This study aimed to quantify sugars and oils in algae through pretreatment and extraction. Algae samples were pretreated with dilute acid at 160°C to break down cell walls and release sugars, which were then quantified by HPLC. Oil was extracted from separate samples using hexane in a Soxhlet apparatus. Results found very low concentrations of xylose and glucose sugars, less than 10% of algae mass. Oil content was also low, less than 3% of mass. The algae used was thus not suitable for biofuel production due to insufficient sugar and oil yields.

1. Identification and Quantification of Sugars and Oils in Algae
Blake Johnson, Ching-Shuan Lau and Dr. Danielle Julie Carrier
Department of Chemical Engineering, Department of Biological Engineering
April 26, 2010
Background:
• Plant material wall is composed of cellulose and hemicellulose
(Figure 1).
• Cellulose and hemicellulose have to be broken down into their basic
monomers (glucose and xylose) before being fermented into any
biofuel or biobased product.
Figure 1: Molecular Structure of cellulose and hemicellulose
• Currently, ethanol production through lignocellulose is not cost
effective. The process is very delicate and more expensive than
producing ethanol through corn or producing gasoline. The process
needs to be reduced in cost before lignocellulosic ethanol
production can be considered.
• Green algae (Figure 2) could be a possible source of carbohydrates
that can be transformed into various biobased products.
Figure 2: Dried green algae harvested from Noland Water Treatment
Facility (Fayetteville, AR)
Project Objectives:
• Release the cell wall material contained in algae through dilute acid
pretreatment and enzymatic hydrolysis.
• Quantify the carbohydrates, the by-products (like formic acid,
hydroxymethylfurfural (HMF) and furfural) and the crude oil present
in the Noland Water Treatment Plant algae.
• Determine if the carbohydrate profile is dependent on seasonal
growth (May – September 2010)
Research Impact:
• Increase ethanol and biodiesel production from algae by refining
the process of extracting sugars and oil.
Materials:
Procedure:
Sugar extraction
1. Measure 2 gram of dried algae and mix in reactor tube with 20 mL of 1% sulfuric acid
solvent.
2. Reactor tubes was placed in sand bath at 160 oC for 1 h.
3. After the reaction, allowed the sample to cool, neutralized to pH 7 and filtered.
4. Analyze the samples by HPLC to identify and quantify the sugars as well as the by-
products (formic acid, HMF and furfural) in pretreated algae samples.
Oil Extraction
1. Place 3 g of dried algae in cellulose Thimble.
2. Extract with hexane in Soxhlet for 2 h to extract oil from algae.
*Keep cooling water running to condense hexane vapor and prevent Soxhlet from
running dry.
3. Allow hexane to evaporate, leaving only extracted oil in flask.
*Let flask and algae sit for 2 days to vaporize the remaining hexane.
4. Measure the gain in mass of the flask after the extraction to determine the amount
of oil collected on the flask after extraction.
Figure 3: Reactor tubes used to
hydrolyze algae in dilute acid
hydrolysis by targeting the
break down of hemicellulose
into xylose monomer.
Figure 4: Sand bath used for the dilute
acid hydrolysis pretreatment at 160o C
Figure 5: HPLC (High
Performance Liquid
Chromatography) used to
identify and quantify the sugars
and by-products in algae.
Figure 7: Soxhlet used to extract oil
from algae. The sample shown on the
right turned yellowish indicating the
presence of oil, as compared to the
sample on the left that remained clear.
Results:
Discussion:
• Algae samples contained xylose and glucose in very small concentration (<10% of
the overall mass).
• Algae samples harvested in June and July contained oil, as shown by the yellowish
color of hexane solvent (Figure 7). However, the mass gained from the oil is not
significantly differently from algae of other months that had no visible oil present
(Table).
• The oil content in algae is less than 3% mass of the overall algae mass, which is
lower than algae used for biofuel that typically contains 30% oil.
• Based on the amount of oil and sugar extracted, this particular type of algae is
not recommended for use in biofuel production.
Conclusion:
• Algae sample used in this study contains very small quantity of oil and sugars in
the form of xylose and glucose.
Mg present in 2000 mg of Algae May June July August September
Liquid Fraction Glucose 69.69 24.63 24.47 - 24.21
After dilute acid Xylose 33.63 43.96 40.83 - 40.15
pretreatment Formic Acid 120.45 75.07 82.00 42.67 169.40
Acetic Acid 55.03 64.34 35.94 15.37 88.13
Furfural 1.83 1.42 0.89 0.20 1.50
Hydroxyl methyl
furan (HMF) 1.98 2.25 1.52 0.70 2.97
Solid Fraction Glucose 37.79 47.75 36.57 - 45.74
After enzymatic Xylose 2.33 35.30 27.42 - 32.12
hydrolysis
Total Glucose 107.48 72.38 61.04 - 69.96
Xylose 35.96 79.25 68.25 - 72.27
Formic Acid 120.45 75.07 82.00 42.67 169.40
Acetic Acid 55.03 64.34 35.94 15.37 88.13
Furfural 1.83 1.42 0.89 0.20 1.50
Hydroxyl methyl
furan (HMF) 1.98 2.25 1.52 0.70 2.97
Yield (g sugar / g algae) Glucose 5.4% 3.6% 3.1% - 3.5%
Xylose 1.8% 4.0% 3.4% - 3.6%
May June July August September
Mass of oil (mg) in recovered
in 3 gram of algae
30.00 62.60 ± 10.75 21.00 51.30 ± 0.42 40.50 ± 7.78
% Oil Recovered / gram algae 1.0% 2.1% 0.7% 1.7% 1.3%
Figure 6: Water bath used for the
enzymatic hydrolysis of pretreated
algae pellet ( 55 oC for 24 hrs).
* Only has only valid set of results. The second set of results has an inaccurate flask reading.
3rd Annual FEP Honors
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