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

00590590

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

48075

Supelco

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

E2385

Supelco

Ethanol standards 10% (v/v), 10 % (v/v) in H₂O, analytical standard

2858

Sigma-Aldrich

Ethanol Fixative 80% v/v, suitable for fixing solution (blood films)

62122

Sigma-Aldrich

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

62118

Supelco

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

17236

Sigma-Aldrich

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

183164

Sigma-Aldrich

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

W263303

Sigma-Aldrich

(R)-(+)-Limonene, ≥93%

1012772

USP

Dehydrated Alcohol, United States Pharmacopeia (USP) Reference Standard

PHR1070

Supelco

Dehydrated Alcohol, Pharmaceutical Secondary Standard; Certified Reference Material

PHR1320

Supelco

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

459844

Sigma-Aldrich

Ethyl alcohol, Pure, 200 proof, ACS reagent, ≥99.5%

E7148

Sigma-Aldrich

Ethyl alcohol, Pure, 190 proof, for molecular biology

494445

Sigma-Aldrich

Acetonitrile, biotech. grade, ≥99.93%

34881

Sigma-Aldrich

Acetonitrile, suitable for HPLC-GC, ≥99.8% (GC)

02857

Sigma-Aldrich

Ethanol, purum, fine spirit, denaturated with 4.8% methanol, F25 METHYL¹, ~96% (based on denaturant-free substance)

32221

Sigma-Aldrich

Ethanol, puriss. p.a., absolute, ≥99.8% (GC)

60004

Sigma-Aldrich

Acetonitrile, ≥99.5% (GC)

34442

Sigma-Aldrich

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

34888

Sigma-Aldrich

Acetonitrile, for HPLC, for UV, ≥99.9% (GC)

439134

Sigma-Aldrich

Acetonitrile, suitable for HPLC, gradient grade, ≥99.9%

493546

Sigma-Aldrich

Ethyl alcohol, Pure, 200 proof, meets USP testing specifications

792799

Sigma-Aldrich

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

PHR1551

Supelco

Acetonitrile, Pharmaceutical Secondary Standard; Certified Reference Material

792802

Sigma-Aldrich

Ethyl alcohol, Pure, 160 proof, Excise Tax-free, Permit for use required

360457

Sigma-Aldrich

Acetonitrile, ACS reagent, ≥99.5%

459828

Sigma-Aldrich

Ethyl alcohol, Pure, 200 proof, HPLC/spectrophotometric grade

E7023

Sigma-Aldrich

Ethyl alcohol, Pure, 200 proof, for molecular biology

792780

Sigma-Aldrich

Ethyl alcohol, Pure, 200 proof, ACS reagent, meets USP testing specifications, Excise Tax-free, Permit for use required