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Amphoteric Complexes Offer Unique Benefits to Alpha
Hydroxyacid (AHA) Skin Care Formulations

Barbara A. Green, R.Ph., Richard H. Wildnauer, Ph.D.,
Robert C. Hwu, Ph.D., David J. Milora, Brenda L. Edison

NeoStrata Company, Inc., Princeton, NJ, USA.
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Introduction
The benefits of alpha hydroxyacids (AHAs) for skin care and therapy are numerous and well documented in the scientific literature.1-5  Despite their considerable benefits, topical use of AHAs can be limited due to stinging and irritant reactions with these agents on sensitive skin.  AHAs occasionally cause objective irritation responses including erythema and swelling, but more frequently cause untoward sensory responses such as stinging, itching, and burning.  Objective irritation responses are a cause for concern and may warrant discontinuation of a product. Negative sensory responses are generally transient and not a cause for alarm, but often result in unnecessary product discontinuation, or poor compliance.

New polyhydroxy acid (PHA) ingredients and novel formulation technologies are being found to reduce the potential irritation of AHAs with minimal or no negative sensory responses, while maintaining clinical efficacy.6,7   One notable technology utilizes naturally occurring amino acids to modulate AHA delivery and irritation profile.  The skin care benefits provided by the addition of amino acids are related to the amphoteric nature of amino acids, which facilitates the formation of a molecular complex with alpha hydroxyacids.8,9  This complex is purported to provide a slow release of the free alpha hydroxyacid without significantly altering the pH, thereby allowing a slower, more gentle penetration of the AHA without stinging and burning, and with a reduced potential for irritation.  Clinical evaluations indicate that significant benefits are achieved with amphoteric AHA systems in comparison to equivalent non-amphoteric AHA systems including a reduction of perceived stinging and a reduced potential for irritation under exaggerated patch test conditions.9-11  Most important, clinical efficacy and desirable formulation aesthetics are maintained with amphoteric AHA formulations.
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Objective
The purpose of this poster is to describe amphoteric hydroxyacid technology and demonstrate the clinical benefits thereof including:
  • Less stinging than non-amphoteric AHA formulations
  • Less irritation than non-amphoteric AHA formulations
  • Equivalent efficacy to similar non-amphoteric AHA formulations
  • Favorable formulation aesthetics utilizing amphoteric technology
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Amphoteric Hydroxyacids
Clinical effectiveness of topical AHA formulations depends on skin penetration of the AHA, which is determined by the bioavailable concentration of the AHA in an optimum vehicle.12  In order to maximize AHA bioavailability, formulation pH must be acidic. When formulated at pH 3.8 (the pKa of glycolic acid), approximately 50% of the amount of glycolic acid (an AHA) would be present in the non-ionized, free-acid form, which is readily bioavailable.12  Rapid penetration of glycolic acid is often associated with stinging and burning.

Most commercial AHA formulations incorporate alkali and ammonium salts as neutralizing agents to adjust pH.  These agents form ionic bonds with the AHA and therefore inhibit penetration into the skin of the neutralized acid; the unneutralized acid remains free to penetrate. Amphoteric hydroxyacid technology utilizes an amphoteric amino acid as part of the neutralizing system, which not only helps adjust formulation pH, but apparently attracts the 'free' glycolic acid, forming temporary complexes to slow the penetration of the free acid. This seemingly prevents sensory receptor threshold activation and thereby reduces stinging and burning.  Since the complex is temporary due to the nature of the bonds, formulation efficacy is not compromised.
Glycolic Acid / Arginine Complex
AHA Amphoteric Intermolecular Forces
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Molecular Complex and pH Profile
Laboratory investigations utilizing Resistance (ohms) as a quantitative measure of ionic species indicate that amphoteric AHA solutions may contain fewer ionic species than a comparable solution neutralized with ammonium hydroxide, thus providing support for the formation of AHA amphoteric complexes.  Since resistance to conduct electric current is inversely proportional to ion concentration, the fewer the number of ions present in solution the higher the resistance.  Solutions utilizing arginine as the pH adjuster have a greater resistance than otherwise equivalent solutions partially neutralized with ammonium hydroxide.

Experiment

  • Equivalent aqueous glycolic acid (50%) solutions were adjusted to pH using either the amphoteric agent arginine or ammonium hydroxide. Resistance was measured using the Accumet pH / Conductivity meter (model 20, Fisher Scientific) equipped with a two-cell conductivity glass probe.

Results

  • Glycolic acid solutions containing arginine are more resistive to current than solutions containing ammonium hydroxide indicating reduced ion content at equivalent pH.
  • Data indicate that there are fewer ions present in the solutions containing arginine in comparison to identical solutions containing ammonium hydroxide, thus indicating the presence of a molecular complex in the arginine containing solutions.
  • Viscosity increases in arginine containing solutions beginning at a 10:1 molar ratio of glycolic acid: arginine.  There is a corresponding increase in resistance due to diminished flow properties.
Resistance of 50% Glycolic Acid Solutions at Variable pH*
* Solutions were adjusted to pH using either arginine
(amphoteric) or Ammonium Hydroxide (NH40H)
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Viscosity of 50% Glycolic Acid / Arginine Solutions at Various Mole Ratios
Mole Ratio = Moles Glycolic Acid / Mole Arginine
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Reduced Stinging
Many AHA product users experience stinging and burning following topical application of lactic acid and glycolic acid. In an attempt to overcome these negative sensory responses and continue to provide the benefits of AHAs, some formulation approaches have been attempted with varying degrees of success.13  The following experiment demonstrates the effectiveness of amphoteric technology in diminishing the stinging response from lactic acid.

Experiment

  • Method: lactic acid sting test
  • Panel of 21 females identified as 'stingers' screened with 10% unneutralized lactic acid
  • Application of test materials to nasolabial fold and cheek area using cotton swab
  • Test materials: solutions of lactic acid (20%, aq.) partially neutralized to pH 3.5 using (1) arginine (amphoteric), (2) Ammonium Hydroxide, (3) NaOH (alkali)
  • Negative control: water

Results

  • Solutions containing arginine induced significantly less stinging than those containing ammonium hydroxide, p<0.05.
  • Solutions containing arginine induced significantly less stinging than those containing sodium hydroxide, p<0.05.
  • Arginine containing solutions were equivalent to the negative control solution (water)
Facial Sting Test
20% Lactic Acid Solutions, pH 3.5
pH adjusted with ammonium hydroxide or arginine
 *Statistical Significance p<0.05
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Facial Sting Test
20% Lactic Acid Solutions, pH 3.5
pH adjusted with sodium hydroxide or arginine
* Statistical Significance p<0.05.
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Facial Sting Test
20% Lactic Acid Solutions, pH 3.5
pH adjusted with arginine
No Statistical Differences.
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Reduced Irritation
Topical formulations are routinely evaluated for irritation potential using patch test models that are exaggerated beyond normal use in order to approximate long term safety in a short period of time.  To demonstrate the benefit of amphoteric AHA technology, a cumulative irritation patch test was conducted in comparison to non-amphoteric AHA solutions.

Experiment

  • Method: 14 day cumulative irritation test
  • Panel of 24 healthy volunteers
  • Products were applied 14 consecutive days including weekends with occlusive patches
  • Test materials: aqueous solutions of 20% glycolic acid partially neutralized to pH 3.5 using (1) arginine (amphoteric), (2) Ammonium Hydroxide, (3) NaOH (alkali)
  • Controls: 0.9% NaCl (negative control) and 0.1% SLS (positive control)

Results

  • Glycolic acid partially neutralized with arginine is significantly less irritating than glycolic acid solutions partially neutralized with either sodium hydroxide or ammonium hydroxide, p<0.05.
  • Moderate levels of irritation (grade 2) occurred after 4 days of patching in the ammonium hydroxide treatment group compared to nearly 11 days in the arginine treatment group.
  • Equivalent levels of irritation are eventually reached toward the end of the study as the glycolic acid continues to penetrate from all test solutions under the severe conditions of occlusion.  This indicates that while the amphoteric system reduces irritation under occlusion for an extended initial period, there is sustained penetration of the glycolic acid and a slower emergence of irritation in comparison to non-amphoteric AHA solutions. This provides support that bioavailability is not compromised by the amphoteric AHA technology.
Cumulative Irritation Study
20% Glycolic Acid, pH 3.5
pH adjusted with ammonium hydroxide, sodium hydroxide or arginine
Statistical differences between AHA solutions, p<0.05:
NH40H vs. Arginine: days 2-11 and overall;
NaOH vs. Arginine: days 2-6, 8-10 and overall
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Cumulative Irritation Study
20% Glycolic Acid Solutions, pH 3.5
pH adjusted with ammonium hydroxide or arginine
4 days to moderate irritation (grade 2) vs. 11 days
Statistical differences, p<0.05. NH40H vs. Arginine: days 2-11 and overall
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Cell Turnover
Although AHA product benefits extend well beyond stratum corneum exfoliation and enhancement of cell turnover, AHA product effectiveness can be quickly screened using the dansyl chloride cell turnover model.  In this assessment, an amphoteric AHA prototype cream containing 6.3% glycolic acid adjusted to pH 3.5 with an arginine containing neutralizing system (molar ratio 15:1, glycolic acid: arginine) was compared to an untreated control to evaluate enhancement of cell turnover and exfoliation.

Experiment

  • Method: dansyl chloride cell turnover
  • Panel of 26 healthy volunteers
  • Product application twice daily to the inner forearms
  • Test materials: amphoteric AHA cream (pH 3.5) containing 6.3% glycolic acid with arginine (molar ratio 15:1, glycolic acid: arginine). The test cream was compared to an untreated control.

Results

  • The amphoteric AHA cream significantly reduced mean stratum corneum turnover time by 15.7% compared to the untreated control, p<0.01.
  • The amphoteric AHA cream significantly reduced mean total fluorescence scores by 18.4% compared to the untreated control, p<0.01.
  • Data demonstrates that amphoteric technology provides cell turnover benefits consistent with non-amphoteric AHA formulations.
Dansyl Chloride Cell Turnover
Amphoteric AHA Cream vs. Untreated
Statistical differences: Prototype amphoteric AHA reduced mean stratum corneum
turnover time and total fluorescence scores compared to untreated, p<0.01.
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Aesthetically Appealing Formulations
Compliance with skin care regimens is largely influenced by product aesthetics and ease of use. In order to ensure that amphoteric AHA technology does not negatively affect formulation attributes, products are routinely compared to benchmark leaders to assure optimum aesthetics. The following product evaluation screen indicates that amphoteric AHA technology does not negatively affect perceived product aesthetics.

Experiment

  • Method: direct comparison, self-assessment
  • Panel of 10 volunteers
  • Test materials: amphoteric AHA cream (pH 3.5), 6.3% glycolic acid with arginine (molar ratio 15:1, glycolic acid: arginine). The test cream was compared to the NeoStrata Skin Smoothing Cream with 8% glycolic acid and two competitive AHA benchmarks.

Results

  • Formulation aesthetics of the amphoteric AHA cream were rated favorably in comparison to the non-amphoteric AHA benchmarks indicating that amphoteric technology does not compromise AHA formulation aesthetics.
Aesthetic Comparison Test
5=excellent, 4=very good, 3=good, 2=fair, 1=poor
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Conclusion
Amphoteric AHA technology provides unique benefits to AHA skin care through its impact on product mildness characteristics.  This poster demonstrates the important benefits of amphoteric AHA technology in reducing AHA stinging potential and irritation, while maintaining efficacy and desirable formulation aesthetics.  As a result, clinically beneficial AHA products may be used by all skin types including sensitive skin through the incorporation of amphoteric agents in the formulation.
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References
  1. Van Scott EJ, Yu RJ. Hyperkeratinization, corneocyte cohesion and alpha hydroxy acids. J Am Acad Dermatol 1984; 11:867-879.
  2. Van Scott EJ, Yu RJ. Substances that modify the stratum corneum by modulating its formation. In: Frost, P. and Horwitz, S., eds. Principles of Cosmetics for the Dermatologist. St. Louis: C.V. Mosby 1982; 70-74.
  3. Yu RJ, Van Scott EJ. Alpha-hydroxy acids: science and therapeutic uses. Cosmetic Dermatology Supplement 1994:1-6.
  4. Ditre CM, Griffin TD, Murphy GF, Sueki H, Telegan B, Johnson WC, Yu RJ, Van Scott EJ. Effects of a-hydroxy acids on photoaged skin: A pilot clinical, histologic, and ultrastructural study. J Am Acad Dermatol 1996:34:187-95.
  5. Bernstein EF, Underhill CB, Lakkakorpi J, Ditre CM, Uitto J, Yu RJ, Van Scott EJ. Citric Acid increases viable epidermal thickness & glycosaminoglycan content of sun-damaged skin. Dermatol Surg 1997; 23: 689-94.
  6. Berardesca E, Distante F, Vignoli GP, Oresajo C, Green B. Alpha hydroxy acids modulate stratum corneum barrier function. Br. J. Dermatol 1997;137:934-8.
  7. Green B, Tseng C, Wildnauer R, Herndon J, Rizer, R. Safety and Efficacy of a Gluconolactone (Poly Hydroxyacid) Containing Regimen on Sensitive Skin and Photodamage Following Controlled Consumer Use. Amer Acad of Derm Poster Exhibit: New Orleans, March, 1999.
  8. US Pat 5,091,171, Amphoteric composition and polymeric forms of alpha hydroxy acids and their therapeutic use. Yu RJ, Van Scott EJ (1992).
  9. Kraechter HU, McCaulley JA, Edison B, Green B, Milora DJ. Amphoteric Hydroxy Complexes: AHAs with reduced stinging and irritation. Cosmetics & Toiletries 2001;116(1):47-52.
  10. Morganti P, Randazzo SD, Fabrizi G, Bruno C. Decreasing the stinging capacity and improving the antiaging activity of AHAs. J. Appl Cosmetol 1996;14:79-91.
  11. Tucci MG, Belmonte MM, Biagini G, Morganti P, Vellucci E, Talassi O, Solmi R, Ricotti G. AHAs and derivatives. Cosmetics & Toiletries 1998;113:55-58.
  12. Yu RJ, Van Scott EJ: Bioavailability of alpha-hydroxy acids in topical formulations. Cosmet Dermatol 1996;9(6):54-62.
  13. Kligman DE, Pagnoni A, Stoudemayer T, Kligman AM. Strontium Nitrate Decreases the Efficacy of Glycolic Acid Peel. Amer Acad of Derm Poster Exhibit: San Francisco, March, 2000.
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Acknowledgements
Sting testing was conducted at Consumer Product Testing Company, Inc., Fairfield, NJ. The cumulative irritation test was conducted at Hill Top Research, Inc., East Brunswick, NJ.
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Products Using the Amphoteric System
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