2.2 Spectrophotometric Apparatus
1. Vortex mixer.
2. Adjustable pipet(s), covering 20 μL to 1 mL.
3. Cuvettes—glass, quartz, or plastic.
4. 13 Â 100 mm glass test tubes or equivalent.
5. Glass marbles or caps to cover or plug the 13 Â 100 mm glass
test tubes.
6. Digital dry block, capable of maintaining 80 Æ 3
C (compatible with glass tubes).
7. Spectrophotometer, set to measure at 620 nm.
2.3 Liquid
Chromatography
Apparatus
1. Vortex mixer.
2. Plastic graduated pipet, 5 and 10 mL with 0.1 mL graduations
(see Note 2).
3. Plastic 2 L graduated cylinder or volumetric flask (see Note 2).
4. Screw-top vials and presplit screw caps compatible with the
HPAEC-PAD injector.
5. High-performance
anion-exchange
chromatography
(HPAEC) system equipped with pulsed amperometric detection (PAD) (see Note 3).
2.4 Spectrophotometric Analysis
Reagents and
Standards
Use 18.2 MegaOhm (MΩ) water for all preparations requiring
water. Carefully follow all relevant chemical handling and disposal
procedures.
1. 02% (w/w, specific gravity 1.6338 at 20
C) sulfuric acid, used
for sample two-step hydrolysis.
2. Prepare glucose standard and calibration verification standard
(CVS) stock solutions (these must be made up separately as the
CVS is used to check the accuracy of the calibration curve)
using a high-purity glucose standard. Weigh approximately
25 mg of glucose into a clean, tared, class A, 100 mL volumetric flask. Record the weight to the nearest 0.1 mg. Bring to
volume with water and cap, and mix thoroughly. The standard
and CVS working solution concentrations should be approximately 0.25 mg/mL glucose. Due to balance limits, start with
a higher glucose stock concentration, and dilute to 0.25 mg/
mL. Always prepare the standards and CVS from separate
stocks so that you can check the accuracy of your calibration
with the CVS. Glucose stock solutions can be stored in a freezer
(À20
C) for up to 2 months or 5 days in the refrigerator
(4
C).
3. Prepare a 0.5 M NaOH solution using sodium hydroxide pellets. Weigh out 4 g of sodium hydroxide pellets. Add about
50 mL of water to a 200 mL volumetric flask. Slowly add pellets
and swirl to dissolve. The mixture will become warm as the
Total Carbohydrate Content Determination of Microalgal Biomass by Acid. . .
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1. Vortex mixer.
2. Adjustable pipet(s), covering 20 μL to 1 mL.
3. Cuvettes—glass, quartz, or plastic.
4. 13 Â 100 mm glass test tubes or equivalent.
5. Glass marbles or caps to cover or plug the 13 Â 100 mm glass
test tubes.
6. Digital dry block, capable of maintaining 80 Æ 3
C (compatible with glass tubes).
7. Spectrophotometer, set to measure at 620 nm.
2.3 Liquid
Chromatography
Apparatus
1. Vortex mixer.
2. Plastic graduated pipet, 5 and 10 mL with 0.1 mL graduations
(see Note 2).
3. Plastic 2 L graduated cylinder or volumetric flask (see Note 2).
4. Screw-top vials and presplit screw caps compatible with the
HPAEC-PAD injector.
5. High-performance
anion-exchange
chromatography
(HPAEC) system equipped with pulsed amperometric detection (PAD) (see Note 3).
2.4 Spectrophotometric Analysis
Reagents and
Standards
Use 18.2 MegaOhm (MΩ) water for all preparations requiring
water. Carefully follow all relevant chemical handling and disposal
procedures.
1. 02% (w/w, specific gravity 1.6338 at 20
C) sulfuric acid, used
for sample two-step hydrolysis.
2. Prepare glucose standard and calibration verification standard
(CVS) stock solutions (these must be made up separately as the
CVS is used to check the accuracy of the calibration curve)
using a high-purity glucose standard. Weigh approximately
25 mg of glucose into a clean, tared, class A, 100 mL volumetric flask. Record the weight to the nearest 0.1 mg. Bring to
volume with water and cap, and mix thoroughly. The standard
and CVS working solution concentrations should be approximately 0.25 mg/mL glucose. Due to balance limits, start with
a higher glucose stock concentration, and dilute to 0.25 mg/
mL. Always prepare the standards and CVS from separate
stocks so that you can check the accuracy of your calibration
with the CVS. Glucose stock solutions can be stored in a freezer
(À20
C) for up to 2 months or 5 days in the refrigerator
(4
C).
3. Prepare a 0.5 M NaOH solution using sodium hydroxide pellets. Weigh out 4 g of sodium hydroxide pellets. Add about
50 mL of water to a 200 mL volumetric flask. Slowly add pellets
and swirl to dissolve. The mixture will become warm as the
Total Carbohydrate Content Determination of Microalgal Biomass by Acid. . .
193