3.1 Synthesis of biodiesel from sunflower seeds
Oil Extraction: Initial experiments were performed using as the solvent: hexane, methanol, or a 50:50 mixture of methanol:hexane. In the last two cases, after sonicating and centrifuging the samples, a whitish layer deposited between the phases of the sample. Sunflower seeds contain mucilage which is a protective layer that some seeds have, and it is extracted along with the oil when extraction is done with polar solvents. Similar behaviour was observed by Baümler et al. [23,24]. They studied and compared the ethanolic extraction of oil from sunflower collets with the extraction with hexane, and they proved that ethanol has a higher extraction power than hexane. Ethanol showed great ability to extract sugar, phospholipids and tocopherols, extracting over 75% of the initial sugar content. The mucilage that we extracted with methanol can be related to these types of compounds. This fact forced us to carry out the extraction only using hexane as a solvent.
Before addressing the synthesis of biodiesel, several experiments were performed with the aim of finding the optimal conditions to extract all the oil content present in sunflower seeds. Table 1 shows the main experiments performed for this extraction, varying the seed/hexane ratio (g/mL) as well as the sonication time. This table also includes the yield of extracted oil and compares it with what was expected for a fat content of the seed of 49.6%.
Table1. Sonochemical extraction of oil from sunflower seeds and hexane as solvent
Sample
|
Sunflower seed/hexane ratio
(g/mL)
|
Sonication time
(min)
|
Content
oil*
(g)
|
Extracted
oil
(g)
|
Oil
yield
(%)
|
1
|
6/30
|
1
|
2.98
|
2.81
|
94.3
|
2
|
6/30
|
3
|
2.98
|
2.65
|
88.9
|
3
|
6/30
|
6
|
2.98
|
2.71
|
90.9
|
4
|
9/30
|
1
|
4.46
|
3.50
|
78.5
|
5
|
9/30
|
3
|
4.46
|
3.30
|
74.0
|
6
|
12/30
|
1
|
5.88
|
4.18
|
71.1
|
7
|
12/30
|
3
|
5.88
|
3.40
|
57.8
|
*Amount calculated for an oil content of 49.6%
The results showed a higher yield (94%) when adding 6 g of seeds and 30 mL of solvent with 1 minute of sonication. The optimal ratio for oil extraction in sunflower seeds is 1/5. The table also shows the yields with different sonication times. Similar yields at 1, 3 or 6 minutes of sonication were observed. Therefore, with these reaction conditions, with only one minute of sonication, it is possible to extract more than 90% of the oil contained in the sunflower seed.
So far, there are few published works on ultrasonic-assisted extraction of sunflower seed oil. Studies on this raw material have mainly been directed to the extraction of other types of compounds, such as phenolics [25]. Milenković et al. [26] applied two ultrasonic-assisted oil extraction techniques to sunflower seeds. A modified Soxhlet apparatus with a ultrasonic generator and a classical batch reactor with ultrasonic generator were used in this research. With the first device, a positive contribution of 1-1.5% was observed in the extraction with the use of ultrasound. However, the extraction times are 6 hours for a seed/solvent ratio of 1/10. Using the second device, most of the oil was extracted in 20 minutes of exposure to ultrasound (> 95%) with the same ratio of 1/10 seed/solvent. Similar results were obtained by Moradi et al. working also with this type of seeds [27]. Using n-hexane as extracting solvent and a seed/solvent ratio 1/12, ultrasound frequency 24kHz and a time of 2 hours for a complete extraction. In our case we have used a seed/solvent ratio of 1/5 and only 1 minute of sonication and the extraction achieved 94.3% of the oil contained in the sunflower seed. The secret is to increase the applied ultrasonic power. ShalmashI [28], using tea seeds, showed that the oil extraction yield of tea increased when increasing the ultrasonic power. When the power was increased from 10 to 50W, the yield of tea seed oil was increased from 46.23% to 85.21%. However, a further increase in the power showed only a moderate rise in yield. Our experience tells us that the exposure time to ultrasound should be controlled and only an appropriate time applied, since longer exposure times can degrade the extracted oil (see Table 1). Most of the works found in the literature use hours to sono-extract the oil from the seeds. However, it is very important to control sonication time when using high power ultrasound. Senrayan et al. [29], using Carica papaya seeds, obtained a maximum oil recovery when ultrasound-assisted extraction was carried out at a 1/13 (g/mL) seed/solvent ratio, 80% amplitude of ultrasonic power and an extraction time of 7 minutes.
Taking these results into account, the production of biodiesel from sunflower seeds must be carried out in two steps: in the first one, hexane extraction is carried out with the subsequent filtration of the seeds, and in the second, after addition of NaOH and MeOH, the sono-transesterification reaction is carried out.
Transesterification reaction. In the transesterification stage, biodiesel synthesis experiments have been carried out at different concentrations of methoxide (0.83, 0.50, 0.25, and 0.00 M) and with different volumes of this (7.50, 5.00, and 2.50 mL) added to the oil mixture dissolved in hexane. In all experiments, the seed/hexane ratio starting amounts were kept constant: 6/30 (g/mL) and analysed qualitatively by TLC and quantitatively by HPLC. With this last method, the percentage of the ester with respect to the triglycerides present in the final mixture was obtained. Table 2 shows this conversion analysed by HPLC for a sonication time of 1 minute. The results show very high conversions in all cases, obtaining, in almost all experiments, percentages greater than 99%. Note that in the absence of a catalyst (sample 10), no conversion to FAME was observed. On the other hand, it should be remembered that when biodiesel is produced, it is very important to achieve a high conversion efficiency to FAME with the minimum amount of catalyst, both to avoid the formation of soaps and to simplify the washing process. Therefore, in view of the results, the conditions of sample 9 can be selected as the most suitable to obtain a high conversion rate to FAME in a sonoreaction time of only one minute.
Few are the published works on the production of biodiesel in situ from sunflower seeds. Siler-Marinkovic and Tomasevic [30] used sunflower seeds with an oil content of 55.6% to produce biodiesel in situ by acid catalysis, using methanol as a solvent and without applying ultrasound. In all their research they reached a biodiesel yield > 90%, although the reaction times were never less than 60 minutes. Siatis et al. [31] shortened conversion times to FAME to 30 minutes, using an ultrasonic bath and a mixture of hexane methanol (50mL/3mL) and 10g of seeds. The conversion yield was 93%. Georgogianni et al. [32], using low frequency ultrasonication (24kHz), alkaline transesterification of sunflower seed oil with a molar ratio of methanol to oil of 7/1, obtained high yields of methyl esters (95%) after a short reaction time (20 minutes). Gama et al. [33] also used in situ transesterification of sunflower seed oil with methanol on KOH as basic catalyst for the production of biodiesel. The activity of KOH was evaluated using an oil to methanol ratio of 1/90 without ultrasound. KOH was shown to be an active catalyst, leading to total conversion in biodiesel after 1 hour reaction time. The shortest reaction times were obtained by Zhen et al. [17] using methanol assisted by diethoxymethane (DEM) as a cosolvent without applying ultrasound in the process. When the in situ transesterification was carried out with a molar ratio of methanol to oil of 101.39/1 and a molar ratio of DEM/oil of 57.85/1; a product containing 97.7% FAME was obtained in a reaction time of 13 minutes. We have managed to shorten the reaction time to just 1 minute.
Table 2: Sono-transesterification, in two steps (1min for extration and 1min for transesterification), of sunflower for the production of biodiesel.
Sample
|
Sunflower
seed/hexane ratio
(g/mL)
|
Methoxide concentration
(M)
|
Methoxide
Volume
(mL)
|
Biodiesel
yield
%
|
1
|
6/30
|
0.83
|
7.5
|
----
|
2
|
6/30
|
0.83
|
5.0
|
99.7
|
3
|
6/30
|
0.83
|
2.5
|
99.8
|
4
|
6/30
|
0.50
|
7.5
|
94.2
|
5
|
6/30
|
0.50
|
5.0
|
98.8
|
6
|
6/30
|
0.50
|
2.5
|
99.7
|
7
|
6/30
|
0.25
|
7.5
|
99.6
|
8
|
6/30
|
0.25
|
5.0
|
99.5
|
9
|
6/30
|
0.25
|
2.5
|
99.3
|
10
|
6/30
|
0.00
|
7.5
|
0.00
|
3.2 Synthesis of biodiesel from soybeans
Oil Extraction: For the extraction of the oil contained in soybean seeds, hexane and methanol were also used as solvents, as well as a 50:50 mixture of these. In this case, the presence of mucilage was also observed, although in smaller quantities than in sunflower seeds. No amount of mucilage was observed when hexane was used as the solvent in the oil extraction process. Thus, although it was observed that the extraction process had a higher yield in the presence of methanol, it was decided to use only hexane as the solvent. Table 3 shows the main experiments performed for this extraction, varying the seed/hexane ratio (g/mL) as well as the sonication time. This table also includes the yield of extracted oil and compares it with that expected for a fat content of the seed of 18.1%.
Table 3. Sonochemical extraction of oil from soybean seeds and n-hexane as solvent
Sample
|
Soybean seed/hexane ratio
(g/ mL)
|
Sonication
time
(min)
|
Theoretical oil*
(g)
|
Extracted oil
(g)
|
Oil
Yield
(%)
|
1
|
3/30
|
1
|
0.54
|
0.32
|
59.3
|
2
|
6/30
|
1
|
1.09
|
0.46
|
42.2
|
3
|
9/30
|
1
|
1.63
|
0.68
|
41.7
|
4
|
12/30
|
1
|
2.17
|
0.92
|
42.4
|
5
|
12/30
|
3
|
2.17
|
0.83
|
38.2
|
*Amount calculated for an oil content of 18.1%
The yields shown in Table 3 are, in all cases, lower than those obtained in sunflower seeds. A limiting effect of the solvent (comparing experiments 2-4) was also observed since, although the amount of oil increases with the grams of seeds added, the yield remains constant. Comparing experiments four and five of this table, it is observed that increasing the exposure time to ultrasound does not improve the oil extraction yield. Probably the longest exposure time to ultrasound degrades the oil already extracted. The results also reveal that the proportions with which higher yields are achieved are 3:30 with one minute of sonication.
There are also few published works on the application of ultrasound for the extraction of oil contained in soybeans. Li et al. [34,35] applied 20kHz high-intensity ultrasound during extraction of oil from two varieties of soybeans. The highest yield result of 12.21g from 100g soybeans (19.2% oil content in soybean seeds) was obtained using 150mL of a 3:2 hexane to isopropanol mixture under 47.6W/cm2 and 3 hours of sonication. In our case, we chose to perform the extraction only in hexane as we already showed in the case of sunflower oil extraction: alcohols favour the extraction of phospholipids, pigments and sugars that could become part of the biodiesel by contaminating it. Kanitkar et al. [36] demonstrated that shorter extraction times are necessary when microwave radiation is used. They exposed a mixture solvent to a feedstock ratio of 3/1 to microwave radiation at different temperatures. The results showed that the oil yield changed with temperature and achieved a maximum value in a relatively short time (order of minutes). Maximum oil yields of 17.3% and 120 °C were achieved by microwave extraction. In our case, with only 1 minute, we have achieved a yield of 59.3% (see Table 3).
Transesterification reaction: After oil extraction, tests were carried out to obtain biodiesel by transesterification in situ in a single step. Since the presence of methanol is necessary for the synthesis of biodiesel and this solvent favours the extraction of mucilage, several experiments were carried out with different amounts of methanol (always less than 20% of the total volume). In all these tests, no traces of this substance were observed, so it was decided to continue with the synthesis in a single step. Several experiments were conducted in which the amounts of seed/hexane (2.5 g/25 mL) and 1min of sonication time remained constant. Two concentrations of starting methoxide (0.12 and 1 M) were used, and different volumes of both methoxide and methanol were added. Table 4 shows the amounts added in each experiment and the results obtained by TLC.
Table 4: Sono-transesterification in one step of soybean for the production of biodiesel. A same starting soybean/n-hexane ratio (2.5g/25mL) was used in all samples. 1min of sonication time.
Sample
|
Methoxide volume
(mL)
|
Methoxide concentration (M)
|
MeOH Volume
(mL)
|
MeOH total volumen
(mL)
|
Ester presence in
TLC
|
1
|
0.50
|
0.12
|
0.00
|
0.50
|
No
|
2
|
0.83
|
0.12
|
0.00
|
0.80
|
No
|
3
|
5.00
|
0.12
|
0.00
|
5.00
|
Yes*
|
4
|
0.50
|
0.12
|
4.50
|
5.00
|
No
|
5
|
5.00
|
1.00
|
0.00
|
5.00
|
Yes
|
6
|
2.50
|
1.00
|
2.50
|
5.00
|
Yes
|
7
|
1.25
|
1.00
|
3.75
|
5.00
|
Yes*
|
8
|
1.25
|
1.00
|
1.25
|
2.50
|
Yes*
|
9
|
0.62
|
1.00
|
0.62
|
1.25
|
Yes*
|
10
|
1.50
|
1.00
|
1.50
|
3.00
|
Yes*
|
* The formation of the ester was observed in TLC, although there are also unreacted triglycerides.
In view of the results shown in this table, only two experiments with higher NaOH content showed a significant amount of ester and lack of oil signal by TLC. Sample 5 (was chosen because it has a lower NaOH content than sample 6) was repeated three times and analysed by HPLC. In all cases, the amount of ester versus triglyceride in the biodiesel formed was greater than 99%.
The work of Kddiran et al. [37] was one of the first published on alcoholysis in situ of soybean seeds in different alcohols. These authors found that maceration before in situ alcoholysis caused an appreciable increase in the amount of the oil. With maceration, approximately 40% of the soybean oil was transferred to the methanol phase, and the methyl ester content of this oil reached 55% for 3 hours reaction time and ultrasonic absence. In a more recent work, Haas and Scott [38,39], using dry flakes of soybean, achieved an optimal transesterification in reactions containing 5g of flakes and 12mL of 0.10 N NaOH in methanol, 10 hours of reaction time and the absence of ultrasound. More recently, Tuntiwiwattanapun et al. [40] showed that Isopropanol is an effective solvent for use in soybean oil extraction. This solvent was used in the in situ transesterification process, with up to 85% of the biodiesel yield and a reaction time of 90 minutes. In none of the above works has ultrasound been applied in the transesterification process. If we compare these results with those shown by us, clearly ultrasound‐assisted reduces the time of biodiesel production from hours to 1 minute.
3.3 Analysis of fatty acids in oil and biodiesel
The oils extracted and biodiesel produced from the soybean and sunflower seeds were analysed by gas chromatography to check if the extraction and transesterification process using ultrasound altered their composition.
Figure 1 and Table 5 shows the oil and biodiesel chromatograms obtained from sunflower seeds.
Table 5: Fatty acid composition of oil and biodiesel obtained from sunflower seeds. tR is the retention time in minutes
Peak
|
tR (min)
|
Sunflower
oil (%)
|
Sunflower biodiesel (%)
|
A
|
12.963
|
6.115
|
5.360
|
B
|
16.477
|
3.039
|
5.399
|
C
|
17.624
|
31.167
|
21.809
|
D
|
19.364
|
59.430
|
67.068
|
E
|
24.497
|
0.248
|
0.364
|
The results of Table 5 show quite a significant difference between the majority peaks C and D. The conversion of oleic acid to linoleic acid probably occurs sonochemically. This conversion could be explained by the presence of OH radicals produced by an intense field of ultrasound [18]. On the other hand, the hydrogen abstraction reaction produced by these radicals in the presence of hydrocarbons is well known. This abstraction generates alkyl radicals that could explain the conversion of oleic acid into linoleic acid observed in sunflower samples.
As can be observed in Figure 2 and Table 6, in the case of soybeans, the fatty acid composition of the extracted oil and biodiesel are very similar and, therefore, ultrasound does not alter its composition.
Table 6: Fatty acid composition of oil and biodiesel obtained from soybean seeds. tR is the retention time in minutes.
Peak
|
tR (min)
|
Soybean
oil
(%)
|
Soybean biodiesel
%
|
A
|
12.962
|
12.387
|
13.248
|
B
|
16.476
|
3.990
|
3.701
|
C
|
17.617
|
22.637
|
21.688
|
D
|
19.355
|
52.569
|
52.957
|
E
|
21.380
|
8.282
|
8.285
|
F
|
24.492
|
0.134
|
0.120
|
From the results obtained in this work, the in situ sono-transesterification is a promising approach to synthesis biodiesel from seeds directly. Few studies on this subject are found in the bibliography. The majority have utilised liquid oil directly instead of solid oilseeds. Within the latter, still fewer were performed on transesterification in situ from edible oil seeds. The results published so far using an in situ transesterification with soybeans and basic catalysis required several hours to complete the transesterification reaction [37-40]. In our case, using a concentration of 0.5 M NaOH, a solvent/seed molar ratio of 638/1, a one-minute reaction time in the presence of an ultrasonic field of 20 kHz frequency, and a calorimetric power of 106 W, we have achieved a conversion rate greater than 99%.
The results with sunflower seeds were also good, although with the inconvenience of having to produce biodiesel through two steps. The best results found in the literature using in situ transesterification in the presence of an ultrasonic field with this type of seeds showed a conversion rate of 95% of methyl esters after approximately 20 min of reaction time [32]. In our case, a percentage higher than 99% has been achieved using room temperature and one minute of sonication.
The most significant of our results is having achieved such high biodiesel yields with just one minute of reaction simply by replacing pure methanol with a hexane/methanol mixture. These results offer us the possibility of designing flow reactors that, with very little residence time within an ultrasonic field, achieve very high biodiesel yields.
In terms of energy consumption, mechanical stirring usually requires higher energy consumption than ultrasonication. It was reported that ultrasonication only required one-third to half of the energy that was consumed by mechanical agitation [21,22]. Therefore, we think that the presence of an ultrasonic field in the process of biodiesel production from seeds has enormous potential when it comes to reducing production costs.