Safety and Feasibility of Topical Application of Limonene as a Massage Oil to the Breast 753
delivery with minimal systemic side effects. Studies have
shown that topical application of 4-hydroxyta- moxifen,
the active metabolite of tamoxifen, to the breast resulted
in effective mammary tissue concentration of 4-hydroxy-
tamoxifen with much lower plasma levels [14]. A re-
cent study showed that topical application of 4-hydroxy-
tamoxifen to the breast resulted in reductions in breast
tumor tissue proliferation indexes similar to that seen
with oral administration of tamoxifen [15].
In our study, similar limonene levels were observed in
the mouse mammary pads after oral and topical limonene
administration as orange oil blended with base oil. Two
different limonene dose levels (equivalent to 6 and 12 mg
limonene dose daily) did not result in a dose dependent
increase in the limonene levels in the mammary pads.
Consistent with its high lipophilicity and previous find-
ings in humans [16] and rodents [17], limonene demon-
strated preferential mammary tissue-to-plasma distribu-
tion in mice. Plasma concentrations of limonene were on
average between 0.015 - 0.08 µM, with mammary tissue
levels over 1000-fold higher.
With the demonstration of mammary tissue bioavail-
ability of limonene following topical application in mice,
we conducted a pilot clinical study to determine the
safety and feasibility of topical limonene application in
healthy women. Our pilot study showed that topical ap-
plication of limonene containing massage oil was met
with excellent compliance and good safety profile. Side
effects associated with orange oil application were
mostly mild and limited to dermatological events at the
application site. The rashes developed at the application
site were most likely attributed to an oxidized limonene
by-product and could be avoided by pre-testing the con-
tent of the oxidized limonene by-product in the orange
oil supplies. Additionally, this study demonstrates that
women are willing to apply a topical agent to their breast
daily, providing a potential alternative for women un-
willing or unable to take a daily oral drug.
Interestingly, limonene was detectable at baseline
specimens in most participants despite of a 4-week
washout period. Because limonene is present in many
household cleaning products [18], in the environment
[19], and in foods [20,21], presence of low levels of
limonene in plasma and NAF at baseline was not unex-
pected. However, limonene levels were not changed in
NAF or plasma after four weeks of daily orange oil ap-
plication even though prior in vitro studies have shown
that topically applied limonene can penetrate through
human epidermis and dermis [8]. It is plausible that topi-
cally applied limonene as massage oil has limited ab-
sorption through human breast skin. It is also likely that
dermally absorbed limonene, if any, may concentrate in
the breast adipose tissue with minimal secretion into
NAF or distribution into systemic circulation. Future
clinical studies should consider incorporation of direct
breast tissue sampling such as in women undergoing
breast reduction surgery to assess tissue limonene levels.
In summary, topical administration of limonene to
mice led to high mammary tissue limonene levels with-
out affecting normal mammary gland morphology. A
similar formulation given to women for topical applica-
tion to the breast was met with high compliance and tol-
erability. Future research should assess the efficacy of
topical limonene application in mammary cancer models
and develop topical formulations that may further en-
hance breast tissue drug delivery.
5. Acknowledgements
We would like to thank Donna Vining and Heidi Fritz for
their assistance in the clinical study conduct.
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