Accepted Manuscript

Non-catalytic liquefaction of microalgae in sub-and supercritical acetone

Binbin Jin, Peigao Duan, Caicai Zhang, Yuping Xu, Lei Zhang, Feng Wang

PII: S1385-8947(14)00724-4
DOI: http://dx.doi.org/10.1016/j.cej.2014.05.137
Reference: CEJ 12232

To appear in: Chemical Engineering Journal

Received Date: 28 October 2013

Revised Date: 19 May 2014
Accepted Date: 30 May 2014

Please cite this article as: B. Jin, P. Duan, C. Zhang, Y. Xu, L. Zhang, F. Wang, Non-catalytic liquefaction of
microalgae in sub-and supercritical acetone, Chemical Engineering Journal (2014), doi: http://dx.doi.org/10.1016/
j.cej.2014.05.137

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 Non-catalytic liquefaction of microalgae in sub-and supercritical acetone

Binbin Jin, Peigao Duan*, Caicai Zhang, Yuping Xu*, Lei Zhang, Feng Wang

Department of Applied Chemistry, Henan Polytechnic University, No. 2001, Century Avenue, Jiaozuo, Henan 454003, P.R.

China

Opening Laboratory of Alternative Energy Technologies, Henan Polytechnic University, No. 2001, Century Avenue, Jiaozuo,

Henan 454003, P.R. China

ABSTRACT

In the present study, a microalga (Chlorella pyrenoidosa) was treated in sub/supercritical acetone

in the absence of catalyst by using a high pressure bath reactor. Influence of process variables such as

temperature (varied from 170 to 350 °C), acetone/microalga ratio (varied from 2/2.5 to 16/2.5), and

time (varied from 5 to 120 min) on the yields of product factions and properties of biocrude has been

studied. Temperature was the most influential factor affecting the products yield and properties of the

biocrude, and the highest biocrude yield of 60.1 wt.% was achieved at 290 °C. Addition of acetone not

only promoted the conversion of microalga but also favored the biocrude yield due to the incorporation

of acetone into the biocrude. Furthermore, liquefaction of microalga in acetone made the conversion

milder that than of in water. The biocrude was less viscous than that of oil produced from hydrothermal

liquefaction under otherwise identical reaction conditions. The biocrudes, which contained significant

carbon and hydrogen than that of the original algal biomass, had higher heating values ranging from

28.7 to 37.1 MJ/kg. The most abundant compounds for the biocrude are unsaturared fatty acids

(9,12,15-octadecatrienoic acid) and hydrocarbons (2-hexadecene, 3,7,11,15-tetramethyl-). CO2 was the

dominant component in the gaseous products under all experimental conditions. Deoxygenation and

deoxygenation are necessary if one to expect to produce transportation fuels from this kind of biocrude.

Keywords: Liquefaction; microalgae; Chlorella pyrenoidosa; sub- and supercritical acetone; biocrude

*Corresponding author. Tel: +86(0391) 3986820; fax: +86(0391) 3987811

E-mail address: pgduan@hpu.edu.cn (P. Duan); xuyuping@hpu.edu.cn (Y. Xu)

1
1. Introduction

In recent years, public criticisms have been raised against the use of biofuels derived from the

food crops, most prominently based on arguments related to the environment, land-use, and public

nutrition [1, 2]. Motivated by such issues, there has been a big push for the production of biofuels from

non-food plants and other biomatters. Microalgae, which are unicellular organisms grown in water and

do not divert crops from consumption to energy usage, may help to alleviate such concerns. Growing

algae for biofuel production not only provides a sustainable source of alternative energy, but also

benefits the environment and reduces arable land use conflicts [3-5].

To date, different methods were developed for converting microalgae into biofuels. They can be

basically classified into two categories: thermochemical (e.g. direct combustion, gasification,

liquefaction, and pyrolysis) and biochemical (e.g. anaerobic digestion, alcoholic fermentation, and

photobiological hydrogen production) conversion [6, 7]. Of those different conversion technologies,

hydrothermal liquefaction (HTL) was considered as one of the most promising ways because it can

readily accept moist or wet biomass, thereby obviating the feedstock dewatering and drying [8-11].

Furthermore, the resulting biocrude has a lower moisture compared to the pyrolysis oil which typically

contains approximate 25-50 wt.% moisture [9, 12,13]. However, only 40 wt.% of carbon and 35 wt.%

of hydrogen in the microalgae feedstock were converted into the biocrude, and a considerable amount

of organics remained in the aqueous phase, and thus resulted in a relatively lower energy recovery [14].

Moreover, the biocrude was very viscous and rich in oxygen and nitrogen, making it hard to handle and

more likely to deteriorate when stored over a long period of time. To improve the yield and quality of

the biocrude, organic solvents such as tetralin, 1-methyl naphthalene, toluene, methanol, ethanol, 1,4-

dioxane, and ethylene glycol[15-19] were employed in the liquefaction of microalgae. All these

solvents showed a significant positive effect on the yield and quality (e.g. lower density and viscosity)

of the biocrude. Furthermore, using organic solvents could shift the liquefaction to a milder

temperature compared to the HTL, thereby reducing the capital cost. More recently, the authors have
2
compared the thermo-chemical liquefaction of Chlorella pyrenoidosa (C. pyrenoidosa) in eleven

different solvents [20]. The results suggested that the solvent polarity significantly affected the

conversion rate of C. pyrenoidosa and the biocrude yield, and higher biocrude yield was always

achieved in those solvents with strong polarity such as ethylene glycol, ethanol, acetone, and ethyl

acetate. Supercritical acetone (Tc=235°C, Pc=4.8MPa) showed the highest biocrude yield of 57.0 wt.%.

However, no further work has been done to determine the effect of other variables such as temperature,

time, and algae/solvent ratio on the yields of product fractions and properties of the biocrude when

employing acetone as the liquefaction medium. These details are expected to provide more insights on

the role of acetone in the liquefaction of microalgae.

This is an extension work of Duan et al.[20] with a focus on the liquefaction behavior of C.

pyrenoidosa in acetone. Particular attention will be given to optimize the reaction temperature,

acetone/microalga ratio (A/M), and time. Finally, the physical and chemical properties of the biocrude

were characterized by using Gas chromatography-Mass spectroscopy (GC-MS), elemental analysis,

and Fourier transform infrared spectroscopic analysis (FT-IR), respectively.

2. Materials and methods

2.1Materials.

C. pyrenoidosa powder rather than its paste was used in present study due to its easy shipment and

storage, which was obtained in the form of non-cracked cell walls from

Shandong Binzhou Tianjian Biotechnology Co., Ltd. (North China). This microalga contains 10 wt.%

moisture,19 wt.% crude lipid, 52 wt.% crude protein, respectively. More details about this microalga

are available in previous publication [20].

A custom made stainless-steel autoclave, which has an internal volume of 58 mL, was used to

perform all experiments. The reactor was seasoned by water at 400 °C for 4 h to eliminate or

significantly reduce the catalytic effect of the reactor wall to the liquefaction reaction. The reactor was

heated by using a molten-salts bath that consists of potassium nitrate and sodium nitrate at a mass ratio
3
of 5:4.

2.2 Liquefaction

Typical experiments were performed at a microalga loading of 2.5 g and acetone loading of 10 mL.

The air inside the reactor was displaced with helium by flashing the reactor with He. No Further He

was charged. The 1atm of helium that remained served as an internal standard for the quantification of

gas yields. The loaded reactor was placed into a molten-salts tank pre-heated to the desired temperature

to initiate the reaction. The reaction temperature was controlled by an Omega temperature controller.

Reaction time was defined as the period of time when the pre-set operating temperature was first

achieved to the time as the reactor was taken out of the molten-salts bath. The pressures inside the

reactor under different reaction conditions are provided as supporting information (see Table S1), is

temperature and A/M ratio dependent. Increasing temperature and A/M ratio increases the pressure

inside the reactor. After the desired reaction time, the reactor was taken out of the molten-salts tank and

immersed into a cooled water bath for about 10 min to terminate the reaction. The cooled reactor was

thoroughly dried by an electric hair dryer and weighed before and after collecting the gas to estimate

the gas formation. The gas was collected for some experiments for components analysis. The reactor

was then opened. Dichloromethane was added to recover the biocrude fraction. The dichloromethane

extract and solid residue were separated by filtration. Details about the isolation of biocrude and solid

residue were similar as previously reported [20]. The separated solid residue was dried in an oven at

110 °C for 12 h, and then weighed. The dichloromethane was removed from the extract by using a

rotary evaporator. To estimate the possible residual solvent amount, a control experiment was

performed wherein a flask containing pure dichloromethane alone was treated to this evaporation

procedure. The mass of residual solvent in the control experiment was close to zero, suggesting that

almost no solvent was remained in the biocrude. The remaining material was biocrude. The yield of

each product fraction was calculated as its mass divided by the mass of microalga powder loaded into

the reactor
4
 At least duplicate independent runs were conducted under each set of conditions to estimate the

uncertainties in the experiments. Results reported herein represent an average of two or three

replications. Uncertainties are reported as the experimentally determined standard deviations.

2.3 Analysis

GC-7900 (Shanghai Techcomp Scientific Instrument Co., Ltd.) gas chromatograph equipped with

a thermal conductivity detector (TCD) was used to analyze the gas products. A 15-ft×1/8-in. i.d.

stainless steel column, packed with 60×80 mesh Carboxen 1000 (Supleco) separated each component

in the mixture. Argon (column pressure of 0.18MPa) served as the carrier gas for the analysis. Two

consecutive analyses of the gas mixture were taken for each reactor. The temperature of the column

was held at 70 °C for 120 minutes. The mole fraction of each gaseous component was determined via

calibration curves generated from analysis of the analytical gas standards with known composition.

The amount of helium added to the reactor was used as an internal standard to determine the molar

amount of each constituent. The yield of each gas species was calculated as its molar amount divided

by the mass of dry microalgae loaded into the reactor.

A Gas chromatograph-Mass spectrometer (7890A GC; Agilent Technologies) equipped with a

capillary column (DB-5MS, 30 m length, 0.25 mm I.D., 0.10 µm film thickness) was used to realize the

compound identification in the biocrude. The biocrude samples were prepared as 10±1wt.% solutions

in dichloromethane. The injector temperature was set at 300 °C. 2 µL of biocrude sample was injected

with a split ratio of 3:1. A two-minute solvent delay was set to protect the filament. The column was

initially held at 40 °C for 4 min. The temperature was ramped to 300 °C at 4 °C· min-1 and held

isothermally for 4 min, giving a total runtime of about 73 minutes. Helium flowing at 3 mL· min-1

served as the carrier gas. A NIST mass spectral library was used to identify the compounds.

The elemental composition of the biocrudes was determined using Elemental analyzer (Elemental

Vario ELⅢ CHNS/O) [20]. The Dulong formula (Heating value (MJ·kg-1) = 0.338C + 1.428(H-O/8) +

5
0.095S) was used to estimate the higher heating value (HHV) of the biocrudes based on their weight %

of each element.

FT-IR was performed on a Vertex 70 FT-IR Spectrometer (Bruker Optics Corporation) to

determine the functional groups in the microalga, solid, and biocrude. 1 g of sample was mixed with

200 mg of KBr powder. The pellet preparation involves grounding the sample and KBr using an agate

mortar and pestle and using a hydraulic press and die to create a thin and transparent disk. FT-IR

spectra (resolution: 4cm-1, scan: 254, range: 4,000-400 cm-1) were taken at a controlled ambient

temperature (25 °C). A background spectrum was also collected under identical conditions.

3. Results and discussion

We first investigate the influence of experimental temperature (170-350 °C), A/M ratio (2/2.5-

16/2.5), and reaction time (5-120 min) on the products yield, and then give a closer look to the

properties of biocrude in terms of elemental and molecular composition.

3.1 Effect of temperature on the yields of product fractions

C. pyrenoidosa was converted to three fractions including biocrude, gas, and solid residue after it

was processed in acetone. To determine the influence of reaction temperature on the yields of product

fractions, experiments were conducted by varying the temperature from 170 to 350 °C with other

conditions fixed (A/M ratio of 4/2.5, 60 min batch holding time). The critical point of acetone is at a

temperature of 235 °C and pressure of 4.8 MPa. Therefore, the liquefaction reactions were conducted at

sub- and supercritical acetone, respectively. We selected this temperature range because it represents

that used in previous work on the thermo-chemical conversion of C. pyrenoidosa in ethanol [20]. The

A/M ratio was defined as the ratio of acetone (mL) to the microalga mass (g). Fig.1 shows all the

results. Under the subcritical conditions of acetone (T<235 °C), microalga conversion is not complete,

which is reflected by high solid and low biocrude yields. Increasing temperature increases the

conversion which reaches the highest value at 310 °C, and thereafter it increased back due to the char

formation from cyclization, condensation, and re-polymerization of oil intermediates. The solid
6
obtained at low temperatures resembles unconverted microalga as indicated by elemental analysis and

FI-IR and discussed later (see section 3.42 and 3.43). The C. pyrenoidosa used in present study

contains 19 wt.% lipid, 52 wt.% protein, and 10 wt.% carbohydrate which exhibited typical

decomposition temperature ranges of 150-230 °C, 230-400 °C, and 400-500 °C, respectively [21].

Therefore, steep decrease in solid products is observed as the temperature increased from 200 to 250 °C

because most of the lipids and proteins were decomposed at this temperature range. The biocrude yield

(17.3 wt.%) obtained at 170 °C is close to the lipid content (19 wt.%) originally contained in the

microalga. Increasing temperature to 230 °C led to a major breakage of the cells [22], and thus

significantly increases the biocrude yield. The highest biocrude yield of 60.1 wt.% is achieved at

290 °C. Above 290 °C, the biocrude yield starts decreasing, mainly as a result of the further

polymerization of intermediates to high molecular weight compounds remaining in the solid residue.

Cracking of the biocrude was also accelerated at more severe temperatures, thereby promoting the

production of light end products which could not be recovered as oil during the solvent vaporization

process due to their high volatility. Similar biocrude yield trend as a function of temperature was also

observed for the liquefaction of C. pyrenoidosa in ethanol [20]. Higher conversion rates were also

observed when liquefying the pinewood and oil palm fruit press fiber in acetone [23, 24]. However, the

biocrude yields from those two biomass feedstocks were both significantly lower than that of from

microalgae, more likely due to their deficient lipid and protein contents. The gas yield increases from 4

to 8 wt.% as the temperature increased from 170 to 250 °C due to the continually degradation of lipid,

protein, and carbohydrate, and levels off at 250 °C. Constant gas yield at higher temperatures might

due to two aspects: 1). positive effect from the temperature increase offset the negative effect from the

pressure increase on the gas formation at temperatures above 250 °C; 2). lower detection limit (0.1 g)

of the balance used to weigh the gaseous products. CO2 was typically present in the highest amount,

which was always the dominant component for the hydrothermal processing of microalgae [13]. The

gas yield mainly consists of CO2 (28.2 mmol/g) at the lowest temperature of 170 °C. Increasing
7
temperature not only significantly increases the yield of CO2 but also increases the amounts of H2, CO,

and CH4 (see Fig.S1). CO2 can be formed from reactions such as steam reforming and water gas shift,

which also produce hydrogen. Methane formation could be via the methanation reaction. Ammonia

was detected in any of the experiments, suggesting the nitrogen atoms released from the microlaga

during liquefaction being in the form of ammonia. The total yield of all products fractions decreased

with increasing temperature, especially at temperatures above 300 °C. The mass balance was 97 wt.%

at 170 °C but declined to 85 wt.% at 350 °C. Recall, the alga has 10 wt.% moisture, and thus the

overall mass balance after the liquefaction reaction should be equal to 90 %. The mass balance

exceeding 90 % at temperatures between 170 and 300 °C was due to the incorporation of acetone in the

reaction. With increasing the temperature, the consumed amount of acetone was smaller than that of the

material loss during the sample transferring and handling, thereby decreasing the mass balance below

90 wt.%.

3.2 Effect of A/M ratio on the yields of product fractions

Effect of A/M ratio on the yields of product fractions was investigated by conducting reactions at

290 °C for 60 min with changing the A/M ratio from 2/2.5 to 16/2.5. Variation of A/M ratio was

realized by changing the acetone loading volume at a fixed loading amount (2.5 g) of alga. Fig.2 shows

that the yield of solid products decreases from 29.7 to 18.7 wt.% as the A/M ratio increased from 2/2.5

to 12/2.5. It seems that organic solvent plays an important role in the decomposition of biomass,

possibly due to its easy penetration into the solid biomass particle. Further increase in A/M ratio

slightly increases the solid yield, possibly due to the contribution coming from the acetone. At lower

A/M ratios, the algae did not form a well-mixed suspension in the reactor due to the limited amount of

acetone, which would result in unfavorable mass- and heat-transfer conditions inside the reactor,

limiting the liquefaction and solvolysis reactions, and thus reducing the biocrude yield. At higher A/M

ratios, the microalga was easily distributed in the solvent phase, which could increase the mass and heat

transfer, thereby resulting in higher conversion [25, 26]. However, large amount loading of acetone has
8
its downside, namely a significant increase in operating temperature (see Table S1) and cost. The yield

of biocrude steadily increases from 52.7 to 80.1 wt.% as the A/M ratio increases from 2/2.5 to 12/2.5.

Further increase in A/M ratio has almost no effect on the biocrude yield. The highest biocrude yield

(80.1wt.%) was even comparable to the organics content (~81 wt.%) of the microalga, suggesting again

that acetone was incorporated into the biocrude during the liquefaction reaction. In the liquefaction of

biomass, ionic and radical reactions were the primary reactions [27]. Increasing the loading amount of

polar solvent dilutes the concentration of the intermediate products and thus decreases the possibility of

the cross linked reactions and reverse reactions, thereby increasing the biocrude yield. The gas yield is

not affected with increasing the A/M ratio from 2/2.5 to 10/2.5. Further increase in A/M ratio decreases

the gas yield. At lower A/M ratios, the reaction behaved like pyrolysis which would produce more

gases than that at higher A/M ratios [28]. Increasing solvent to biomass ratio also decreased the gas

yield when liquefying powdered poplar wood in water [29]. However, contrary results were observed

in the liquefaction of C. pyrenoidosa in ethanol [20]. The only difference in these two experiments was

the temperature (350 vs 290 °C for the present study). Possibly, the gas suppression from the pressure

increase was predominant at lower temperatures, thereby decreasing the gas yield. In addition to

ammonia, the gas yield mainly consists of H2, CO, CH4, and CO2 at an A/M ratio of 2/2.5. Increasing

A/M ratio significantly increases the amount of CH4 and CO2. The yields of H2 and CO are

independent to the A/M ratio (see Fig.S1).

3.3 Effect of reaction time on the yields of product fractions

Fig.3 shows the yield of different product fractions from the thermo-chemical processing of C.

pyrenoidosa at 290 °C and A/M ratio of 10/2.5 with varying the holding time from 5 to 120 min. It is

noted that the pre-heating time was about 20 min, during which large proportion of the microalga were

decomposed as a lower solid yield was achieved at a reaction time of 5 min. The solid yield slightly

decreases with increasing the reaction time and hits the lowest value of 20.0 wt.% at 60 min. Further

increase in reaction time would gradually increase the solid yield due to the tar formation from the
9
condensation or re-polymerization of the oil. As inferred from Fig.3, reaction time has a more

pronounced effect on the biocrude yield than that of solid products. The biocrude yield increases from

63.7 to 78.9 wt.% as the reaction time increased from 5 to 60 min. Further extending the reaction time

decreases the biocrude yield due to the increasing chances of the secondary and tertiary reactions of

biocrude, suggesting the biocrude reached its saturation point at a reaction time of 60 min. Similar yield

trend of biocrude was also observed when liquefying the C. pyrenoidosa in supercritical ethanol at

different reaction time [20]. The gas yield kept constant within the time range examined in this study,

indicating gas formation was entirely a thermo-controlled process. The gas phase consists mainly of

CO2, CO, and CH4. CH4 becomes significant at longer reaction time of 120 min (see Fig.S1).

It is worth notice that the mass balances under all experimental conditions are larger than that of

the organic content (~81 wt%) in the microalga, suggesting acetone was not only a solvent but also a

reactant during the liquefaction reaction. The same results were also observed in the liquefaction of

pinewood and oil palm fruit press fiber in acetone [23, 24].

3.4 Characterization of biocrude

3.4.1 GC-MS analysis

The inlet temperature of the GC was 300 °C. At this temperature, only part of the material in the

oil samples was volatilized, as the rest had a higher boiling point. Thus, the data provided are only for

the volatile fraction and are not necessarily representative for the total oil.

Fig. 4 compares the total ion chromatograms of biocrudes produced from Run 1, Run 7, Run 11,

Run 15 (see run numbers in Table 1), respectively. Clearly, reaction conditions also significantly

influenced the product distribution of the biocrude. The biocrude produced at a temperature of 170 °C

showed fewer major peaks prior to 40 min than that of biocrude produced at higher temperatures such

as 290 and 370 °C, respectively. Since the GC column separates largely on the basis of boiling point or

volatility of compounds, that is, those compounds with lower boiling point will elute from the column

earlier. Therefore, increasing temperature favored the cracking reactions, and thus increased the
10
proportion of low-boiling range species in the biocrude. This outcome suggests that temperature plays a

vital role in determining the compounds distribution of the biocrude, which is consistent with previous

study [20]. Furthermore, the biocrude obtained from a larger A/M ratio showed very similar

chromatogram as that produced at a smaller A/M ratio, but increased the concentration of particular

compounds.

To gain a better understanding of the characteristics and properties of the biocrudes, a mass

spectral library and computer matching were used to facilitate the compound identification. The area %

for each compound identified was defined by the percentage of the compound’s chromatographic area

out of the total area. Therefore, the area % only represents the comparative or semi-quantitative content

of each component in the biocrude. Table S2 shows the tentative identities of the major (peak area at

least 0.4 % of the total) individual molecular components in the biocrudes as shown in Fig.4. The

identified compounds in the biocrudes mainly include N-heterocyclics, ketones, un-saturated

hydrocarbons, saturated and unsaturated fatty acids, and fatty acid amides. In contrast, the fatty acid

esters are the largest portion identified in biocrudes produced from the thermo-chemical processing of

C. pyrenoidosa in alcohols [20]. Significant differences in products distribution were also observed

from the thermo-chemical conversion of pinewood and oil palm fruit press fiber in acetone [23, 24].

These previous studies suggested that solvent type and biochemical composition of biomass played a

very important role in determining the products category and their relative proportion in the biocrude.

As inferred from Table S2, the most abundant compounds for the biocrude are unsaturated fatty acids

(9,12,15-octadecatrienoic acid) and hydrocarbons (2-hexadecene, 3,7,11,15-tetramethyl-), likely due to

the direct decomposition of triglycerides and dehydration of phytol, respectively. Significant quantity

of 9,12,15-octadecatrienoic acid was also detected from the thermo-chemical conversion of C.

pyrenoidosa in ethanol at a temperature of 170 °C for 60 min [20], suggesting thermal decomposition

was the dominant reaction at lower temperatures. Increasing temperature decreased the relative

proportion of unsaturated compounds and increased the abundance of nitrogen-containing compounds

11
in the biocrude. The higher content of nitrogen-containing compounds in the biocrude was due to the

higher protein content (65 wt.%) in the starting microalga. Increasing temperature favored the

conversion of protein, and thus increased the nitrogen content in the biocrude. Increasing the solvent

loading increased the content of ketones (e.g. 3-Penten-one, 4-methyl) and derivatives of pyridine in

the biocrude and decreased the relative proportion of fatty acid amides. 3-Penten-one, 4-methyl was not

detected in the biocrude produced from the thermochemical conversion of C. pyrenoidosa in ethanol,

indicating that it might be derived from the interaction of acetone with the biocrude.

3.4.2 Elemental analysis

The biocrude produced from the thermo-chemical conversion of C. pyrenoidosa in acetone is less

viscous (on the basis of visual inspection of flowability) than that of oil produced from the HTL under

otherwise identical reaction conditions. Possibly, the incorporation of acetone in the biocrude reduced

the viscosity. The ultimate analysis and HHV of the biocrudes produced at different reaction conditions

along with the starting material are listed in Table 1. In addition, the ratios of H and chemical energy in

the biocrude relative to those quantities in the microalgal feedstock for processing at different reaction

conditions are also provided.

Table 1shows that increasing temperature from 170 to 350 °C led to an increase in carbon content

from 67.2 to 73.4 wt.%. The hydrogen content of biocrude except that produced at 170 and 200 °C

increases from 9.1 to 9.8 wt.% as the temperature increased from 230 to 290 °C, and thereafter, it

maintained constant at around 9.7 wt.%. The abnormal higher hydrogen content of biocrude produced

at 170 and 200 °C might due to the partial reaction of microalgae. Microalgae usually contain a high

proportion of nitrogen due to its presence in chlorophyll and in protein. For the microalga in our

experiments, the nitrogen content is 8.4 wt.%. Increasing temperature initially increases the nitrogen

content in the biocrude due to the gradual conversion of protein. The highest nitrogen content of 9.4

wt.% was observed at 290°C. Further increase in temperature decreases the nitrogen content, likely due

to the in situ denitrogenation of biocrude at more severe temperatures. Consistently, GC-TCD analysis
12
of the gas phase suggested that ammonia was present. An increase in temperature reduces oxygen

content, indicating deoxygenation proceeded during the liquefaction reaction. The net effects for the

thermo-chemical conversion of microalgae are that the carbon and hydrogen content of the biocrude

increased, the oxygen content was significantly reduced. Thermo-chemical conversion significantly

increases the energy densification from 22.6 MJ/kg for the algal biomass to around 37.1 MJ/kg for the

biocrude. The H/C molar ratio was nearly constant over the temperature range of 250 to 300 °C, and

then a reduction was observed with further increase in temperature, likely as a result of increasing

content of aromatic compounds in the biocrude [22]. Furthermore, the N/C and O/C molar ratios

showed similar trends as that of N and O as a function of temperature. A ratio of unity denotes recovery

of the same number of atoms or amount of chemical energy in the products that was originally present

in the feedstock. Table 1 shows that increasing temperature significantly improves the energy recovery

into oil. The maximum energy recovery of 0.96 was obtained at 290 °C. Further increasing temperature

adversely affected the energy recovery due to the difficult recovery of light ends in the oil fraction. The

last column also shows the energy recoveries with the energy inputs of acetone included. Clearly, the

energy recovery considering acetone was much smaller than that the energy recovery without

considering acetone, suggesting only small amount of acetone was incorporated in the biocrude.

The biocrude produced at A/M ratio of 2/2.5 presented the lowest carbon and hydrogen and

highest oxygen, it also exhibited the lowest carbon and hydrogen and highest oxygen among all of the

biocrudes. The carbon content decreases while the oxygen content increases as the A/M ratio increased

from 4/2.5 to 16/2.5. Therefore, lower energy density of biocrude was also observed at larger A/M

ratios. Increasing the A/M ratio however, resulted in a biocrude with improved pour behavior (on the

basis of visual inspection of flowability). The hydrogen and nitrogen content of biocrude was

insensitive to the A/M ratio. Table 1 shows that the energy recovery significantly increases with

increasing the A/M ratio until the highest value of 1.17 is achieved at an A/M ratio of 12/2.5, and then

levels off. Energy recoveries exceeding unity attributed to the incorporation of acetone in the biocrude.
13
The energy recovery considering acetone shows the similar trend as the energy recovery without

considering acetone.

The carbon and hydrogen content increases while the nitrogen and oxygen content decreases with

increasing the reaction time, suggesting denitrogenation and deoxygenation was promoted with

increasing the reaction time. In Table 1, the energy recovery increases from 0.89 to 1.16 as the reaction

time increased from 5 to 70 min, and thereafter, it maintained almost constant. Higher HHV of

biocrude was always achieved at a longer reaction time.

Although higher biocrude yield and energy recovery were achieved when processing microalgae

in acetone, the high nitrogen content of biocrude is an important drawback in view of fuel applications,

as the nitrogen will lead to NOx when burned. Furthermore, the nitrogen in the biocrude has potential

poisons for the catalyst during the processing. Similarly, some deoxygenation might be necessary to

decrease the possible corrosion of biocrude to the reactor system. Therefore, denitrogenation and

deoxygenation are necessary if one to expect to produce transportation fuels from this kind of biocrude.

3.4.3 FT-IR

FT-IR analysis was performed to determine the structure of the microlaga and liquefaction products

(solid and biocrude) obtained after the liquefaction. Fig. 5a, 5b, and 5c show the FT-IR spectra of solid

and biocrude derived from the liquefaction experiments at different reaction conditions.

Since the KBr to sample ratio is 200:1, relative differences between heights can be taken as indication

for relative differences in concentration of the corresponding function group between the samples. As

shown in Fig.5a, algae structure is dominated by protein (N-H at 3300 cm-1, C=O at 1600-1690 cm-1,

C-N stretching and NH bending at1480-1575 cm-1) [30] and lipid [C=O at1740 cm-1, C-H at 2900 cm-1]

[31]. The solid obtained at the lowest temperature of 170 °C shows low peak height at 3300 cm-1,

possibly due to the reaction of protein and lipid. As the temperature increases, the protein and lipid

bands decrease. The broad bands around 3300 cm-1 and 1600-1690 cm-1 at 350 °C are significantly

reduced, suggesting most of the protein and lipid were converted. Fig. 5b and 5c show the FT-IR
14
spectra of biocrude produced at different temperature and A/M ratios. Compared to the biocrude

produced at 170 °C, the biocrude produced at 350 °C shows strong absorbance at 2900 cm-1 which

correspond to C-H stretch in methyl and methlyene. The biocrude produced at an A/M ratio of 16/2.5

presents much higher absorbance at 1600-1750 cm-1 than that of biocrude produced at a lower A/M

ratio of 2/2.5, indicating the incorporation of acetone into the biocrude.

4. Conclusion

Liquefaction of microalgae in acetone had a beneficial effect on the yield and flowability of the

biocrude and made the conversion milder that than of in water. The biocrude had a much higher energy

density that that of the original algal biomass, but it also had a high viscosity and contained significant

quantity of nitrogen. Therefore, upgrading for denitrogenation and viscosity reduction will be required

if this kind of oil can be used as transportation fuels. Nevertheless, this study showed that microalgae

can be effectively converted into oil in acetone in the absence of any catalysts.

Acknowledgement

We gratefully acknowledge financial support from the Henan Polytechnic University (B2011-008)

and from the Program for New Century Excellent Talents in University (NCET-13-775).

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18
Fig.1 Effect of temperature on the yields of product fractions at 60 min and A/M ratio of 4/2.5.

Fig.2 Effect of A/M ratio on the yields of product fractions at 290 °C and 60 min.

Fig.3 Effect of time on the yields of product fractions at 290 °C and A/M ratio of 10/2.5.

Fig.4 Total ion chromatograms for biocrudes produced from Run 1, Run 7, Run 11, and Run 15.

Fig.5 FT-IR spectra for microalga, solid and biocrude obtained at different reaction conditions

19
Fig.1

Solid Oil Gas

100
4.0
6.0 6.0 8.0 8.0
8.0 8.0 8.0 8.0
17.3 8.0
80 8.0
25.0
Yield(wt.%)

44.0

60 52.9
58.2 59.3 60.1
59.6 58.8 54.8 50.3

40
75.6
64.3

45.3
20
34.4
27.9 27.1 26.9 25.4 25.7 26.5
24.4

0
170 200 230 250 270 280 290 300 310 330 350
Temperature(°C)
Fig.2

Solid Oil Gas

8.0 6.0 4.0 4.0

100
8.0
8.0
8.0
8.0
80
Yield(wt.%)

60 78.9 80.1 80.2

60.1 66.6 80.1
52.7 73.5

40

20
29.7 26.9 24.4 20.7
20.0 20.0 18.7 19.3

0
2/2.5 4/2.5 6/2.5 8/2.5 10/2.5 12/2.5 14/2.5 16/2.5
A/M ratio (mL/wt.)
Fig.3

Solid Oil Gas

8.0 8.0 8.0 8.0 8.0

100 8.0
8.0 8.0 8.0

80
Yield(wt.%)

63.7 66.4 73.6 76.2 74.3

60 70.1 78.9 76.8 75.9

40

20
29.7 28.0 24.9 25.8 24.2 22.7 25.8
20.0 21.2

0
5 10 30 40 50 60 70 90 120
Time(min)
Fig.4
2.50E+07
Run 1 2.50E+07 Run 7

2.00E+07 2.00E+07

1.50E+07 1.50E+07

1.00E+07 1.00E+07

5.00E+06 5.00E+06

0.00E+00 0.00E+00
2 15 28 41 54 67 80 2 15 28 41 54 67 80
Time(min) Time(min)

3.00E+07 Run 11 3.00E+07 Run 15

2.50E+07 2.50E+07

2.00E+07 2.00E+07

1.50E+07 1.50E+07

1.00E+07 1.00E+07

5.00E+06 5.00E+06

0.00E+00 0.00E+00
2 15 28 41 54 67 80 2 15 28 41 54 67 80
Time(min) Time(min)
Fig.5
(a) Algae Run 1-solid Run 11-solid (b) Algae Run 12-oil Run 18-oil
100
100
90 90

80

% Transi,ittance
80
%Transmittance

70
70
60
60
50
50 40

40 30

20
30
4000 3500 3000 2500 2000 1500 1000 500
4000 3500 3000 2500 2000 1500 1000 500
Wave numbers(cm-1) Wave number (cm-1)

(c) Algae Run 1-oil Run 11-oil

100

90

80
%Transmittance

70

60

50

40

30

20
4000 3500 3000 2500 2000 1500 1000 500
Wave numbers(cm-1)
Table 1 Elemental composition (wt.%) and HHV of biocrudes produced from the conversion of C.
pyrenoidosa in acetone under different reaction conditions.

HHV E biocrude E biocrude

Run
Conditions C H N Oa H/C N/C O/C (MJ/kg
numbers E algae E algae + E acetone
)
C. pyrenoidosa 46.8 6.9 8.4 16.9 1.76 0.154 0.275 22.6 -
Temperature(°C) (60 min, 2.5 g microalgae, and 4 mL acetone)
1 170 67.2 9.6 4.3 18.9 1.72 0.055 0.211 33.1 0.25 0.09
2 200 68.0 9.7 6.3 16.0 1.71 0.079 0.177 34.0 0.38 0.14
3 230 68.1 9.1 8.1 14.7 1.60 0.102 0.162 33.4 0.65 0.24
4 250 68.3 9.4 8.8 13.4 1.65 0.111 0.148 34.1 0.80 0.29
5 270 70.3 9.7 9.4 10.8 1.65 0.114 0.115 35.5 0.92 0.34
6 280 70.5 9.7 8.6 11.2 1.65 0.105 0.119 35.7 0.94 0.34
7 290 70.9 9.8 8.2 11.1 1.65 0.099 0.118 35.9 0.96 0.35
8 300 70.8 9.7 8.3 11.1 1.66 0.101 0.118 35.9 0.95 0.35
9 310 71.9 9.7 7.9 10.4 1.63 0.094 0.109 36.3 0.95 0.35
10 330 72.6 9.7 7.9 9.8 1.60 0.093 0.101 36.7 0.89 0.33
11 350 73.4 9.7 7.9 9.0 1.59 0.092 0.092 37.1 0.83 0.30
A/M (mL/g) (290°C, 60 min, and 2.5 g microalgae)
12 2/2.5 61.8 8.4 7.1 22.8 1.62 0.099 0.276 28.7 0.67 0.25
7 4/2.5 70.9 9.8 8.2 11.1 1.65 0.099 0.118 35.9 0.96 0.34
13 6/2.5 69.8 9.1 7.0 14.1 1.57 0.086 0.152 34.1 1.00 0.37
14 8/2.5 69.0 8.8 7.4 14.9 1.53 0.091 0.162 33.2 1.08 0.40
15 10/2.5 68.3 8.9 6.8 16.0 1.57 0.085 0.176 33.0 1.15 0.42
16 12/2.5 67.3 9.2 7.6 15.8 1.64 0.097 0.177 33.1 1.17 0.43
17 14/2.5 67.9 8.8 7.4 15.9 1.55 0.094 0.175 32.6 1.16 0.42
18 16/2.5 66.8 9.1 7.7 16.5 1.63 0.099 0.185 32.6 1.16 0.42
Time(min) (290°C, 2.5 g microalgae, and 10 mL acetone)
19 5 66.1 8.7 7.4 17.8 1.58 0.096 0.202 31.6 0.89 0.33
20 10 66.7 8.6 7.1 17.6 1.54 0.091 0.198 31.6 0.93 0.34
21 30 66.2 9.0 6.5 18.3 1.63 0.084 0.207 32.0 0.99 0.36
22 40 66.5 9.0 6.5 18.0 1.63 0.083 0.203 32.2 1.05 0.38
23 50 67.9 8.9 6.2 16.9 1.57 0.079 0.187 32.7 1.10 0.40
15 60 68.3 8.9 6.8 16.0 1.57 0.085 0.176 33.0 1.15 0.42
24 70 69.7 9.3 6.4 14.6 1.60 0.079 0.157 34.2 1.16 0.43
25 90 70.3 9.2 6.3 14.2 1.57 0.077 0.151 34.4 1.16 0.42
26 120 71.1 9.4 6.2 13.3 1.59 0.074 0.141 35.1 1.15 0.42
a. Calculated by difference

20
> Microalga can be effectively converted into oil in acetone in the absence of catalyst

> Temperature significantly affects the products yield and properties of the biocrudes

> Liquefaction of microalga in acetone made the conversion milder that than of in water

> Acetone promoted the conversion of microalga and benefited the biocrude yield

> The biocrudes had higher heating values ranging from 28.7 to 37.1 MJ/kg

21
 Graphical Abstract

Chlorella pyrenoidosa

Sub/super-critical
acetone

Biocrude
Autoclave

Extraction and separation