Method of scavenging free radicals using orange extract

The present invention relates to the use of orange extract for the preparation of pharmaceutical, cosmetic, food or agricultural compositions with an anti-radical activity. The orange extract is substantially free from ascorbic acid.

BACKGROUND OF THE INVENTION 
The present invention relates to a method of scavenging free radicals by 
incorporating orange extract into a composition. 
The deleterious effects of free radicals are, in many cases, known but are 
continually being investigated. For example, it is believed that one of 
the causes of the deterioration of skin over time is the formation of free 
radicals within the elements forming the skin such as the fibers, cells 
and enzymes. The formation of free radicals may be caused, in part, by the 
action of external radiation of sunlight. The free radicals thus formed 
may attack the skin elements through a chain reaction that continues as 
long as the free radical exists, i.e., as long as the free radical does 
not combine with something (typically called a scavenger or inhibitor) 
that produces a stable molecule. The adverse action of these free radicals 
at the cutaneous integument level may cause destruction of cell DNA and an 
erosion of the connective tissues (collagen and elastin). Thus, the skin 
looks more aged. 
As a result, substances that interrupt or inhibit the formation of free 
radicals during the free radical chain reaction are continually being 
sought. At the present, well known scavengers (also referred to as 
inhibitors or antioxidants, and as used in the following specification and 
appended claims should be considered synonymous with scavenger) include 
phenols, aromatic amines, sulfur compounds, raw seed oils, wheat germ oil, 
tocopherols, and gums. It is also believed that phosphoric acid, citric 
acid, and ascorbic acid have their own scavenging activity or enhance the 
activity of the known scavengers mentioned above. 
Thus, French Demande 2,597,337-A2 describes a cosmetic to slow down the 
aging process by attacking free radicals. The cosmetic includes a first 
water soluble active ingredient, which may include a source of vitamin C 
such as from acerola cherry extract, and a second fat soluble ingredient, 
which includes free radical scavengers such as vitamin E. 
While the above French Demande describes the use of acerola cherry extract, 
it is particularly with respect to the presence of vitamin C, known to be 
present in relatively high amounts when compared to other natural sources 
of vitamin C. Likewise, Japanese Laid-open applications 2-200610 and 
70-61915 describe that an extract obtained from fruits of acerola may be 
incorporated into cosmetics. Those applications teach that the acerola 
cherry contain vitamin C or ascorbic acid and application no. 2-200610 
describes the extract as containing at least 1% L-ascorbic acid and it is 
the presence of the L-ascorbic acid that provides the desired whitening 
effect. This is not surprising since, it is well known that L-ascorbic 
acid provides a whitening effect. 
There is, however, no appreciation that an orange extract, which is 
substantially free of vitamin C, has free radical scavenging activity. The 
present inventors have now found that an orange extract exhibits free 
radical scavenging activity and, as a result may find particular 
application in the preparation of pharmaceutical, food and agricultural 
compositions, and more particularly in cosmetics for external skin 
application. Thus, the ability of the orange extract used in the present 
invention to scavenge free radicals is surprising and unexpected. 
SUMMARY OF THE INVENTION 
It is an object of the present invention to provide compositions that 
inhibit the formation of or scavenge the presence of free radicals. The 
present invention also contemplates that compositions containing known 
free radical scavengers can be enhanced by incorporating orange extract. 
Thus, the present invention includes a composition of the type containing 
free radical scavengers wherein the improvement comprises adding orange 
extract. Preferably, the composition is a cosmetic composition suitable 
for use on human skin. For example, the orange extract may have 
application alone or in conjunction with skin whitening agents for use as 
a skin whitener. 
The present invention also includes a method of providing free radical 
scavenging activity in a composition comprising admixing to the 
composition an effective amount of orange extract to increase the free 
radical scavenging activity of the composition. 
It is known that the superoxide ions produced during oxidation reactions 
caused by molecular oxygen are very active and in particular attack 
proteins and nucleic acids. Thus, the present invention is based on the 
anti-radical activity of orange extract towards the superoxide radical. 
The compositions of the present invention incorporating the orange extract 
are particularly suitable for protecting the skin due in particular to the 
protection against phototoxic reactions and UV aggression. 
The present invention also includes a method of scavenging free radicals in 
human skin comprising topically applying to the skin a composition 
comprising orange extract. Preferably, the composition further includes a 
pharmaceutically acceptable carrier. 
The orange extract useful in the present invention is substantially free 
from ascorbic acid and contains only minor amounts of organic acids 
particularly carboxylic acids. 
The term "substantially free from ascorbic acid" as used in the 
specification and accompanying claims means that any ascorbic acid present 
in the orange extract is present in an amount no greater than about 1% by 
weight, preferably no greater than about 0.1%, and more preferably no 
greater than about 0.01% of the orange extract. Most preferably, the 
ascorbic acid, if present, is present in an amount no greater than about 
0.005% of the orange extract. 
The term "minor amounts of organic acids" and "minor amounts of carboxylic 
acids" as used in the specification and accompanying claims means that any 
organic acid and any carboxylic acid, which includes saturated and 
unsaturated carboxylic and dicarboxylic acids, hydroxymonocarboxylic 
acids, hydroxydicarboxylic acids, and hydroxytricarboxylic acids, is 
present in an amount no greater than about 5% by weight, preferably no 
greater than about 3%, and more preferably no greater than about 2% of the 
orange extract. 
In the composition according to the present invention, the amount of orange 
extract to be used can not be absolutely specified because it varies 
according to the form of preparation. However, it is generally used in an 
amount from about 0.01% to about 50%, preferably from about 0.01% to about 
5%, desirably from about 0.05% to about 2%, more preferably from about 
0.1% to about 1% based on the whole weight of the composition. 
It is noted that, unless otherwise stated, all percentages given in this 
specification and the appended claims refer to percentages by weight. 
DETAILED DESCRIPTION OF THE INVENTION 
In accordance with the present invention, a free radical scavenger 
composition is provided which comprises orange extract as an active 
ingredient. 
In the present invention, however, the orange extract is substantially free 
from any substantial amount of vitamin C (ascorbic acid or its 
derivatives). In other words, the orange extract does not contain more 
than about 1% by weight, preferably no greater than about 0.1%, and more 
preferably no greater than about 0.01% of the orange extract. Most 
preferably, the ascorbic acid, if present, is present in an amount no 
greater than about 0.005% of the orange extract. 
The orange extract can be made in any suitable manner to achieve an extract 
that is substantially free of ascorbic acid. Preferably, the orange 
extract is obtained from orange peel, flowers, and/or leaves on an orange 
tree. It is believed that the orange extract is prepared by drying the 
material, pretreating it, cold extracting the the pretreated material by 
alternating maceration and percolation for 24 to 48 hours, filtrating, 
microfiltrating, and adding a suitable preservtive such as parabens or 
phenoxyethanol. Most preferably, the orange extract is Bitter Orange Peel 
Extract HB from Centerchem, Inc. or Herbasol Orange Extract from 
Cosmetochem. It is believed that the Centerchem material is obtained from 
the orange peel while the Cosmetochem material is obtained from the orange 
flower. 
In the composition according to the present invention, the amount of orange 
extract to be used can not be absolutely specified because it varies 
according to the form of preparation. However, it is generally used in an 
amount from about 0.01% to about 50%, preferably from about 0.01% to about 
5%, desirably from about 0.05% to about 2%, more preferably from about 
0.1% to about 1% based on the whole weight of the whitener composition. 
The composition of the present invention may be prepared in various forms. 
For example, it may be in the form of a cosmetic preparation such as 
cream, cosmetic lotion, pack or powder, or as an emulsion, lotion, 
liniment or ointment. In each formulation, various known conventional 
cosmetic ingredients may be incorporated. For example, cosmetic 
ingredients such as alcohols, fats and oils, surfactants, fatty acids, 
silicone oils, humectants, viscosity modifiers, emulsifiers, stabilizers, 
coloring agents, and perfumes. 
In another aspect of the composition of the present invention there is 
provided an improved free radical scavenging composition wherein the 
improvement comprises adding orange extract, as fully defined above. In 
this aspect, it may be desirable to incorporate the orange extract in, for 
example, a skin whitening composition. In this instance, the free radical 
scavenging activity of the orange extract can inhibit the conversion of 
dopaquinone with the result of inhibiting the production of eumelanin. 
Consequently, it is contemplated that the free radical scavenging activity 
of the orange extract can augment the known tyrosinase inhibitors and 
thus, provide enhanced skin whitening properties. 
The present invention also contemplates a method of enhancing the free 
radical scavenging activity of known free radical scavengers that 
comprises adding orange extract to compositions containing known free 
radical scavengers. 
The present invention also includes a method of providing free radical 
scavenging activity in a composition comprising admixing to the 
composition an effective amount of orange extract to increase the free 
radical scavenging activity of the composition. 
In another aspect of the present invention, a method of scavenging free 
radicals in human skin is provided and comprises topically applying to the 
skin orange extract. More preferably, the method comprises topically 
applying to the skin a composition comprising orange extract and a 
pharmaceutically acceptable carrier. The term "pharmaceutically 
acceptable" means those drugs, medicaments, or inert ingredients which are 
suitable for use in contact with the tissues of humans and lower animals 
without undue toxicity, incompatibility, instability, irritation, and the 
like, commensurate with a reasonable benefit/risk ratio. 
To demonstrate the free radical scavenging activity of the orange extract 
of the present invention, the test described in Cosmetics & Toiletries, 
Vol. 10, pp. 51-56, October 1995) was conducted. 
A control solution was prepared by measuring 39.43 mg of 
2,2-diphenyl-1-picrylhydrazyl hydrate (DPPH), a known source of free 
radicals, and diluting it to 1000 ml with ethanol. A control cuvette was 
charged with 1.75 ml of the control and 1.75 ml of ethanol. A blank 
cuvette was charged with 3.5 ml of ethanol. The cuvettes were placed in an 
oven for 30 minutes at 37.degree. C. The absorbency was measured at 516 nm 
using a UV/vis spectrophotometer by placing the blank cuvette into the 
reference slot and the control cuvette into the sample slot.

EXAMPLE 1 
An orange extract was prepared according to the method described above. A 
test cuvette was charged with 1.75 ml of the control solution and 1.75 ml 
of a 1% solution of orange extract. A reference cuvette was charged with 
1.75 ml of the orange extract and 1.75 ml of ethanol. The cuvettes were 
placed in an oven for 30 minutes at 37.degree. C. The absorbency was 
measured in the manner described above with the test cuvette placed in the 
sample slot and the reference cuvette placed in the reference slot. 
The free radical scavenging activity was calculated as a percentage of 
inhibition of the generation of the free radicals using the following 
formula: 
EQU Percent Inhibition=100.times.1-(DT/DR)! 
DT is the measured absorbance of the test cuvette with respect to the 
reference cuvette and DR is the measured absorbance of the control cuvette 
with respect to the blank cuvette. 
The orange extract exhibited 52% inhibition. 
This test shows that the orange extract has anti-radical activity and would 
therefore be expected to show beneficial results when incorporated into 
compositions. 
EXAMPLE 2 
Scavenging activity tests were conducted in the manner described in Example 
1 using 1% solutions of (a) tocopheryl acetate, (b) rose hips oil, (c) 
tioxolone, and (d) Lanachrys 2B. The results are presented in Table 1 
below. 
TABLE 1 
______________________________________ 
WHITENER % INHIBITION 
______________________________________ 
Tocopheryl Acetate 
67 
Rose Hips Oil 63 
Tioxolone 50 
Lanachrys 2B 40 
______________________________________ 
It should be understood that a wide range of changes and modifications can 
be made to the embodiments described above. It is therefore intended that 
the foregoing description illustrates rather than limits this invention, 
and that it is the following claims, including all equivalents, which 
define this invention.