2.15 Determination
of Number and
Configuration of
Double Bonds in MAME
by Argentation
Chromatography
1. Silica gel G60 precoated plates.
2. TLC tank.
3. 10% (wt/vol) aqueous AgNO 3 .
4. HPLC-grade dichloromethane (see Note 2).
5. 0.2% dichlorofluorescein in 50% ethanol in distilled water.
6. Oven at 100
C.
2.16 Location of
Double Bonds in MAME
by PeriodatePermanganate
Oxidation
1. NaIO 4 .
2. KMnO 4 .
3. NaIO 4 (214 mg) and KMnO 4 (39.5 mg) in 100 ml water
(stock oxidant solution).
4. Aqueous 0.002 M Na 2 CO 3 .
5. HPLC-grade solvents: Toluene, ter-butyl alcohol, and diethyl
ether (see Note 2).
6. Sodium metabisulfite.
7. 20% H 2 SO 4 in water.
8. Air or nitrogen stream.
9. Shaker for tubes at 37
C.
10. Screw cap glass tubes.
11. TLC as described in Subheading 2.4 with dichloromethane as
running solvent.
12. GC/MS as in Subheading 2.5.
2.17 Characterization of Hydroxyl
Functions by Mass
Spectrometry After
Trimethylsilylation
1. Glass screw-capped tubes.
2. Pasteur pipettes.
3. Anhydrous pyridine.
4. Hexamethyldisilazane.
5. Trimethylchlorosilane.
6. HPLC-grade petroleum ether.
7. Air or nitrogen stream.
2.18 Polarimetry
1. Apparatus: Automatic digital readout polarimeter at 20
C.
2. Solvents HPLC grade: Chloroform and diethyl ether (see Note
2).
3. Pasteur pipettes.
2.19 MALDI-TOF
Mass Spectrometry
of MAME
1. Apparatus 5800 MALDI TOF/TOF-MS Analyzer (Applied
Biosystems/AB Sciex) equipped with an Nd:Yag laser
(349-nm wavelength).
2. Matrix solution: 2,5-dihydroxybenzoic acid 10 mg/mL in
chloroform/methanol, 1/1 (v/v) (see Note 2).
3. 2-μL automatic pipette.
Lipid and Lipoarabinomannan
121
of Number and
Configuration of
Double Bonds in MAME
by Argentation
Chromatography
1. Silica gel G60 precoated plates.
2. TLC tank.
3. 10% (wt/vol) aqueous AgNO 3 .
4. HPLC-grade dichloromethane (see Note 2).
5. 0.2% dichlorofluorescein in 50% ethanol in distilled water.
6. Oven at 100
C.
2.16 Location of
Double Bonds in MAME
by PeriodatePermanganate
Oxidation
1. NaIO 4 .
2. KMnO 4 .
3. NaIO 4 (214 mg) and KMnO 4 (39.5 mg) in 100 ml water
(stock oxidant solution).
4. Aqueous 0.002 M Na 2 CO 3 .
5. HPLC-grade solvents: Toluene, ter-butyl alcohol, and diethyl
ether (see Note 2).
6. Sodium metabisulfite.
7. 20% H 2 SO 4 in water.
8. Air or nitrogen stream.
9. Shaker for tubes at 37
C.
10. Screw cap glass tubes.
11. TLC as described in Subheading 2.4 with dichloromethane as
running solvent.
12. GC/MS as in Subheading 2.5.
2.17 Characterization of Hydroxyl
Functions by Mass
Spectrometry After
Trimethylsilylation
1. Glass screw-capped tubes.
2. Pasteur pipettes.
3. Anhydrous pyridine.
4. Hexamethyldisilazane.
5. Trimethylchlorosilane.
6. HPLC-grade petroleum ether.
7. Air or nitrogen stream.
2.18 Polarimetry
1. Apparatus: Automatic digital readout polarimeter at 20
C.
2. Solvents HPLC grade: Chloroform and diethyl ether (see Note
2).
3. Pasteur pipettes.
2.19 MALDI-TOF
Mass Spectrometry
of MAME
1. Apparatus 5800 MALDI TOF/TOF-MS Analyzer (Applied
Biosystems/AB Sciex) equipped with an Nd:Yag laser
(349-nm wavelength).
2. Matrix solution: 2,5-dihydroxybenzoic acid 10 mg/mL in
chloroform/methanol, 1/1 (v/v) (see Note 2).
3. 2-μL automatic pipette.
Lipid and Lipoarabinomannan
121