Merck

GC Analysis of Sterols (Silylated Derivatives) in Olive Oil (Free Sterol Fraction) on SLB®-5ms after SPE using Discovery DSC-Si

GC Analysis of Sterols (Silylated Derivatives) in Olive Oil (Free Sterol Fraction) on SLB®-5ms after SPE using Discovery DSC-Si suitable for GC, application for SPE

Materials

analytical column

Product No.
Description
Pricing

SLB®-5ms Capillary GC Column

L × I.D. 20 m × 0.18 mm, df 0.18 μm

CONDITIONS

sample preparation

SPE (Solid Phase Extraction)

sample/matrix

olive oil spiked with a mixed sterol standard at 5 μg/g

SPE tube/cartridge

Discovery DSC-Si, 1 g/6 mL (52656-U)

condition

2 x 5 mL hexane

sample addition

1 mL

elution

esters fraction: 2.5 mL of hexane:ethyl acetate (90:10); free sterols fraction; 5 mL of hexane:ethyl acetate (90:10) and 3 mL x 5 of ethanol:diethyl ether:hexane (50:25:25)

eluate post-treatment

Evaporate to dryness and perform silylation as described in the Analysis Note.

column

SLB-5ms, 20 m x 0.20 mm I.D., 0.20 μm (28564-U)

oven

125 °C (1 min), 10 °C/min to 325 °C (10 min)

inj. temp.

250 °C

detector

MSD, rn/z 500 - 600

MSD interface

325 °C

carrier gas

helium, 0.6 mL/min, constant

injection

1 μL, splitless

liner

4 mm I.D. single taper

sample

Olive oil sample; fractionated, transesterified, and silylated

Description

Analysis Note

Sample pre-treatment
Weigh 0.25 gm oil into test tube. For spiked sample add 12.5 μL of 65-100 μg/mL mixed sterol std. Add 1 ml of hexane:ethyl acetate (90:10). After SPE fractionation, evaporate solvent from each fraction, add 1 ml of 30% methanolic sodium methoxide:MeOH:MTBE solution (13:27:60), mix and let sit at room temp for 30 min. Add 1 ml of water and 2 mL of n-heptane. Withdraw aqueous phase (bottom) and replace w/ 1 mL of 1% citric acid solution. Withdraw organic phase and dry over anhydrous sodium sulfate. Silylation: Evaporate organic phase to dryness. Add 125 μL of Sylon BFT and 125 μL of pyridine. Heat at 70°C for 20 min. The sample is then ready for GC-MS analysis.

Sterol content is one in a battery of tests used to analyze the composition and determine the grade and authenticity of olive oil. Test results are compared against standards such as those established by the International Olive Oil Council (IOC) and US Dept. of Agriculture. In vegetable oils, sterols exist in the free form, or esterified to fatty acids. In this application, sterols were analyzed in olive oil separately as free and esterified. This was accomplished by taking advantage of the polarity difference between these two groups to fractionate them using silica gel SPE. In place of saponification, the fractions were transesterified to liberate the esterified sterols. The fractions were then silylated, and analyzed by GC-MS. The silylated sterols all contained a significant molecular ion in their mass spectum, which made it possible to perform peak identification free of matrix interference. The sterols isolated in the free fraction are shown here.

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