Ultra-High Performance Liquid Chromatography (UHPLC) Method for the Determination of Limonene in Sweet Orange (Citrus sinensis) Oil: Implications for Limonene Stability.

Ultra-High Performance Liquid Chromatography (UHPLC) Method for the Determination of Limonene in Sweet Orange (Citrus sinensis) Oil: Implications for Limonene Stability.

Journal of AOAC International (2015-04-11)

Matthew W Bernart

ABSTRACT

The citrus-derived bioactive monoterpene limonene is an important industrial commodity and fragrance constituent. An RP isocratic elution C18 ultra-HPLC (UHPLC) method using a superficially porous stationary phase and photodiode array (PDA) detector has been developed for determining the limonene content of sweet orange (Citrus sinensis) oil. The method is fast with a cycle time of 1.2 min, linear, precise, accurate, specific, and stability indicating, and it satisfies U.S. Pharmacopeia suitability parameters. The method may be useful in its present form for limonene processing, or modified for research on more polar compounds of the terpenome. A forced-degradation experiment showed that limonene is degraded by heat in hydro-ethanolic solution. PDA detection facilitates classification of minor components of the essential oil, including β-myrcene.

MATERIALS

Product Number

Brand

Product Description

51976

Sigma-Aldrich

Ethanol, BioUltra, for molecular biology, ≥99.8%, (absolute alcohol, without additive, A15 o¹)

PHR1320

Supelco

Residual Solvent - Acetonitrile, Pharmaceutical Secondary Standard; Certified Reference Material

00590590

(R)-(+)-Limonene, primary reference standard

1012772

USP

Dehydrated Alcohol, United States Pharmacopeia (USP) Reference Standard

62118

Supelco

(R)-(+)-Limonene, analytical standard

62122

Sigma-Aldrich

(R)-(+)-Limonene, technical, ~90% (sum of enantiomers, GC)

W263303

Sigma-Aldrich

(R)-(+)-Limonene, ≥93%

183164

Sigma-Aldrich

(R)-(+)-Limonene, 97%

574724

Sigma-Aldrich

Acetonitrile solution, contains 0.05 % (v/v) trifluoroacetic acid

574732

Sigma-Aldrich

Acetonitrile solution, contains 0.1 % (v/v) trifluoroacetic acid, suitable for HPLC

576956

Sigma-Aldrich

Acetonitrile solution, contains 0.1 % (v/v) formic acid, suitable for HPLC

48075

Supelco

Ethanol solution, certified reference material, 2000 μg/mL in methanol

685461

Sigma-Aldrich

Acetonitrile solution, contains 10.0% acetone, 0.05% formic acid, 40.0% 2-propanol

17236

Sigma-Aldrich

3-(Benzyldimethylammonio)propanesulfonate, BioXtra, ≥99.0% (HPCE)

02890

Sigma-Aldrich

Ethanol, purum, fine spirit, denaturated with 2% 2-butanone, F25 MEK¹, ~96% (based on denaturant-free substance)

792810

Sigma-Aldrich

Ethyl alcohol, Pure, 140 proof, Excise Tax-free, Permit for use required

792829

Sigma-Aldrich

Ethyl alcohol, Pure, 140 proof, meets water USP testing specifications, Excise Tax-free, Permit for use required

792799

Sigma-Aldrich

Ethyl alcohol, Pure, 190 proof, ACS reagent, meets USP testing specifications, Excise Tax-free, Permit for use required

V800024

Sigma-Aldrich

Acetonitrile, AR, ≥99.5%

L010300

Sigma-Aldrich

Ultrapure Acetonitrile

110086

Sigma-Aldrich

Acetonitrile, ReagentPlus^®, 99%

82063

Sigma-Aldrich

Acetonitrile, for preparative HPLC, ≥99.8% (GC)

E7023

Sigma-Aldrich

Ethyl alcohol, Pure, 200 proof, for molecular biology

E7148

Sigma-Aldrich

Ethyl alcohol, Pure, 190 proof, for molecular biology

60004

Sigma-Aldrich

Acetonitrile, ≥99.5% (GC)

34442

Sigma-Aldrich

Acetonitrile, suitable for DNA synthesis, ≥99.9% (GC)

PHR1551

Supelco

Acetonitrile, Pharmaceutical Secondary Standard; Certified Reference Material

493538

Sigma-Aldrich

Ethyl alcohol, Pure, 190 proof, meets USP testing specifications

271004

Sigma-Aldrich

Acetonitrile, anhydrous, 99.8%

437557

Sigma-Aldrich

Acetonitrile, ≥99.5%, ACS reagent