Method for testing skin for presence of moisturizer

A method for testing skin for presence of moisturizing substances is provided which method includes the steps of applying to the skin a moisturizer formulation which includes protein, urea and/or glucose and/or other reducing sugar such as fructose all of which bind with water and form a film or coating on the skin which inhibits moisture loss from the skin, and subsequently testing the skin for the presence of protein, urea and/or glucose and/or other reducing sugar remaining on the skin which is directly correlated to the amount of moisturizer formulation remaining on the skin. When it is determined through the testing that there has been at least a predetermined loss of protein, urea and/or glucose or other reducing sugar, additional moisturizer is then applied to the skin.

FIELD OF THE INVENTION 
The present invention relates to a method for testing skin for the presence 
of certain moisturizing substances such as protein, urea and/or glucose or 
other reducing sugar by the use of test strips sensitive to these 
materials. 
BACKGROUND OF THE INVENTION 
The presence of water in the outer dead layers of the epidermis is 
essential to prevent what is generally referred to as "dry skin". If water 
is depleted from the stratum corneum more rapidly than it is received from 
the lower layers of the epidermis, the skin becomes dehydrated and loses 
its flexibility. Moisturizing compositions are essential and quite 
effective in restoring water lost from the stratum corneum and thereby 
prevent the above conditions. 
One of the more important properties of a moisturizer composition is its 
substantivity or staying power, that is, the ability to form a film or 
coating which will remain on the skin even after washing. The actives 
which may be present, such as protein, urea and/or glucose and/or other 
reducing sugars, bind with water and aid in forming the film or coating. 
However, many moisturizer compositions available today are non-greasy 
compositions which are difficult to detect once applied to the skin. Thus, 
the user of such compositions may not be aware that the moisturizing film 
or coating has been washed off or otherwise removed and as a result may 
develop dry skin. To avoid this problem many users of such moisturizer 
composition may randomly apply same to the skin several times throughout 
the day regardless of the substantivity of the moisturizing composition. 
Until now, there was no known accurate, simple, quick technique available 
for determining the presence of moisturizer on the skin and when 
additional moisturizer is necessary to avoid dry skin and therefore should 
be applied. 
DESCRIPTION OF THE INVENTION 
In accordance with the present invention, a method is provided for 
determining the presence of moisturizing substances on skin, which method 
includes the steps of forming a moisturizer composition which includes one 
or more actives such as protein, urea and/or glucose and/or other reducing 
sugar or carbohydrate which binds with water, applying the moisturizer 
composition to desired areas of the skin to form a film or coating on the 
skin which inhibits moisture loss from the skin, and subsequently testing 
skin previously treated with such moisturizer composition for the presence 
of protein, urea and/or glucose and/or other reducing sugar or 
carbohydrate remaining on the previously treated skin the presence of 
which is directly correlated to the amount of moisturizer remaining on the 
skin. 
The testing step for detecting the presence of actives such as protein, 
urea and/or glucose and/or other reducing sugar or carbohydrate is 
accomplished through the use of known test indicators for these materials. 
For example, the presence of protein on the skin as a result of the 
previous application of moisturizer composition may be detected through 
the use of the protein test device disclosed in the working Examples of 
U.S. Pat. No. 3,438,737 to Atkinson et al; the presence of urea on the 
skin as a result of the previous application of moisturizer composition 
may be detected through the use of the urea test device disclosed in the 
working Examples of U.S. Pat. Nos. 3,461,036 to Harvill et al, 3,427,225 
to Harvill et al, 3,395,082 to Mast, and 3,145,086; and the presence of 
glucose and/or other reducing sugar on the skin as a result of the 
previous application of moisturizer may be detected through the use of the 
glucose test device disclosed in the working Example of U.S. Pat. No. 
3,814,668 to Blake et al. 
The moisturizer compositions may contain one or more actives such as the 
protein, urea and/or glucose and/or other reducing sugars or 
carbohydrates. Thus, the actual testing step carried out will vary 
depending upon which one or more of these indicators were present in the 
moisturizer composition as previously applied. For example, where only one 
of the above materials is present in the moisturizing composition, a 
single test strip device as disclosed in the appropriate patent for 
testing of the material, may be employed. Where two or more of protein, 
urea and/or glucose and/or other reducing sugar are present, then one or 
more appropriate test strips each separately or combined into a single 
test strip or device may be employed. 
The moisturizer composition will contain at least one of the protein, such 
as albumin, urea and/or glucose and/or other reducing sugar such as 
fructose, lactose or maltose and may be of conventional composition. Thus, 
the moisturizer composition will contain from about 0.1 to about 10% and 
preferably from about 0.1 to about 2% by weight of a protein such as 
albumin, from about 0.1 to about 10% and preferably from about 0.1 to 
about 5% by weight urea, and from about 0.1 to about 10% and preferably 
from about 0.1 to about 5% by weight glucose or other reducing sugar. The 
protein, urea, glucose and/or other reducing sugar will serve to bind 
water and form a film or coating which may be occlusive and prevent loss 
of moisture from the skin. Each test strip or section of combined test 
strip will be specific to one of the above materials and will vary in 
color depending upon the concentration of these materials remaining on the 
skin. The actual color change or lack of color change obtained will 
determine whether additional moisturizer composition should be applied. 
The moisturizer composition for use in the method of the invention may take 
the form of a cream or lotion and will include actives or highly effective 
moisture regulators based on naturally occurring carbohydrates, such as 
glucose and/or other reducing sugars, as well as proteins, such as serum 
albumin or egg albumin and/or urea which are similar to those found in the 
outer layers of skin. The actives combine with water to form an active 
complex in the form of a film or coating which effectively adheres to skin 
to assure sustained moisturizer activity. Such complex is even effective 
at low humidities when skin is most likely to suffer from dryness. 
Employing the method of the invention, using the test strips described in 
the aforementioned patents, a person will be able to confirm the 
effectiveness of the moisturizer. The test strips react with the active 
complex contained in the cream or lotion to change color. To test, the 
subject merely wets the test strip and places it on his skin for 30 
seconds or more. No color change in the test strip indicates reapplication 
of moisturizer is necessary. Where there is a color change in the test 
strip, such color change will be approximately proportioned to the amount 
of active complex remaining on the skin from previous applications of 
moisturizer compositions. 
The moisturizer formulations employed in the method of the invention will 
also contain conventional moisturizer ingredients necessary in formulating 
a desirable product, such as, one or more diluents, thickeners, 
stabilizers, coloring agents, humectants, preservatives, emollients, 
bodying agents, sunscreen agents and the like. The moisturizer 
formulations of the invention may contain one or more diluents such as 
deionized water in an amount within the range of from about 40 to about 
90% and preferably from about 50 to about 80% by weight, optionally one or 
more thickeners, such as magnesium aluminum silicate, Carbomer 934 and 
xanthan gum in an amount within the range of from about 0.1 to about 2% 
and preferably from about 0.1 to about 0.5% by weight, optionally one or 
more skin protecting agents, such as panthenol, which serves as a skin 
moisturizer and humectant, in an amount within the range of from about 0.1 
to about 5% and preferably from about 0.1 to about 1% by weight, one or 
more other humectants such as polyethylene glycols (for example, Carbowax 
400), sodium 2-pyrrolidone carboxylic acid, sorbitol, propylene glycol or 
glycerine in an amount within the range of from about 1 to about 20% and 
preferably from about 1 to about 5% by weight, one or more preservatives 
such as parabens including methyl paraben, propyl paraben, butyl paraben, 
Glydant (dimethyldimethoyl hydantoin), benzyl alcohol, imidazolidinyl urea 
and the like usually employed in amounts within the range of from about 
0.1 to about 1% by weight and preferably from about 0.5 to about 0.8% by 
weight, one or more emollients or emollient oils such as mineral oil, 
avocado oil, petrolatum, propylene glycol dicaprylate/dicaprate and 
isopropyl myristate in an amount within the range of from about, 10 to 
about 20% by weight and preferably from about 10 to about 15% by weight, 
one or more co-emulsifiers such as PEG 20 sorbitan monolaurate 
(Polysorbate 20), diethanolamine cetyl phosphate, glyceryl stearate, 
polyethylene glycol 100 stearate, and PEG 20 stearyl ether (Brij 78, 
Steareth 20) or PEG 150 distearate in an amount within the range of from 
about 0.1 to about 7% by weight and preferably from about 0.2 to about 5% 
by weight; one or more bodying agents such as stearic acid, glyceryl 
monostearate, and the like in an amount within the range of from about 1 
to about 10% by weight and preferably from about 1 to about 5% by weight, 
optionally one or more sun screen agents such as octyl dimethyl 
p-aminobenzoic acid, octyl salicylate, benzophenone 3 and the like in an 
amount within the range of from about 0.5 to about 10% by weight and 
preferably from about 1 to about 5% by weight, and optionally one or more 
antioxidants such as dl-alpha-tocopherol in an amount within the range of 
from about 0.05 to about 0.5% by weight and preferably frm about 0.05 to 
about 0.2% by weight. In addition, the formulations of the invention may 
contain one or more fragrances, solubilizing agents and emulsifiers for 
the fragrances such as polyoxyethylene (13) octyl phenyl ether. 
The actual structure and compositions of the test devices employed are 
disclosed in U.S. Pat. Nos. 3,814,668 (reducing sugar such as glucose), 
3,438,737 (protein such as albumin), and 3,461,036, 3,427,225 and 
3,395,082 (urea) and the subject matter thereof is incorporated herein by 
reference. As described in the above patents, the presence and 
concentration of each of the above actives may be detected by wetting one 
or more appropriate test strips, applying the test strips to the skin and 
observing color changes produced on the test strips. 
The moisturizer composition may be prepared as follows. 
Deionized water (diluent) together with glycerine and/or other 
water-soluble humectants, optionally one or more wetting agents, one or 
more actives such as glucose (and/or other reducing sugar), urea and/or 
protein, and optionally one or more thickeners, optionally one or more 
preservatives such as methyl paraben are mixed together to form a first 
mixture (A). Then, one or more water-insoluble humectants, emollients, 
thickeners, emulsifiers, as well as preservatives etc. are mixed together 
to form the oil phase or a second mixture (B) which is mixed with mixture 
(A) to form mixture (AB). A third phase formed of an aqueous solution of 
preservative and surfactant may be added to form the final mixture which 
is cooled to form the moisturizer composition.

The following Examples represent preferred embodiments of the present 
invention. All temperatures are expressed in degrees Centigrade unless 
otherwise indicated. 
EXAMPLES 1 AND 2 
Moisturizer formulations in the form of creams having the following 
compositions are prepared as described below. 
______________________________________ 
Ingredient Parts by Weight 
Example No. 1 2 Control 
______________________________________ 
Blend A 
Deionized water 78.5 79.5 79 
(emollient) 
Glycerine (humectant) 
2 2 2 
Methyl paraben 0.2 0.2 0.2 
(preservative) 
Glucose (active) 2 -- -- 
Urea (active) -- 1 -- 
Blend B 
Octyl palmitate 5 5 5 
(emollient) 
Dimethicone (emollient) 
2 2 2 
Polyethylene glycol 20 
3 3 3 
sorbitan monostearate 
(emulsifier) 
Glyceryl monostearate and 
2 2 2 
Sodium lauryl sulfate 
(bodying agent) (95:5) 
Cetyl, stearyl alcohol and 
4.5 4.5 4.5 
polyethylene glycol 
stearyl ether (1:1) 
(emulsifier) 
Propyl paraben 0.1 0.1 0.1 
Blend C 
Deionized water 0.5 0.5 0.5 
Quaternium 15 0.1 0.1 0.1 
(Dowicil 200) 
______________________________________ 
The ingredients of each of the three Blends A, B, and C were mixed while 
heating to form respective Blends A, B and C. Blend B was added to Blend A 
with homomixing for one hour. Blend C was then mixed with Blend AB for 15 
minutes under slow speed mixing while heating to 75.degree. C. The mixture 
was cooled to 30.degree. C. to form the moisturizer formulations. 
A glucose test strip is prepared as outlined in the Example in U.S. Pat. 
No. 3,814,668 and as set out below. 
The following components are mixed, stepwise for the preparation of 100 ml 
of mix. 
______________________________________ 
Component Quantity 
______________________________________ 
(1) Potassium iodide 1.0 g 
(2) FD and C Blue No. 1 10.4 mg 
(3) Distilled water 53 ml 
(4) Citric acid, anhydrous 
0.595 g 
(5) Sodium citrate.2H.sub.2 O 
5.23 g 
(6) Gantrez AN 139 copolymer of methyl 
10 ml 
vinyl ether and maleic anhydride 
(10% aqueous solution) 
(7) Polyvinylpyrrolidone (10% 
5 ml 
aqueous solution) 
(8) Peroxidase .sup.1 50 
mg 
(9) Glucose oxidase .sup.2 33 
mg 
______________________________________ 
.sup.1 3000 Worthington units/mcg 
.sup.2 1000 units/ml 
Whatman 3 MM paper is impregnated with the above solution, allowed to drain 
and dried in a tunnel dryer set at 85.degree. C. for 7 minutes. After 
removal from the drying tunnel, the impregnated paper is dipped into a 
solution of 0.85 grams of ethyl cellulose in 100 ml of benzene. After 
removal from this second impregnating solution, the paper is again dried, 
cut into small squares approximately 5 mm by 5 mm and attached to an end 
of a strip of fluid impervious plastic film 5 mm wide and approximately 6 
cm long. 
The Example 1 and Control moisturizer compositions are applied to the skin. 
Excellent glucose quantitation is achieved employing a formulation as 
described above. 
A urea test strip is prepared as outlined in Example 1 of U.S. Pat. No. 
3,461,036. 
The composition is formulated as follows: 
______________________________________ 
Gelatin 0.5 g 
Urease 0.5 g 
Acetamide 1.0 g 
4% "Carbowax 4000" (polyethylene 
11.5 ml 
glycol) 
0.1 M ammonium citrate buffer 
2.5 ml 
(dibasic) 
1.6% aqueous solution bromothymol 
3.8 ml 
blue 
______________________________________ 
The gelatin is added to 11.5 ml of water and heated until completely 
dissolved. The remaining ingredients are combined and then mixed with the 
solution of gelatin until a clear solution is obtained. The temperature of 
the final solution is approximately 30.degree. C. The solution pH is then 
adjusted to a pH of about 6.5 by the addition of small quantities of 
dilute sodium hydroxide. Paper strips measuring 2 inches by 1/4 inch are 
then dipped in the solution and dried at a temperature of 85.degree. C. 
The dried strips are then coated with a semi-permeable polymeric film by 
dipping them into a 1.25% solution of ethyl cellulose in benzene and air 
drying until the benzene is completely evaporated. 
The Example 2 and Control A moisturizer compositions are applied to the 
skin. 
Excellent urea quantitation is achieved employing a formuation as described 
above. 
EXAMPLES 3 TO 7 
Moisturizer formulations in the form of creams having the following 
compositions are prepared as described in Examples 1 and 2. 
______________________________________ 
Ingredient Parts by Weight 
Example No. 3 4 5 6 7 
______________________________________ 
Blend A 
Deionized water (diluent) 
78.5 78.5 79 79.5 79 
Glycerine (humectant) 
2 2 2 2 2 
Methyl paraben (preservative) 
0.2 0.2 0.2 0.2 0.2 
Glucose (active) 2 -- 2 -- 2 
Urea (active) 1 -- -- 1 1 
Albumin (active) -- 1 1 1 1 
Blend B 
Octyl palmitate (emollient) 
5 5 5 5 5 
Dimethicone (emollient) 
2 2 2 2 2 
Polyethylene glycol 20 sorbitan 
3 3 3 3 3 
monostearate (emulsifier) 
Glyceryl monostearate and 
2 2 2 2 2 
sodium lauryl sulfate (1:1) 
(bodying agent) 
Cetyl, stearyl alcohol and 
4.5 4.5 4.5 4.5 4.5 
polyethylene glycol (emulsifier) 
stearyl ether (1:1) 
Propyl paraben 0.1 0.1 0.1 0.1 0.1 
Blend C 
Deionized water 0.5 0.5 0.5 0.5 0.5 
Quaternium 15 (Dowicil 200) 
0.1 0.1 0.1 0.1 0.1 
______________________________________ 
A protein test strip is prepared as outlined in U.S. Pat. No. 3,438,737. 
The compositions set out below in the Table are prepared as follows: a 
buffer solution is first prepared by mixing the sodium citrate, citric 
acid and water. The solution is then combined with the indicated amount of 
a 0.1% aqueous solution of tartrazine and the combination mixed with a 
solution comprising the indicated amounts of ethanol and tetrabromphenol 
blue. Strips of Eatman and Dikeman No. 641 filter paper are dipped into 
the various compositions, allowed to drain and dried at 100.degree. C. for 
11 minutes. When 40 square inches of the impregnated strip is extracted 
with 15 ml water, the strips exhibit substantially the same pH as that of 
the buffer solution. Upon contact with various solutions containing from 0 
to 1000 mg percent of protein, the strips prepared from the test 
composition are capable of accurately detecting the quantity of protein 
present to a lower concentration limit of from about 5-10 mg percent. The 
hue change is from yellow or a slight off-yellow to blue with various 
intermediate hues depending on the amount of protein present in the 
solution. 
______________________________________ 
Amount 
______________________________________ 
Sodium citrate (g) 10 
Citric Acid (g) 14 
Water (ml) 140 
Tetrabromphenol Blue (mg) 
48.5 
Ethanol (95%) (ml) 48.5 
______________________________________ 
______________________________________ 
Amount of protein in 
solution (mg.-percent): 
Hue of strip 
______________________________________ 
0 Pyrethrum yellow (51) 
5-20 Apple green (57) 
30 Variscate green (66) 
100 Turquoise green (72) 
300 Turquoise blue (75) 
1000+ Blue turquoise (78) 
______________________________________ 
The Examples 3 to 7 moisturizer composition are applied to different test 
subjects. The test devices for glucose and urea described in Examples 1 
and 2 and the test device for protein set out above are wetted with water 
and contacted with skin treated with appropriate moisturizer compositions. 
For example, the protein (albumin) test device is used for testing 
subjects treated with Examples 4, 5, 6 and 7 and Control compositions, the 
glucose device is used for testing subjects treated with Examples 3, 5 and 
7 compositions and the urea device is used for testing subjects treated 
with Examples 3, 6 and 7 compositions. 
The results obtained from the Examples 4, 5, 6 and 7 subjects are that 
after 1 minute, the amount of protein is shown as 0 for the Control A 
device and satisfactory for the Examples 4, 5, 6 and 7 subjects since the 
Examples 4, 5, 6 and 7 test strips turned blue turquoise. 
The glucose test strips for the Examples 3, 5, and 7 subjects turn brown 
and the urea test strips for the Examples 3, 6 and 7 subjects turn 
characteristic color for urea.