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Printed by: USP NF Official Date: Official as of 01-May-2020 Document Type: NF & DS @2024 USPC
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1
Table 2 (continued)
Eucalyptus Oil
Relative
Retention
CAS RN®: 8000-48-4. Name Time
DEFINITION β-Pinene 0.80
Eucalyptus Oil is obtained by steam distillation and
rectification from the fresh leaves or the fresh terminal Sabinene 0.83
branchlets of various species of Eucalyptus rich in 1,8-cineole. (R)-(–)-α-Phellandrene 0.91
The species mainly used are Eucalyptus globulus Labill.,
Eucalyptus polybractea R.T. Baker, and Eucalyptus smithii R.T. (R)-(+)-Limonene 0.98
Baker. It contains NLT 70.0% and NMT 95.0% of eucalyptol Eucalyptol 1.00
(1,8-cineole, C10H18O).
Camphor 1.58
IDENTIFICATION
Change to read: Suitability requirements
Resolution: NLT 1.5 between (R)-(+)-limonene and
• A. ▲SPECTROSCOPIC IDENTIFICATION TESTS á197ñ, Infrared eucalyptol
Spectroscopy: 197F ▲ (CN 1-May-2020) Analysis
• B. IDENTITY BY AROMA SUBSTANCE PROFILE: The retention Samples: Standard solution A, Standard solution B, and
times of the (+)-α-pinene, β-pinene, (R)-(–)-α-phellandrene, Sample solution
(R)-(+)-limonene, and eucalyptol peaks of the Sample Identify the peaks in the Sample solution based on those in
solution correspond to those of the Standard solution, as Standard solution A as well as in Table 3.
al
obtained in the test for Content of Aroma Substances in the
Assay. Table 3
Relative
ASSAY Retention
• CONTENT OF AROMA SUBSTANCES Name Time
Standard solution A: 1.0 µL/mL of (+)-α-pinene, 0.5 µL/mL
of β-pinene, 0.5 µL/mL of sabinene, 0.5 µL/mL of
(R)-(–)-α-phellandrene, 1.0 µL/mL of (R)-(+)-limonene, 5
µL/mL of USP Eucalyptol RS, and 5 mg/mL of USP Camphor
ci β-Myrcene
γ-Terpinene
p-Cymene
0.86–0.90
1.05–1.07
1.12–1.13
RS in heptane
Standard solution B: 0.01 µL/mL of (R)-(+)-limonene in Terpinen-4-ol 1.68
ffi
heptane α-Terpineol 1.82
Sample solution: 20 µL/mL of Eucalyptus Oil in heptane
Chromatographic system
(See Chromatography á621ñ, System Suitability.) β-Myrcene elutes before (R)-(–)-α-phellandrene and after
Mode: GC sabinene; γ-terpinene and p-cymene elute between
Detector: Flame ionization eucalyptol and camphor; and terpinen-4-ol and
α-terpineol elute after camphor.
O
Column: 0.25-mm × 60-m fused-silica capillary; 0.25-µm
layer of phase G16 Calculate the percentage of (+)-α-pinene [β-pinene,
Temperatures sabinene, (R)-(–)-α-phellandrene, (R)-(+)-limonene, or
Injection port: 220° eucalyptol] in the portion of Eucalyptus Oil taken:
Detector: 220°
Column: See Table 1. Result = (rU/rS) × (CS/CU) × 100
Table 1
rU = peak response of (+)-α-pinene [β-pinene,
sabinene, (R)-(–)-α-phellandrene,
Hold Time (R)-(+)-limonene, or eucalyptol] from the Sample
Initial Temperature Final at Final
Temperature Ramp Temperature Temperature solution
(°) (°/min) (°) (min) rS = peak response of (+)-α-pinene [β-pinene,
sabinene, (R)-(–)-α-phellandrene,
60 — 60 5
(R)-(+)-limonene, or eucalyptol] from the
60 5 200 5 Standard solution
CS = concentration of (+)-α-pinene [β-pinene,
Carrier gas: Helium sabinene, (R)-(–)-α-phellandrene,
Flow rate: 1.5 mL/min (R)-(+)-limonene, or USP Eucalyptol RS] in the
Injection type: Split ratio, 1:50 Standard solution (µL/mL)
Injection volume: 1 µL CU = concentration of Eucalyptus Oil in the Sample
System suitability solution (µL/mL)
Sample: Standard solution A
[NOTE—The relative retention times are listed in Table Calculate the percentage of camphor in the portion of
2.] Eucalyptus Oil taken:
Result = (rU/rS) × [CS/(CU × D)] × 100
Table 2
Relative rU = peak response of camphor from the Sample
Retention solution
Name Time
rS = peak response of camphor from the Standard
(+)-α-Pinene 0.64 solution
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Printed on: Fri Jan 05 2024, 09:30:38 PM(EST) Status: Currently Official on 06-Jan-2024 DocId: GUID-FEA84FA5-7C15-464F-85D5-F8D18C04F05D_2_en-US
Printed by: USP NF Official Date: Official as of 01-May-2020 Document Type: NF & DS @2024 USPC
Do Not Distribute DOI Ref: 5pb4g DOI: https://doi.org/10.31003/USPNF_M387_02_01
2
CS = concentration of USP Camphor RS in the Standard camphor, β-myrcene, γ-terpinene, p-cymene,
solution (mg/mL) terpinen-4-ol, and α-terpineol]: NLT 98.0%
CU = concentration of Eucalyptus Oil in the Sample
solution (µL/mL) IMPURITIES
D = density of Eucalyptus Oil (mg/µL) • TEST FOR ALDEHYDE
Alcoholic hydroxylamine solution: Dissolve 3.5 g of
Calculate the percentage of β-myrcene (γ-terpinene, hydroxylamine hydrochloride in 95 mL of 60% alcohol
p-cymene, terpinen-4-ol, or α-terpineol) in the portion of (v/v) and add 0.5 mL of a 2-mg/mL solution of methyl
Eucalyptus Oil taken: orange in 60% alcohol (v/v) and sufficient 0.5 M potassium
hydroxide in 60% alcohol (v/v) to give a pure yellow color.
Result = (rU/rT) × 100 Dilute with 60% alcohol (v/v) to 100 mL.
Analysis: In a ground-glass-stoppered tube, 25 mm in
rU = peak response of β-myrcene (γ-terpinene, diameter and 150 mm long, add 10 mL of Eucalyptus Oil.
p-cymene, terpinen-4-ol, or α-terpineol) from the Then add 5 mL of toluene and 4 mL of Alcoholic
Sample solution hydroxylamine solution. Shake vigorously, and titrate
rT = sum of the total peak responses, except for the immediately with 0.5 M potassium hydroxide in 60%
peaks due to solvent, from the Sample solution alcohol (v/v) until the red color changes to yellow.
Continue the titration with shaking; the endpoint is reached
Acceptance criteria: See Table 4. Disregard any peak with when the pure yellow color of the indicator is permanent
an area less than the major peak area from Standard solution in the lower layer after shaking vigorously for 2 min and
B, corresponding to 0.05%. allowing separation to take place. The reaction is complete
in about 15 min.
Table 4 Repeat the titration using a further 10 mL of Eucalyptus Oil
al
Acceptance
and, as a reference solution for the endpoint, the titrated
Criteria, liquid from the first determination, to which has been
Name NMT (%) added 0.5 mL of 0.5 M potassium hydroxide in 60%
(+)-α-Pinene 0.2–10.0
alcohol (v/v).
ci Acceptance criteria: NMT 2.0 mL of 0.5 M potassium
β-Pinene 0.05–1.5 hydroxide in 60% alcohol (v/v) is required in the second
Sabinene 0.3 titration.
(R)-(–)-α-Phellandrene 0.05–1.5 SPECIFIC TESTS
• SPECIFIC GRAVITY á841ñ: 0.906–0.927, at 20°
(R)-(+)-Limonene 2.0–15.0 • REFRACTIVE INDEX á831ñ: 1.450–1.470, at 20°
ffi
Eucalyptol 70.0–95.0 • OPTICAL ROTATION á781ñ: 0°–10°, at 20°
Camphor 0.1 ADDITIONAL REQUIREMENTS
• PACKAGING AND STORAGE: Preserve in tight containers. Do
Percentage of γ-terpinene: 0.1%–6.0% not store above 25°.
Percentage of p-cymene: 0.5%–15.0% • USP REFERENCE STANDARDS á11ñ
O
Total percentage of all identified aroma substances USP Camphor RS
[(+)-α-pinene, β-pinene, sabinene, USP Eucalyptol RS
(R)-(–)-α-phellandrene, (R)-(+)-limonene, eucalyptol, USP Eucalyptus Oil RS
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