A GUIDE TO CAROTENOID ANALYSIS IN FOODS

34 A Guide to Carotenoid Analysis in Foods
DO’S AND DON’TS IN CAROTENOID ANALYSIS
For emphasis, the necessary measures that should
be taken to ensure the reliability of carotenoid data
are summarized below.
Do’s
• Before starting any analytic work, familiarize yourself
with the nature of food carotenoids and the
physicochemical properties of these compounds.
With this background information, you will save
time and money and prevent analytic errors.
• If you have the opportunity, undergo training for at
least 15 days in an experienced carotenoid laboratory.
This will put you ahead in your work much
faster.
• If you are going to use a high-performance liquid
chromatographic (HPLC) method, be sure that you
know how to use this chromatographic technique
very well. It is very easy to make errors with this
technique, and because the results are reproducible
(high precision), erroneous data are not easily
perceived.
• Search the literature for previous carotenoid work
with the food samples you intend to analyze. This
will give you an idea of what to expect, but remember
that natural variations exist among samples
of the same food (as a function of factors such as
stage of maturity, variety, climate or season, and
portion taken as edible) and analytic errors persist
in the literature.
• Work under subdued artificial light from the extraction
step on. If the laboratory is used by other
analysts for other types of analyses (as is usually
the case in developing countries) and it is not possible
to put the whole laboratory under dim light,
choose the part of the laboratory most protected
from sunlight, turn off the lights in this area, and
shield carotenoids from diffuse light by covering
vessels and equipment (e.g., open columns and thinlayer
chromatography development tanks) with
aluminum foil or black cloth. It may also be neces-
sary to put blinds on windows, use tinted windows,
or cover some windows with aluminum foil or other
appropriate material.
• Test your selected analytic method exhaustively
before you use it to gather data. The time, resources,
and effort you spend in testing will be more
than compensated by avoidance of erroneous or
meaningless results.
• Use reagent-grade solvents and reagents for the
prechromatographic steps and for open-column
chromatography. If only technical-grade solvents
are available, distill them before use. Use HPLCgrade
solvents for HPLC. Purification of solvents
for HPLC in the laboratory may be time consuming
and, in the end, may be more costly.
• Test ethyl ether and tetrahydrofuran for peroxides;
if present remove peroxides, for example, by distillation
over reduced iron powder. Stabilize tetrahydrofuran
with an antioxidant (e.g., butylated hydroxytoluene
BHT); if stabilized tetrahydrofuran
is bought, observe the time limit for its use.
• Carry out all operations as rapidly as possible, especially
the steps that introduce risks of oxidation
and isomerization. Take advantage of the color of
carotenoids and monitor carotenoid losses in the
different steps by carefully observing color changes.
• Exclude oxygen as much as possible. Store carotenoids
in air-tight containers under vacuum or nitrogen.
• Obtain and prepare samples for analysis according
to an adequate plan that meets the objectives
of your work and submits homogeneous and representative
samples for analysis. Unless sampling
and sample preparation are done properly, it is useless
to spend time and effort in carrying out the
analysis per se with great accuracy.
• Plan your work so that samples are analyzed immediately
after collection. Changes are difficult to
prevent during storage of samples, even at low temperature.
If you have to store samples, store them