HYDROGEN-DONATING MECHANISM OF ROSMARIQUINONE
1149
following conditions: (i) fluorescent radiation, (ii) rose ben-
gal (1.5 mmol; a photosensitizer) and fluorescent radiation,
or (iii) rose bengal (1.5 mmol), molecular oxygen and fluo-
rescent radiation. A second model study included the addi-
tion of RQ (500 mg; 1.77 mmol) to methyl oleate (5 g; 16.86
mmol) and was stored in a 60°C oven for 65 d.
All reactions were monitored by thin-layer chromatogra-
phy (TLC) and exposed to the appropriate conditions until
TLC indicated that RQ had been consumed by the reaction
conditions. The methyl oleate and RQ mixtures were sepa-
rated using flash chromatography on silica gel, eluted with
hexane/ethyl acetate (100:0; 98:2; 95:5; 90:10; 80:20) and
methanol (100%), and collected in 25-mL fractions. This
method proved successful for the separation of RQ, arucadiol
(AD), and the oxidized methyl oleate.
local supermarket and stored in the dark at −18°C until needed.
The oil was stripped using the modified method of Hall et al.
(4), which included a batch process rather than a column (17).
The stripping process continued until tocopherols, chlorophyll,
and carotenoids were no longer detected. Solvent was evapo-
rated under vacuum at 30°C, and the stripped SBO was stored
at −18°C for no longer than four weeks.
Oxidation of bulk soybean oils. Stripped SBO (100 g) was
weighed into 110-mL glass jars. Rosmariquinone, arucadiol,
and TBHQ were added separately to stripped SBO at a con-
centration of 200 ppm (0.02%). Each sample was thoroughly
mixed to assure complete dispersion of the antioxidants.
TBHQ served as the positive control, and an untreated sam-
ple served as the negative control.
The jars were randomly placed under two, 15-W cool flu-
orescent lamps at an illuminance of 4200 lx of fluorescent ra-
diation at 25 1°C, or they were placed in an oven (60°C).
Peroxide values (PV) were analyzed every 24 h (12 h when
necessary), until a peroxide level of 20 meq/kg was reached
in the stripped oil, using the American Oil Chemists’ Society
(18) official methods.
Statistical Analysis. All studies were completed in tripli-
cate and analyzed by analysis of variance (ANOVA) with Sta-
tistical Analysis System (SAS) software (19). Least signifi-
cant differences (20) were determined between the mean val-
ues of the treatments at the 95% confidence level (P < 0.05).
1H nuclear magnetic resonance (NMR) and 13C NMR spec-
tra were recorded on a 300-MHz NMR instrument (Oxford In-
struments, Palo Alto, CA; 300 MHz for 1H, 75 MHz for 13C).
Chemical shift data were reported as parts per million (δ) and
referenced to the residual chloroform peak of deuteriochloro-
form (δ 7.25 ppm) for 1H NMR and to the center peak of the
residual chloroform triplet (δ 77.0 ppm) for 13C NMR. All OH
functional groups were confirmed using deuterium oxide. In-
frared (IR) absorption spectra were recorded on an Analect
model RFX-30 FT-IR (Analect Instruments, Inc., Irvine, CA)
spectrophotometer with a potassium bromide cell. UV data
were collected using a Milton Roy Genesys 5 UV/VIS spec-
trophotometer [Spectronic Instruments, Inc. (formerly Milton
Roy Co.), Rochester, NY]. Mass spectra were obtained using a
Hewlett-Packard (Wilmington, DE) 5790 gas chromatogra-
phy–mass spectrometry (GC–MS) system and a Chiral-val col-
umn (Alltech, State College, PA) using both chemical and elec-
tron impact ionization. Uncorrected melting point determina-
tions were completed in open pyrex capillary tubes and a
Mel-Temp apparatus (Fisher Scientific, St. Louis, MO). All
flash column chromatography was completed using E. Merck
silica gel 60 (Aldrich, Milwaukee, WI) and TLC on commer-
cial silica gel plates (Analtech Silica HLF 250 m; Analtech,
Inc., Newark, DE).
Analysis of soybean oil components. Tocopherol (α, δ), β-
carotene, and chlorophyll standards were obtained from
Sigma Chemical Co. (St. Louis, Mo). Gamma (γ)-tocopherol
was obtained from Eastman Chemical Products Inc.
(Kingsport, TN). The tocopherols were determined by high-
performance liquid chromatography (12). Chlorophyll and β-
carotene were spectrophotometrically analyzed by Associa-
tion of Official Analytical Chemists (AOAC) methods
(13,14) using the modifications of Hall and Cuppett (15).
Antioxidants. Tertiary butylhydroquinone (TBHQ) was
obtained from Eastman Chemical Products Inc. (Kingsport,
TN). RQ was synthesized in this laboratory by a modified
method of Lee et al. (16), which included optimization of the
reactants. Arucadiol, the red crystalline compound from the
model systems, isolated as previously described, was also
tested. All compounds were tested at the 200 ppm level.
Stripping of SBO. Commercial SBO was purchased from a
RESULTS AND DISCUSSION
Spectral data. Arucadiol (miltiodiol) was purified from the
crude reaction mixtures of the model test systems to give a
red-orange crystal: IR (KBr) λmax: 3471, 2957, 2925, 2866,
1633, 1590, 1506, 1461, 1416, 1320, 1177, 1031, 965 cm−1;
1H NMR: δ 10.65 (s, 1 H, OH), 7.95 (d, J = 7.97 Hz, 1H),
7.32 (d, J = 7.97 Hz, 1 H), 7.25 (s, 1 H), 6.90 (s, 1 H, OH),
3.24 (sept, J = 6.82, 1 H), 2.90 (t, J = 6.82 Hz, 2 H), 2.10 (t, J
= 6.82 Hz, 2 H), 1.45 (s, 6 H), 1.35 (d, J = 7.97, 6 H), (Fig.
1B); 13C NMR: δ 204.3, 158.2, 144.9, 138.1, 137.7, 136.5,
127.8, 125.3, 120.3, 120.2, 118.6, 36.2, 35.9, 35.4, 29.6 (2
C), 27.6, 22.2 (2 C) (Fig. 2B); low-resolution MS (chemical
ionization, methane): m/z 298.1 (M), 299.1 (M + 1), 300.1 (M
+ 2); UV-Vis (methanol): λmax (ε) 275 (17,730), 410 (2184).
Evaluation of model test systems. All methyl oleate–RQ
model systems gave two major oxidation compounds: sev-
eral oxidized methyl oleate products and AD. Two minor
compounds were also isolated but not characterized. The AD
structure was characterized by IR, UV/Vis, NMR, and mass
spectrometry and compared to the spectral data of RQ. The
spectral data of AD were identical to previously reported data
(10,11). The infrared spectrum of AD had a broad peak at
3471 cm−1, indicating the presence of a hydroxyl group. This
peak was lacking, as expected, for RQ, and the increase in the
molecular weight by 16 m/z on the mass spectrum indicated
the addition of an oxygen atom. UV/Vis λmax had shifted
from 257 nm in RQ to 275 nm in AD, indicating the presence
of phenolic hydrogens at C-11 and C-12 positions. The phe-
JAOCS, Vol. 75, no. 9 (1998)