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For more information, or to download product instructions,
Methoxamine (MOX) Reagent
Use this reagent for preparing oximes of steroids and
ketoacids prior to silylation.
Thermo Scientific MOX Reagent (M.W. 83.51) converts keto
groups to methoxime derivatives. It prevents the formation
of multiple derivatives (which interfere with quantitation)
when enols are present during silylation. Our MOX Reagent
is a 2% solution of methoxyamine•HCI in pyridine, and it is
used primarily with steroids.
The procedures below are used to prepare methoxime
derivatives of steroids and ketoacids prior to silylation.
Forming methoximes is based on the work of Fates and
Luukkainen, with further applications by Horning,
et al
. Both
procedures have been used successfully by Horning,
et al
.
PROTOCOL 1
This simplified procedure is for stable ketones that are readily soluble
in organic solvent.
1. Combine 2 mg sample and 0.5 ml MOX Reagent in a 10 ml Thermo
Scientific Reacti-Vial Small Reaction Vial.
2. Cap vial and heat at 60°C for 3 hours.
3. Add 2 ml water.
4. Extract with three 5 ml portions of high-purity benzene.
5. Combine benzene extracts and wash with 1 N HCI, followed by
bicarbonate solution.
6. Dry over anhydrous magnesium sulfate and evaporate to 0.5 ml with
nitrogen.
7. Analyze by gas or thin layer chromatography.
PROTOCOL 2
This procedure is for polar steroids, such as corticoids, that have
several hydroxyl groups.
1. Combine 2 mg sample and 0.5 ml MOX Reagent in a 10 ml
Thermo Scientific Reacti-Vial Small Reaction Vial.
2. Cap vial and let stand overnight at room temperature.
3. Add 2 ml saturated NaCI solution.
4. Extract with three 5 ml portions of high-purity ethyl acetate.
5. Combine ethyl acetate extracts and wash with salt saturated 0.1 N HCI
wash, followed by a 5% NaHCO
3
salt saturated wash.
6. Dry over anhydrous magnesium sulfate and evaporate with nitrogen
to 0.5 ml.
7. Analyze by gas or thin layer chromatography.
NOTE:
After completing the methoxime reaction, some researchers have
silylated the reacted mixture without further treatment. The resulting
mixture was centrifuged to remove solids, and aliquots of the sample were
used for gas chromatography.
Ordering Information
Product # Description
Pkg. Size
TS-45950
Methoxamine (MOX) Reagent
10 ml
(2% methoxyamine•HCI in pyridine)
Hypo-Vial Sample
Storage Vial
Tri-Sil HTP (HDMS:TMCS:pyridine) Reagent
Our reagent-catalyst solvent mixture for one-step
derivatization.
Thermo Scientific Tri-Sil HTP Reagent is composed of HMDS,
TMCS and high purity pyridine. It is useful for rapid
production of TMS derivatives of polar compounds for gas
chromatographic determination and biochemical synthesis.
The Tri-Sil HTP Reagent is ideal for GC determinations of:
• Sugars
• Alcohols
• Phenols
• Steroids
• Sterols
• Bile acids and other organic acids
• Some amines
Our Tri-Sil HTP Reagent is based on the procedure of
Sweeley,
et al.
and is used for the optimal conversion of
organic hydroxyl and polyhydroxyl compounds into TMS
ethers. The reaction proceeds as:
3 ROH + Me
3
SiNHSiMe
3
Me
3
SiCl – 3 ROSiMe
3
+ NH
4
Cl
The versatility, speed and ease of use of our Tri-Sil HTP
Reagent has made it the most widely used silylation
formulation available.
PROTOCOL
1. Combine 5-10 mg sample and 1.0 ml Tri-Sil HTP Reagent in a 3.0 ml Thermo
Scientific Reacti-ViaI Small Reaction Vial.
2. Shake the reaction vigorously for 30 seconds or warm to 75-85°C to
dissolve.
3. React at room temperature for 5 minutes.
4. Analyze by gas chromatography.
NOTE:
A majority of hydroxyl and polyhydroxyl compounds will be
completely derivatized in less than 5 minutes including sugars, phenols,
organic acids, some amines and alcohols. Highly hindered compounds,
such as some steroids, may require 15 minutes to 8 hours. Extremely
intractable compounds may require refluxing for several hours.
Ordering Information
Product # Description
Pkg. Size
TS-48999
Tri-Sil HTP Reagent
10 x 1 ml
HMDS:TMCS:Pyridine (2:1:10)
ampules
✖
TS-49001
Tri-Sil HTP Reagent
50 ml
HMDS:TMCS:Pyridine (2:1:10)
Hypo-Vial Sample
Storage Vial
✖
Additional hazardous handling charge.
Thermo Scientific Silylation Reagents
