Peppermint oil

Botanical orign
Mentha piperita

Chemical constitutents
 Menthol
 Menthone
 1,8-Cineole
 b-Caryophyllene
 Limonene

Iodine value
 The reagent is prepared by mixing 8 g. of pyridine and 10 g of concentrated sulfuric acid
with 20 ml. of glacial acetic acid, and to this solution adding 12 g. of bromine in 20 ml.
of glacial acetic acid.
 After thorough mixing, the solution is diluted to one liter with glacial acetic acid
 To determine the iodine number for a sample, 0.5 ml. of oil, accurately weighed, is
treated with 25 ml. of the reagent and allowed to stand for 30 minutes.
 Two grams of potassium iodide is then added and the mixture is diluted with 50 ml. of
water. The mixture is allowed to stand for 10 minutes and the iodine liberated by the
excess bromine is titrated with N/10 sodium thiosulfate solution.
 With each group of samples a blank is run under similar conditions. The iodine value is
calculated using the following formula:

Iodine Value = (B — A) (0.01269)100

where

 B is the number of milliliters of N/10 sodium thiosulfate solution required for the blank,
 A is the number of milliliters of N/10 sodium thiosulfate solution for back titration,
 0.01269 is the iodine factor for N/10 iodine solution,
 W is the weight of sample. No marked variation of iodine values with length of exposure
of oils to the bromine solution was observed

Acid value
Maximum 1.4, determined on 5.0 g diluted in 50 mL of the prescribed mixture of solvents
Mint oil
A. Thin-layer chromatography
Test solution
Mix 0.1 g of the substance to be examined with toluene R and dilute to 10 mL with the same
solvent.
Reference solution
Dissolve 50 mg of mentholR, 20 uL of cineole R, 10 mg of thymolRand 10 ul, of menthyl
acetate R in toluene R and dilute to 10 mL with the same solvent.
Plate TLC silica gelF254 plateR (5-40 um) [or TLC silica gel
F254 plate R (2-10 umj].
Mobile phase Ethyl acetate R, toluene
Application 10 ul, [or 1 )lL] of the reference solution and 20 uL [or 2ul] of the test solution, as
bands of 10 mm [or 8mm].
Development Over a path of 15 cm [or 6 cm].
Drying In air.
Detection A Examine in ultraviolet light at 254 nm.
Detection B Treat with anisaldehyde solution R and heat at 100-105 DC for 5-10 min; examine
immediately in daylight.
Results B The chromatogram obtained with the test solution shows no blue zone between the
zones due to 1,8-cineole and menthol.
B
Examine the chromatograms obtained in the test for chromatographic profile. .
Results the chromatogram obtained with the test solution does not show a peak with the retention
time of isopulegol that has an area of more than 0.2 per cent of the total area.
Test solution
Mix 0.20 mL of the substance to be examined with heptane R and dilute to 10.0 mL with the
same solvent.
Reference solution (a)
Dissolve 10 uL of limonene R, 20 uL of cineole R, 40 uL of menthane R, 10 uL of menthofuran
R, 10ul of (+)-isomenthone R, 40 ul, of menthyl acetate R, 20 ulL of isopulegol R, 60 mg of
mentholR, 20ul of pulegone R, 10 uL of piperitone R and 10 ul, of carvone R in heptane R and
dilute to 10.0 mL with the same solvent.
Reference solution (b)
Dissolve 5 uL of isopulegol R in heptane R and dilute to 10.0 mL with the same solvent.
Dilute 0.1 mL of the solution to 5.0 mL with heptane R.
Column:
Material: fused silica;
Size: 1= 60 m, 0 = 0.25 mm;
Stationary phase: macrogol 20 000 r (film thickness 0.25 µm).
Carrier gas helium for chromatography R.
Flow rate 1.5 ml/min.
Split ratio 1:50.
Temperature:

Detection flame ionization.

Injection 1 uL.
Elution order indicated in the composition of reference solution (a); record the retention times
of these substances.
Identification of peaks Using the retention times determined from the chromatogram obtained
with reference solution (a), locate the components of reference solution (a) in the chromatogram
obtained with the test solution.
System suitability Reference solution (a)
Resolution: minimum.1.5 between the peaks due to limonene and 1,8-cineole; minimum 1.5
between the peaks due to piperitone and carvone.
Determine the percentage content of each of the following components.
The limits are within the following ranges:
 Limonene: 1.0 per cent to 3.5 per cent;
 1,8-cineole: 3.5 per cent to 8.0 per cent;
 Menthone: 14.0 per cent to 32.0 per cent;
 Menthofuran: 1.0 per cent to 8.0 per cent;
 Isomenthone: 1.5 per cent to 10.0 per cent;
 Menthyl acetate: 2.8 per cent to 10.0 per cent;
 Isopulegol: maximum 0.2 per cent;
 Menthol: 30.0 per cent to 55.0 per cent;
 Pulegone: maximum 3.0 per cent,
 Carvone: maximum 1.0 per cent;
 Disregard limit: the area of the principal peak in the chromatogram obtained with
reference solution (b) (0.05 per cent).
 The ratio of 1,8-cineole content to limonene content is minimum 2
ASSAY
Protocol Bp and USP
Total esters
Sample : 109 of Oil
Analysis: Placethe Sample in a 250-mL conical flask
 Add 10 mL of neutralized alcohol and 2 drops of phenolphthalein TS,
 Then add, dropwise, 0.1 N sodium hydroxide until a faint pink color appears.
 Add 25.0 mL of 0.5 N alcoholic potassium hydroxide VS, connect the flask to a reflux
condenser and heat on a boiling water bath for 1 h.
 Allow the mixture to cool, add 20 mL of water, and add phenolphthalein TS.
 Titrate the excess alkali with 0.5 N hydrochloric acid Vs.
 Perform a blank determination, disregarding the 0.1 N sodium hydroxide
  Each mL of 0.5 N alcoholic potassium hydroxide consumed in the saponification is
equivalent to 99.15 mg of total esters calculated as C12H22O2
 Acceptance criteria: NLT 5.0% of esters, calculated as C12H22O2
Total menthol
Sample: 10 mL of Oil
Analysis: Place the Sample in an acetylation flask of 100mL capacity.
 Add 10 mL of acetic anhydride and 1 g of anhydrous sodium acetate.
 Boil the mixture gently for 1 h, accurately timed, cool, disconnect the flask from the
condenser, Transfer the mixture to a small separator,
 Rinsing the acetylation flask with three 5-mL portions of warm water.
 Add the rinsings to the separator.
 When the liquids have completely separated, discard the water layer, and wash the
remaining oil with successive portions of sodium carbonate TS, diluted with an equal
volume of water, until the last washing is alkaline to phenolphthalein TS.
 Dry the resulting 011 with anhydrous sodium sulfate, and filter.
 Transfer 5 mL of the dry acetylated oil to atared, 100-mL conical flask, and weigh.
 Add 50.0 mL of 0.5 N alcoholic potassium hydroxide VS, connect the flask to a reflux
condenser, and boil the mixture on a steam bath for 1 h, accurately timed.
 Allow the mixture to cool, and add 10 drops of phenolphthalein TS.
 Titrate the excess alkali with 0.5 N hydrochloric acid VS.
 Perform a blank determination Residual Titrations).
Calculate the percentage of total menthol in the Oil tested:
Result= 7.813 × A × (1 - 0.0021 × E)/(W- 0.021 × A)
A =result obtained by subtracting the number of mL of 0.5 N hydrochloric acid required in the
above titration from the number of mL of 0.5 N hydrochloric acid required in the residual
titration blank.
E = percentage of esters calculated as menthyl acetate C12H22O2
W =weight of acetylated Oil taken (g)
Acceptance criteria: NLT 50.0% of C10H200, free and as Esters
References
https://en.wikipedia.org/wiki/Peppermint

Lawrence H. Baldinger, University of Notre Dame, Iodine Number of Oil of Peppermint, 108-
109

United state pharmacopoeia 5866

British pharmacopoeia 376-377