated in Fig 1. This process has previously been demonstrated in too much literature using the conventional methods [26-32], and the process usually uses pre-extracted algae oil as raw material [33-36]. To get the algal oil out of the microalgae cells, oil extraction, typically by an organic solvent, is needed. This step is expensive because it involves cell rupture, solvent extraction, oil/solvent separation and reuse, etc. So, the combination of the oil extraction and transesterification steps in one novel process named "reactive extraction" or "in-situ transesterification" is considered the available alternative that could minimize the biodiesel production cost [37-40]. It includes the simultaneous addition of the acid catalyst (e.g. conc. H2SO4) and pure alcohol (e.g. methanol) to dried powder microalgal biomass. The

Research Journal of Chemical and Environmental Sciences Res J. Chem. Environ. Sci. Vol 2 [4] August 2014: 10-21 Online ISSN 2321-1040 CODEN: RJCEA2 [USA] ©Academy for Environment and Life Sciences, INDIA Website: www.aelsindia.com/rjces.htm

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alcohol extracts the microalgae lipids "oil" and, catalyzed by the acid, concurrently transesterifies the extracted lipids to produce fatty acid methyl esters, FAME "biodiesel" [41, 42]. In-situ transesterification of microalgae to produce biodiesel provides benefits rather than the conventional method, and summarized in Table 1 [6, 13, 22, 38].

Fig 1: Transesterification reaction equation The reactive extraction method was first demonstrated by Harrington and D’Arcy-Evans [40] using sunflower seeds as raw material, and an improvement in biodiesel yields up to 20% compared to the conventional process were done by these authors. This conversion improvement was considered to be attributable to the improved accessibility of the oil in the biomass by the acidic medium. The in situ transesterification "reactive extraction" of macerated sunflower seeds was also studied by Siler- Marinkovic et al. [43] who investigated two temperature levels of 30°C and 65°C and a range of test reaction conditions: the alcohol (methanol)-to-oil molar ratio varied from 100:1 to 300:1, and the sulphuric acid catalysts concentration ranged from 16% to 100% (on the mass-basis of the oil) and a reaction time of 1 - 4 h. Under these conditions, the best biodiesel yields was 98.2%, based on the oil content of the sunflower seeds and obtained at oil-to-methanol of 300:1, an acid catalyst concentration of 100% and 1h as a reaction time. Also, this process was used by E.A. Ehimen et al. [39] on Chlorella microalgae oil using methanol containing 100% (wt/wt oil) sulphuric acid as a catalyst. Steps involved with producing biodiesel from algal biomass are shown in Fig 2.

Table 1: Comparison of in-situ method with traditional one for biodiesel production Item No. In-Situ Transesterification Extraction-Transesterification 1 Very simple process in operation Complex process 2 Avoided lipids loss Lipid loss during operation 3 High yield Low yield 4 Minimize the wastewater pollutants Large wastewater quantities that

pollutes the environment 5 Heating value is high Low heating value 6 Less time consuming process High time consuming process 7 Production cost is low Production cost is relatively high

The ultimate goal of this research is to study the engineering variables that affecting the biodiesel production from Spirulina-platensis microalgae using a novel process "reactive extraction" that combines the oil extraction and esterification/transesterification in a single step. This ultimate goal is to be realized with the specific objectives of (1) characterizing the microalgae oil of the selected strain, (2) optimization of the main reaction factors (catalyst concentration, oil-to-alcohol molar ration, agitation influence, processing time and temperature) that strongly affect the cost of reactive extraction process for algal biodiesel production, and (3) investigating systematically the properties of the produced biodiesel in a comparison with the EN 14214 standards.

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Fig 2: Illustrative scheme of algal biodiesel production Experimental Background Materials Microalgae biomass Dried microalgae of Spirulina-Platensis strain were supplied from Microbiology Department, Soils Water and Environment Res. Inst., Agricultural Research Center, Giza, Egypt. The cultivation system was similar to that used in Ref. [1]. Conditions of 30oC, fluorescent lamps of 3.5klx and pH of 8.5±0.5 were used for its cultivation. Reagents Sulphuric acid (98% purity) as a catalyst in the trans-esterification process and, Methanol (99.9% purity) as the reacting alcohol were used in this study. All chemicals were purchased from El-Nasr Pharmaceutical Chemicals Co. Experimental Procedure First of all, Spirulina-platensis oil was extracted using the Soxtherm extraction system described by El-Shimi, H [1] and Jie Sheng et al. [44]. An optimization study on the microalgae oil extraction was performed by El-Shimi, H [1]; to determine the total lipids content on dry basis. Then, the extracted oil was analyzed for characterization. Different sulphuric acid, H2SO4 concentrations (30, 50, 100 and 200% wt./wt. oil), were used throughout this study. The amount of catalyst is weight out by the electronic balance, as well as the amount of microalgae biomass. Step 1: Preparation of catalyst- alcohol solution The amount of methanol (CH3OH) is measured in a test-tube and poured into Erlenmeyer flask. The pre-weighted amount of catalyst is added to the flask. The acid-methanol solution is freshly formed by dissolution of H2SO4 into methanol on a magnetic stirrer for 5 min. The solution was prepared freshly; in order to maintain the catalyst activity. Step 2: Mixing of reactants Fifteen grams (15gm) of microalgae biomass were previously dried at 110oC; to remove water traces, and then added carefully to the acid-alcohol mixture in the agitated vessel. The vessel was then heated and maintained at the desired temperature. At this moment, reactive extraction process (simultaneous extraction and transesterification reaction) starts. Step 3: Reactive extraction process In this step, it takes place simultaneous extraction and transesterification reaction "in-situ transesterification process" explained previously in the theoretical part. Where the catalyst/alcohol solution attacked the triglycerides (oil) in the Spirulina-platensis strain and cleaved off a fatty acid chain. The amount of oil in the biomass reacted with methanol in presence of H2SO4 and biodiesel with glycerin are produced. Other non desirable products like soap can be produced in the reaction, altering methyl esters yields. This chemical reaction is given itself for the required time. Time is also changed to study the effect in biodiesel yield. The reaction time studied is from 2 - 10 h. The major reactive extraction processes and product purification steps used are shown in Fig 3.

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Step 4: Separation of products When the reaction is finished, the warm reaction mixture was allowed to cool for 20 min. the resulting mixture is removed from the vessel, then filtered and the residues are washed three times by re-suspension in methanol; to recover any traces of FAME product left in the residues and then poured into a 500ml separating funnel. Water (60ml) was added to the filtrate; to facilitate the decantation of the hydrophilic components of the extract. Further extraction of the FAME product was achieved by extracting three times for 15 min using 60 ml of hexane, which resulted in generation of two layers: hydrophobic layer (hexane, FAME and glycerides), and hydrophilic layer (water, glycerol and excess methanol). The separation can take about 12 hours to carry out correctly.

Fig 3: Schematic diagram of biodiesel production from Spirulina-platensis microalgae used in this research

When the product has fully settled, two distinct layers were separated. These two layers are biodiesel and glycerin. The biodiesel on top looked as a clear, lighter in color and thin. The glycerin settled to the bottom looked clear, darker amber color and thick. Most of the settling occurred within the first hour. The reaction was demonstrated to be successful by observing the glycerin settling by gravity in the bottom. Once the glycerol and biodiesel phases have been separated, the bottom layer which contains glycerol,

Algae Residues

Filtrate

Hexane

Water

Spirulina-platensis Powder

"Acidic Methanol"

Biodiesel

Hydrophobic layer: Hexane + Biodiesel

Water layer: glycerol + catalyst

methanol

Glycerol

Reaction Processing

Reaction mixture (Biodiesel, glycerol, algae

biomass, methanol, catalyst)

Filtration

Decantation

Biodiesel Cleaning

Fractional Distillation

Hexane

Evaporation Methanol

Dissolution Methanol Conc. H2SO4

Water Dirty water

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trace water, catalyst, and excess methanol was drawn into a pre-weighted beaker and dissolved in pure water; to purify the glycerol layer, and then subjected to a flash evaporation process, in which excess alcohol and water are removed. The recovered alcohol was recycled and reused. Now, the layer contains only the by-product glycerol and the catalyst, therefore the weight of pure glycerol can be detected by the well-known catalyst weight. This procedure was performed in each experiment of the work, since we took a 15 g of microalgae biomass in each experiment, which expected to contain lipids of 1.6425 g, and based on just 60% reaction conversion, around 0.99 g glycerol will be obtained and can be weighted; using four digits balance. Step 5: Biodiesel cleaning When biodiesel and glycerin are clearly separated, the biodiesel layer was washed with water and filtered into a clean, dry side-arm flask; to evaporate the methanol and the hexane using a fractional distillation apparatus. The amount of collected biodiesel is difficult to be measured; because it mixed with the unreacted-glycerides. With the forward reaction resulting in FAME production and the process is near to completion, the weight of the purified glycerol as a co-product is expected to increase until a constant value, signifying an equilibrium conversion of the microalgae lipids to the methyl esters. Engineering factors affecting the reactive extraction process

i. Effect of reaction time and temperature Fifteen grams (15g) of powder microalgae was mixed with 80ml methanol containing 0.82ml H2SO4 at various temperatures (27, 40, 50 and 65oC) with continuous stirring at 650rpm. At each of the interest temperature, the reactive extraction process is given itself for the required time (2, 4, 8 or 10 h); to investigate the effect in biodiesel yield and it was repeated in duplicate. The glycerol purification and its weight were carried out as explained previously.

ii. Effect of catalyst concentration and oil-to-alcohol molar ratio Spirulina-platensis powder of 15g was mixed in the reaction vessel with different alcohol volumes (40, 60, 80 and 100ml) that corresponding to (1857:1, 2786:1, 3714:1 and 4643:1 respectively), containing various acid-catalyst concentrations (30, 50, 100 and 200%) on the mass-basis of microalgae oil content, this was performed at 65oC for 8h with 650rpm continuous agitation, assumed as an optimum conditions. A minimum volume of 40 ml methanol was selected since it was the suitable amount that facilitated a complete submersion of 15 g of biomass. The respective FAME products and the co-product glycerol at different investigated variable levels were obtained and their weights determined. Also the in situ transesterification reaction was performed at the same conditions without catalyst; to provide a greater insight on the effect of the catalyst presence in the transesterification process.

iii. Effect of agitation The agitation is an important engineering factor that affecting the reaction conversion; because it facilitate a complete suspension of the particles in the reaction vessel. So, the reactive extraction process was run with and without agitation for comparison, while the other variables kept constant; 15g of microalgae was mixed with 80ml methanol containing 0.82ml H2SO4 as a catalyst at 65oC for 8h. The reaction co-product (glycerol) was purified and its weight was determined. Analytical method Fatty acids composition of the extracted algae oil was determined using gas chromatographic analysis of the oil ethyl esters. Modification of the oil to its ethyl esters was made using 2% H2SO4 as catalyst in the presence of dry ethyl alcohol in excess. The chromatographic analysis was made using Hewlett Packard Model 6890 Chromatograph. A capillary column 30 m length and 530 μm inner diameter, packed with Apiezon® was used. Detector temperature, injection temperature and the column temperature were 280°C, 300°C and 100°C to 240°C at 15°C/min, respectively. RESULTS AND DISCUSSION Characterization of pure microalgae oil From the optimization study on the microalgae oil extraction that done by El-Shimi, H [1], Spirulina-platensis oil was determined to be 0.11g/g of biomass. The oil was characterized and compared to ASTM standards as shown in Table 2; since the extracted oil may be used as a fuel without further processes in case, if it has low viscosity.

Table 2: Characterization of algae oil and a comparison with the standards Characteristics Spirulina-platensis Oil ASTM biodiesel’s Standards Density (g/cm3) 0.892 0.86 – 0.90 Viscosity (cp , 40oC) 58 3.5 – 5.0 Acidity (mg KOH/g oil) 37.4 Max. 0.5

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The results obtained denote satisfactory need to the oil transesterification process; to reduce its viscosity, while the density is in compliance with the standards as illustrated in Table 2. The acid value of the oil was 37.4 mg KOH/g oil, which is very higher; therefore the choice of acidic over alkaline catalysis for the reactive extraction process was justified. The fatty acids composition of Spirulina-platensis oil is detected via GC-MS and tabulated in Table 3. It's clear that the predominant fatty acid was the palmitic acid C16:0 (49.6% by mole), which makes the oil a promising feedstock for biodiesel fuel synthesis. The percentages of the major fatty acids in the Spirulina used in this study were in accordance with previous works by other authors [45-47] the principal fatty acids present were palmitic, gamma-linolenic and linoleic acid as reported in Table 3. The fatty acid profiles extracted by Bligh& Dyer, Folch, methanol, ethanol or by static hexane are expected to be similar [48], even when the total amounts of lipid extracted differed.

Table 3: Fatty acids profile of Spirulina-platensis oil measured by GC-MS Fatty acid % in sample (by mole) Mystic (C14:0) 22.67

Palmitic (C16:0) 49.58

Palmitoleic (C16:1) 2.75 Stearic (C18:0) 5.56 Oleic (C18:1) 2.24 Linoleic (C18:2) 5.03 Linoleuic(C18:3) 7.41 Ecosanoic (C20:0) 1.06 Eicosic (C20:1) 3.69

The average molecular mass of Spirulina-platensis oil and that of biodiesel were calculated to be 845.19 and 284 g/gmole respectively with the same procedure published by H.I.El-Shimi et al. [5]. It's well-known that, the reaction yield is calculated from “Equation 1”, which needs to the amount of produced methyl esters. And that can be calculated from the stiochiometric equation of the transesterification reaction (Fig 1), by knowing the weight of glycerol detected as from the experimental work mentioned previously.

퐵푖표푑푖푒푠푒푙 푌푖푒푙푑 % =푊푒푖푔ℎ푡 표푓 푏푖표푑푖푒푠푒푙

푊푒푖푔ℎ푡 표푓 푚푖푐푟표푎푙푔푎푒 표푖푙 푥 100 (1)

3.2. Effect of catalyst concentration on biodiesel yield In this study the reactive extraction process was investigated at four catalyst loadings (30%, 50%, 100% and 200% H2SO4 wt/wt algae oil content) as illustrated in Table 4 and presented in Fig 4. Higher yields of microalgae methyl esters were computed to be 84.716% with ±9% standard error using 0.82 ml H2SO4 (100% by wt. of oil) dissolved in 80ml methyl-alcohol at 65°C for 8 hr, with further increase in catalyst concentration the conversion efficiency more or less remains constant. These results agree with El-Shimi, H [5] and also with the methanolysis using 100% H2SO4 (wt./wt. of oil) resulted in successful conversion of Chlorella oil giving the best yields and viscosities of the esters by E.A. Ehimen et al. [39].

Table 4: Catalyst concentration effect on the biodiesel yield H2SO4 Vol. (ml) H2SO4 Conc. (%) Biodiesel Yield (%)

0.25 30 55.142 0.41 50 77.23 0.82 100 84.716 1.64 200 84.716

The use of statistical analysis by Excel-program allowed expressing the amount of methyl esters produced as a polynomial model [49]. As observed in Fig 4, the concentration of acid catalyst has significant effect on the response (biodiesel yield) and the data are fitted to a second order with determination coefficient of 0.99 as expressed in "Equation 2".

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푌 = − 0.002 푋 + 0.736 푋 + 39.8 , 푅 = 0.99 (2) From the results published in Fig 4, the optimum catalyst concentration is around 100% wt./wt.oil. This result is compliance with previous results obtained by Vicente et al. (1998), and also by A. Bouaid et al. [50] on jojoba oil.

Fig 4: Investigation of catalyst concentration effect on biodiesel yield (at 65oC for 8h using 80ml methyl-alcohol)

Effect of oil-to-alcohol molar ratio on biodiesel yield The oil-to-methanol molar ratio is an important engineering factor affecting the conversion of microalgae lipids to biodiesel. This factor was tested in the range of 1:1857 to 1:4643 as reported in Table 5, while the other variables remain constant. The stiochiometric balance of the trans-esterification reaction that illustrated in Fig 1 requires three moles of methanol and one mole of triglycerides to yield three moles of fatty acid methyl esters (microalgae biodiesel) and one mole of glycerol [5, 51]. However, transesterification is an equilibrium reaction in which an excess of alcohol is required to drive the reaction to the right [1, 50, 52]. The studied range includes and exceeds that of a similar investigation of the in situ transesterification of Chlorella oil by E.A.Ehimen [39], and that of sunflower oil by Siler- Marinkovic and Tomasevic [43]. The biodiesel yield % was computed based on the total weight of pure glycerol obtained as mentioned before, since maximum yield achieved was 84.7%, at 65oC during 8h reaction time using 0.82ml acid catalyst dissolved in 80ml alcohol volume and continuous agitation at 650rpm, concerning experimental and standard error of ±3.5%. The results are presented in Fig 5 and indicate an improvement of the microalgae oil conversion with methanol increase. Dependence of microalgae biodiesel on the reacting alcohol volume was investigated to be a polynomial model of second order as shown in with 99.5% statistical coefficient of determination (R2 = 0.995). However, with the use of alcohol volumes over 80 ml (i.e. a reacting oil-to-alcohol molar ratio of 1:3714) for the reactive extraction of 15 g microalgae biomass, no significant trends were observed for the FAME yields.

Table 5: Effect of oil-to-alcohol molar ration on microalgae biodiesel yield Alcohol Vol. (ml) Oil-to-Alcohol Molar Ratio (1:X) Microalgae Biodiesel Yield (%)

40 1857 73.2 60 2786 81.79 80 3714 84.7

100 4643 84.7

0

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0 50 100 150 200 250

Biod

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Acid Catalyst Conc. %

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Fig 5: Dependence of Biodiesel Yield % on Oil-to-Alcohol Molar Ratio

Effect of reaction time and temperature on biodiesel yield Reactive extraction of Spirulina-platensis microalgae was carried out at various temperatures (27-65oC) over range of reaction time (2-10hr), while the other conditions kept constant using the measured weight of pure glycerol as a conversion indicator for the yielded biodiesel. Really, the elevated temperatures improve the initial miscibility of the reacting species, leading to a significant reduction in the reaction time, as published by all concerned scientists and researchers [5, 32, 53, 54]. The progress of the in-situ transesterification of microalgae oil to biodiesel of this study is reported in Table 6. For the samples investigated at room temperature (no process heating), asymptotic FAME yield % was not reached within the time boundaries of the study. According to Table 6, when the reactive extraction process was carried out at 65°C under continuous stirring at 650rpm, using 0.82ml concentrated H2SO4 as a catalyst dissolved in 80ml methyl alcohol, higher equilibrium conversion levels of biodiesel of 76.22% and 84.7%were attained after reaction time of 4 h and 8 h respectively. Within the investigated experimental conditions, equilibrium of FAME conversions was observed to reach similar asymptotic values after a reaction time of 8 and 10 h for temperatures of 50 and 65°C as shown in Fig 6. While at lower temperatures of 27°C, the process was incomplete and no microalgae biodiesel was observed. Although faster conversion rates could be observed by use of reaction temperatures higher than the boiling point of the reacting alcohol (say, 80°C), the process heating and pressure requirements may inhibit the use of such temperature levels, and when we consider the total energy consumption and the economical situation of the whole biodiesel production system, the reaction temperature of 65°C may prove more beneficial.

Table 6: Influence of reaction time and temperature on biodiesel yield Reaction Time (hr) Biodiesel Yield (%)

27oC 40oC 50oC 65oC 2 1.35 25.11 38.2 43.1 4 10.62 45.81 70.5 76.22 8 30.22 62.3 81.54 84.7

10 34.71 62.512 81.63 84.82

y = -2E-06x2 + 0.020x + 44.39R² = 0.995

60

65

70

75

80

85

90

0 1000 2000 3000 4000 5000

Mic

roal

gae

Biod

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l Yie

ld (%

)

Oil-to-Alcohol Molar Ratio (1:X)

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It's observed from Fig 6, temperature has detectable effect on the ultimate conversion to ester and the data are tend to a polynomial of second degree with high correlation factor (R2 >97%).

Fig 6: Effect of reaction time at different temperatures on biodiesel yield %

However, higher temperatures decrease the time required to reach maximum conversion of microalgae lipids, and the best reaction conditions seem to be 65°C for 8 h. Effect of agitation on microalgae biodiesel yield It's sure that, the agitation is a potential engineering factor affecting the biodiesel production process; since when the reactive extraction of microalgae was performed without stirring, no reaction occurred and zero biodiesel yields was obtained. This indicates that agitating is required to enhance the reaction progress, evidently by aiding the initial miscibility of the reacting species. However, the reaction was carried out at 65oC using 80ml methanol and 100% H2SO4 (wt/wt oil) as catalyst, and the mixture was stirred at 650rpm for 8hr intermittently (1h on and 1 h off) to obtain just 74.3% as a biodiesel yield, and also it subjected to continuously agitation to achieve 84.7% of biodiesel, which prove the positive influence of agitation during reaction. Evaluation of produced algal biodiesel Once biodiesel is produced, some of the important quality control parameters are measured and compared with the EN 14214 standards. These parameters such as density, viscosity, flash point, pour point, cloud point and cetane number have acceptable values and are in agreement with the petroleum diesel standards as presented in Table 7. The viscosity is an important property that influencing the process conversion; as the viscosity of produced biodiesel is 4.8 cSt, and that of microalgae oil was 58 cSt, also it affects the fuel injection through the equipment. Viscosity and density of algal biodiesel are confirming the standards as illustrated in Table 7.

Table 7: Evaluation of algal biodiesel compared to EN 14214 standards Property Value Limits Density (g/ml at 15oC) 0.886 0.86-0.90 Viscosity (cSt at 40oC) 4.8 3.5-5.0 Cetane number 61 Min. 51 Flash point (oC) 172 Min. 101 Cloud point (oC) 5 - Pour point (oC) -1 -

1.35

10.62

30.2234.71

25.11

45.81

62.3 62.512

38.2

70.5

81.54 81.63

43.1

76.22

84.7 84.82

y = -0.199x2 + 6.706x - 11.84R² = 0.995

y = -0.853x2 + 14.80x - 0.650R² = 0.997

y = -1.342x2 + 21x + 3.703R² = 0.973

y = -1.375x2 + 21.09x + 8.884R² = 0.978

0

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20

30

40

50

60

70

80

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100

0 2 4 6 8 10 12

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ld %

Reaction Time (hr)

27C

40C

50C

65C

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Cetane number of biodiesel is always higher than petrol diesel; because it has saturated molecules and longer fatty acids carbon chains. The measurement of cetane number is important; since lower cetane numbers result in poor ignition quality, delay and excessive engine knock. Cetane number of microalgae biodiesel was measured to be 61, which is better than 38 for jatropha methyl-esters [55] and higher than 46 for rapeseed biodiesel [56]. The flash point of produced microalgae biodiesel was estimated to be 172°C. this value exceed the standard and it's high compared to 160°C for jatropha biodiesel [23] and much higher than that of 58°C for petrol-diesel, which makes the biodiesel-based algae, and its blends safer fuels to handle and store near ignition sources. The cold flow properties "cloud and pour points" of algal biodiesel are measured to be "5 and -1" respectively. These values are not good compared to that given in literatures for biodiesel from waste vegetable oils (3 and -6 respectively) and that for sunflower biodiesel (2 and -3 respectively); the reason is that, algal methyl-esters composed of saturated fatty acids as estimated in Table 3, and as reported by El Shimi, H et al, [5] , Lin Rulong et al, [57] and Alan Scragg [58], the unsaturated fatty acids give better cold flow properties than saturated fatty acids. CONCLUSIONS From this research, it may conclude that:

Biodiesel can be produced from plant oils, animal fats and waste cooking oils. The demand of biodiesel production is increasing every year, and oil crops are compromising food crops. So, other sources of biodiesel such as microalgae will have to be commercialized.

Oil extraction from microalgae is one of the most important processes that determine the sustainability of algae-based biodiesel. Lipid content of Spirulina-platensis was detected to be 11% (by wt.). Fatty acid analysis report that C16:0 is the dominance in Spirulina-platensis oil that makes it a promising feedstock for biodiesel production.

Acid value and viscosity of microalgae oil are very high (37.4 mg KOH/g oil and 58 cp, respectively), so the choice of acid-catalysis reactive extraction process was justified to get biodiesel from Spirulina-platensis microalgae.

Reactive extraction process "in-situ transesterification" is that combines the steps of lipids extraction and transesterification to produce biodiesel in single step.

The results of biodiesel production at the studied conditions using reactive extraction methodology show that a reaction time of 8hr at 65oC using 100% concentrated sulphuric acid (wt/wt microalgae oil) as a reaction catalyst dissolved in 80ml methanol (1:3714 oil-to-alcohol molar ratio) agitated continuously at 650rpm, were enough to get high yield of biodiesel (84.7%).

Microalgae biodiesel is close to the practical diesel; as its properties (density, viscosity, flash point, cloud point, pour point and cetane number) confirm the EN 14214 standards.

ACKNOWLEDGEMENTS Financial support from Faculty of Engineering, Cairo University-Egypt [FECU] is appreciated. Also National Research Center [NRC] at Giza-Egypt is gratefully acknowledged for technical information determination. REFERENCES 1. Hassan El-Shimi, "Biodiesel production from Spirulina-platensis microalgae using reactive extraction

methodology". A master thesis, Chemical Eng. Dept., Cairo University, 2013. 2. Demirbas, A. (2009). "Production of Biodiesel from Algae Oils." Energy Sources Part a-Recovery Utilization and

Environmental Effects 31(2): 163-168. 3. Chisti, Y. (2007). "Biodiesel from microalgae." Biotechnology Advances 25(3): 294-306. 4. Michael B. Johnson, " Microalgal Biodiesel Production through a Novel Attached Culture System and Conversion

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Reactive Extraction of Microalgae for Biodiesel Production ...aelsindia.com/rjcesaugust2014/2.pdfUnder these conditions, the best biodiesel yields was 98.2%, based on the oil content