2. Mix 1 mg of isolated bioactive compound with 100 mg potassium bromide and compress the mixture to prepare a small
pellet.
3. Analyze the pellet under FT-IR spectrophotometer in the
range of 349.053–7800.65 cm
À1 at room temperature.
5.3.4 Liquid
Chromatography
Electrospray Ionization
Mass Spectroscopy (LCESI-MS)
1. Analyze the bioactive compound by Agilent 1100 series
LC-MSD with electro spray ionization (ESI) and quadrupole
mass analyzer.
2. Use ammonium hydroxide (0.75 M) as buffering reagent.
3. Perform the analysis at positive ion mode under at a flow rate
1 ml/min, nebulizer pressure 25 psi, capillary voltage 3 kV,
fragmentor voltage 75 V, and drying gas temperature 350
C.
4. Scan a spectrum of bioactive compound over range of
0–650 m/z.
5.3.5 Gas
Chromatography Mass
Spectrometry (GC-MS)
1. Run the GC-MS analysis of the F1 extract on a GC–MS system.
2. The carrier gas is helium (1 ml/min).
3. Use the program: 4 min isothermal at 70
C, then 4
C/min to
180
C and 10 min isothermal.
4. The injection port temperature is 250
C and the detector
temperature is 280
C.
5. Complete ionization of the sample components in the EI mode
(70 eV).
5.3.6 Structural
Elucidation
Nuclear Magnetic
Resonance (NMR)
Spectroscopy
1. NMR spectroscopy is done to examine the structure of isolated
bioactive compound.
2. Dissolve 25 mg of sample in 0.5 ml of DMSO-d 6 .
3. Record
1
H and
13 C NMR spectra on NMR—Jeol/AL400 MHz NMR instrument using deuterated dimethyl
sulfoxide.
4. Use the region from 0 to 12 ppm for
1
H NMR and 0 to
200 ppm for
13 C NMR for scanning. Use trimethylsilane
(TMS) as internal standard.
5. The chemical shifts (δ) are expressed as parts per million (ppm)
and the coupling constants (J) are indicated as hertz (Hz).
6 Biological Assay
The efficacy, virulence, and pathogenicity of symbiotic bacteria,
Xenorhabdus and Photorhabdus, are evaluated under laboratory
conditions.
Biological Assay
139
pellet.
3. Analyze the pellet under FT-IR spectrophotometer in the
range of 349.053–7800.65 cm
À1 at room temperature.
5.3.4 Liquid
Chromatography
Electrospray Ionization
Mass Spectroscopy (LCESI-MS)
1. Analyze the bioactive compound by Agilent 1100 series
LC-MSD with electro spray ionization (ESI) and quadrupole
mass analyzer.
2. Use ammonium hydroxide (0.75 M) as buffering reagent.
3. Perform the analysis at positive ion mode under at a flow rate
1 ml/min, nebulizer pressure 25 psi, capillary voltage 3 kV,
fragmentor voltage 75 V, and drying gas temperature 350
C.
4. Scan a spectrum of bioactive compound over range of
0–650 m/z.
5.3.5 Gas
Chromatography Mass
Spectrometry (GC-MS)
1. Run the GC-MS analysis of the F1 extract on a GC–MS system.
2. The carrier gas is helium (1 ml/min).
3. Use the program: 4 min isothermal at 70
C, then 4
C/min to
180
C and 10 min isothermal.
4. The injection port temperature is 250
C and the detector
temperature is 280
C.
5. Complete ionization of the sample components in the EI mode
(70 eV).
5.3.6 Structural
Elucidation
Nuclear Magnetic
Resonance (NMR)
Spectroscopy
1. NMR spectroscopy is done to examine the structure of isolated
bioactive compound.
2. Dissolve 25 mg of sample in 0.5 ml of DMSO-d 6 .
3. Record
1
H and
13 C NMR spectra on NMR—Jeol/AL400 MHz NMR instrument using deuterated dimethyl
sulfoxide.
4. Use the region from 0 to 12 ppm for
1
H NMR and 0 to
200 ppm for
13 C NMR for scanning. Use trimethylsilane
(TMS) as internal standard.
5. The chemical shifts (δ) are expressed as parts per million (ppm)
and the coupling constants (J) are indicated as hertz (Hz).
6 Biological Assay
The efficacy, virulence, and pathogenicity of symbiotic bacteria,
Xenorhabdus and Photorhabdus, are evaluated under laboratory
conditions.
Biological Assay
139