ISO 23349:2020 pdf free.Animal and vegetable fats and oils一Determination of sterols and stanols
in foods and dietary supplements containing added phytosterols.
5.6 Sodium hydroxide solution, 2,3 N.
Weigh 92 g of sodium hydroxide (5.5) into a 1 000 ml volumetric flask (6.14). Add a stir bar (6.20) and 750 ml of methanol (SA). Place the flask on a magnetic stirrer (6.21) to disperse. Dilute to the mark with methanol, and invert to mix. The sodium hydroxide solution shall be stirred each time before use because it will settle out with time.
5.7 Hydrochloric acid, 3,0 N.
5.8 Epicoprostanol (5f3-cholestan-3a-ol), of purity> 95 %.
5.9 Epicoprostanol internal standard (IS) solution, 5 mg/mi.
Accurately weigh 5,0 g of epicoprostanol (5) into a glass weighing scoop Iranster the epicoprostanol to a 1 000 ml volumetric flask (6.14). Rinse the scoop with internal standard dissolving solvent (5..3) into the flask. Dilute to the mark with toluene or ethyl acetate and invert to mix. The epicoprostanol IS solution of concentration 5 mg/mi is used for sterol/stanol concentrates, steryl/ stanoi ester concentrates and some dietary supplements.
5.10 Epicoprostanol internal standard (IS) solution, 2 mg/mi.
Accurately weigh 2,0 g of epicoprostanol (5) into a glass weighing scoop (). Transfer the epicoprostanol to a 1 000 ml volumetric flask (6.14). Rinse the scoop with internal standard dissolving solvent (5.3) into the flask. Dilute to the mark with toluene or ethyl acetate and invert to mix. The epicoprostanol IS solution of concentration 2 mg/ml is used for analysis of foods and some dietary supplements.
For new formulations (phytosterol type and matrix), determine the optimal protocol by triplicate analysis according to the following protocols: a) 120 mm alkaline, b) 15 mm alkaline and c) 45 mm acid/15 mm alkaline. Select the protocol that produces the highest phytosterol recovery and lowest variability. Further refine this protocol, as needed, by decreasing the saponification time from 120 mm to 60 mm to 30 mm, or the acid hydrolysis time from 45 mm to 30 mm to 15 mm, with triplicate analyses for each. The optimal protocol will show a repeatability coefficient of variation (Cv,.) of less than 5 % for triplicate analyses over a period of five days of testing.
The 120 mm alkaline protocol is appropriate for the analysis of most foods and dietary supplements containing added phytosterols. The 45 mm acid/15 mm alkaline protocol can be required for complete liberation of phytosterol esters from certain matrices, such as some cereals.
Use this protocol for some foods and dietary supplements containing added phytosterol esters. Acid hydrolysis prior to saponification is needed for the complete liberation of phytosterol esters from certain matrices, such as some cereals (see 91=).
a) Accurately weight the test portion into a tarred round bottom flask (6.7.1). Record the exact weight.
b) Add a few boiling chips (6.19) and place a PTFE sleeve (6.7.2) in the neck of the flask.
c) Add 5 ml of hydrochloric acid (51).
d) Add 5,00 ml of the appropriate epicoprostanol IS solution (5,9 or 5.10).
e) Boil the sample at 100 °C for 45 mm.
f) Remove the flask from the heat source, insert a stopper (6.7.3) and let cool to room temperature.
g) Add 40 ml of sodium chloride solution (5.14), insert a stopper (6.7.3) and shake to mix. Allow the two phases to separate.
h) Transfer the organic phase (as much as possible without disrupting the solids on the surface) to a new round bottom flask (6.7.1).
i) Add 5 ml of sodium hydroxide solution (5k).
j) Boil the sample at 100 °C for 15 mm.
k) Remove the flask from the heat source, insert a stopper (6.7.3) and let cool to room temperature.
1) Add 7 ml of hydrochloric acid (51) insert a stopper (6.7.3) and shake to mix.ISO 23349 pdf download.