ISOLATION AND PURIFICATION OF TRIMYRISTIN FROM
NUTMEG WITH TLC ANALYSIS (5/20/09)
(7/20/04)
The isolation and characterization of organic compounds from living organisms is most
often referred to as natural products chemistry. In fact, in its early days, organic
chemistry was almost exclusively natural products chemistry. Even today, the isolation,
identification, and synthesis of natural products remain important areas of research for
organic chemists. This interest in natural products derives from their usefulness as
medicines, dyes, insecticides, and flavorings.
Historically, most natural products have been isolated from plants. Natural product
isolations are normally quite tedious and complex because even the simplest plants
contain a multitude of organic compounds, often in very small quantities. However, in
some instances, a natural product occurs in relatively high concentrations. In these cases
the isolation of the natural product is fairly easy, involving a simple extraction of a part
of the plant with a suitable organic solvent. The isolation of trimyristin (1) from nutmeg
is one of these easy isolations. Nutmegs are hard, aromatic-smelling seeds of an East
Indian tree (Myrictica fragans). Trimyristin is a triester of glycerol (2) and myristic acid
(3), a fatty acid. Glycerol triesters are also called triglycerides. Naturally occurring fats
and oils are generally triglycerides that contain a mixture of long chain (fourteen to
twenty carbon atoms) carboxylic acids (fatty acids). Fats are solid triglycerides which
contain mostly saturated fatty acids, whereas oils are liquid triglycerides which contain
mostly unsaturated fatty acids. Trimyristin is unusual as a naturally occurring
triglyceride in that it contains exclusively myristic acid.
O
CH2O
C
(CH 2) 12CH3
CH2OH
O
O
CHO
C
(CH 2) 12CH3
CHOH
(CH 2) 12CH3
CH2OH
HO
C
(CH 2) 12CH3
O
CH2O
C
1
2
3
Trimyristin
Glycerol
Myristic acid
In this experiment, a whole nutmeg will be crushed and extracted with diethyl ether to
obtain crude trimyristin. The crude trimyristin will be purified by crystallization, and its
purity will be checked its melting point and by thin layer chromatography. When
trimyristin is hydrolyzed with a base, a process known as saponification, glycerol and
myristic acid are produced. Myristic acid has a melting point very close to that of
trimyristin so that a melting point alone is not sufficient for distinguishing trimyristin
from myristic acid. However trimyristin and myristic acid may be readily distinguished
by thin layer chromatography.
Experimental Procedure
First Lab
Crush and grind a whole nutmeg with a mortar and pestle for approximately 5 min. (no
need to pulverize). Weigh the ground nutmeg and transfer it to a 50 mL round-bottomed
flask. Add 15 mL of diethyl ether, attach a water-cooled condenser to the flask, and heat
the mixture under a gentle reflux for approximately 15 min. using a thermowell. Allow
the mixture to cool to room temperature. Without disturbing the settled solids, decant and
gravity-filter the ether solution into a 125 mL Erlenmeyer flask. Add another 10 mL of
diethyl ether to the solid residue, swirl for a few minutes, and transfer the contents to the
filter. Prepare a TLC plate and apply a small amount of the filtrate (crude product).
(Wait until the following week to develop in a chamber.) Finally, quantitatively transfer
the filtrate to a round bottom flask and remove the solvent using a rotary evaporator.
Second Lab
Add sufficient reagent grade acetone to dissolve the remaining crude solid product
(approximately 3-5 mL of acetone, the amount will vary depending upon the weight of
nutmeg extracted). Allow to cool to room temperature. Collect the crystalline trimyristin
by vacuum filtration and allow it to air-dry. Weigh the product and calculate the percent
of trimyristin in the nutmeg.
Place a small amount (the tip of a spatula) of the purified trimyristin in a test tube, then
add a drop or two of chloroform. Do likewise with the crude product. Next, obtain a
small amount of prepared myristic acid (saponified trimyristin) and dissolve in
chloroform (same amounts/procedure as used for trimyristin & crude product). Spot the
TLC plate with each of the chloroform solutions and develop the plate using chloroform
as the solvent. Place the plate under UV light and observe. Next, develop the plate in an
iodine chamber. Note that myristic acid adsorbs less iodine than the surrounding silica
gel and appears as a white spot on a yellow-brown background.
Place some dry trimyristin, just enough to see, in a melting point capillary. Do likewise
with a small amount of prepared myristic acid and determine the melting point of the two
compounds. The melting point should be near 56oC. Remember that melting point
determinations are most accurate when the temperature rise near the melting point
(within 10qC) is approximately 1-2oC/min.
Prepare the TLC plate in a manner consistant with those prepared in the lab’s
introductory experiment, “TLC of Analgesics”.
Do not discard TLC plates. Attach to the postlab exercise form.
¥ Perform purification, TLC and melting point analysis during 2nd week of this
experiment.
Always use a flask (not a beaker) to contain volatile reagents. A flask must always be
used for crystallization/recrystallization procedures!
WASTE DISPOSAL NOTES: Nutmeg, Trimyristin, and Myristic Acid should all be disposed of in
the Non-hazardous Solid Waste Container (located in the Waste Hood)
REFLUX APPARATUS
NOTE: Use a hose clamp to anchor hoses to the condenser inlet and outlets!
Name:__________________________________ Section:_______ Date:_______
POSTLAB EXERCISE FOR:
Extraction of Trimyristin from Nutmeg
with TLC Analysis (7/20/2004)
>> Due no later than the 2nd Friday following the scheduled experiment; no exceptions will
be made! Please answer questions on this form. (30pts TOTAL) <<
A. PRODUCT INFORMATION (15pts)
TRIMYRISTIN (10pts)
%Recovery (4pts):
(4pts)
Melting Point
(4pts):
(2pts)
Appearance
(describe) (4pts):
(4pts)
MYRISTIC ACID (2pts)
X (0pts – n/a)
(2pts)
X (0pts – n/a)
(3pts) Indicate the appearance of your TLC plate (below) and attach originals to this
form. Be sure to identify as many of the components as possible!
B. SUMMARY QUESTIONS (15pts)
1) (4pts) Is melting point determination or thin-layer chromatography analysis a
better method to determine the purity of trimyristin? Explain.
2) (4pts) Was the melting point or thin-layer chromatography a better method for
distinguishing between the compounds trimyristin and myristic acid? Explain.
3) (4pts) Why was the whole nutmeg crushed and ground before it was extracted
with diethyl ether?
4) (3pts) Would water be a suitable solvent for extracting trimyristin from nutmeg?