http://informahealthcare.com/phb ISSN 1388-0209 print/ISSN 1744-5116 online Editor-in-Chief: John M. Pezzuto Pharm Biol, 2014; 52(10): 1362–1365 ! 2014 Informa Healthcare USA, Inc. DOI: 10.3109/13880209.2014.884608
SHORT COMMUNICATION
Topically applied glycyrrhizic acid causes hair removal in rats Julijana Ivosevic-Zaper1*, Matthias Hofmann1*, Ayna Kakadjanova1, Eva Valesky1, Markus Meissner1, Ju¨rgen Bereiter-Hahn2, Roland Kaufmann1, August Bernd1, and Stefan Kippenberger1 1
Department of Dermatology, Venereology and Allergology, Johann Wolfgang Goethe University, Frankfurt/Main, Germany and 2Kinematic Cell Research Group, Johann Wolfgang Goethe University, Frankfurt/Main, Germany Abstract
Keywords
Context: Anecdotic reports from Turkmenistan suggest an epilatory effect of sweet licorice extract after topical application. Objective: This study examines hair removal after topical application of glycyrrhizic acid, the main compound of sweet licorice. Materials and methods: An aqueous solution containing 15% of the ammonium salt of glycyrrhizic acid, 10% urea, and 20% ethanol was topically applied two times per day on the neck areas of Wistar rats using a toothbrush. Results: After 3 d, 20–30% of the treated areas were free of hair. After treatment for 6–12 d, 90–95% of the hair was gone. Clinical as well as immunohistological examinations showed no signs of inflammation even after long-term treatment for more than 9 months. Interestingly, long-term treatment reduced the regrowth of hair of about 20%. Examination by scanning electron microscopy showed a smoothed hair cuticle that might facilitate detachment of the hair shaft from the follicular wall. Discussion and conclusion: Our findings suggest glycyrrhizic acid as an interesting molecule for treating hypertrichosis in humans.
Epilation, hirsutism, licorice
Introduction Particularly in modern societies, most women feel compelled to remove unwanted hair from legs and armpits. Recently, more males also desire to have hairless breasts and legs. Although, the transition between cosmetic and clinical conditions is somehow blurred, there is no doubt that patients with hirsutism or hairy congenital nevi suffer from social stigmatization and require efficient hair removal by physicians. The currently used methods for hair removal are often inefficient, expensive, and associated with adverse effects (Table 1). Hair growth is cyclic comprising periods of active growth (anagen), regression (catagen), and resting (telogen). In humans, the lengths of each period depend from the body site and can differ considerably from an anagen of 2 to 6 years for scalp hair to only few months for arms and thigh hair (Olsen, 1999). In the orchestration of the hair cycle, numerous growth factors, adhesion molecules, cytokines, hormones, neuropeptides and their cognate receptors as well as transcription factors and enzymes are involved (Stenn & Paus, 2001). However, the complete mechanism is still not fully *These authors contribute equally to the paper. Correspondence: Prof. Dr. Stefan Kippenberger, Department of Dermatology, Venereology and Allergology, Johann Wolfgang Goethe University, Theodor-Stern-Kai 7, D-60590 Frankfurt/Main, Germany. Tel: 49 69 6301 7734. Fax: +49 69 6301 6466. E-mail: kippenberger@ em.uni-frankfurt.de
History Received 20 November 2013 Accepted 14 January 2014 Published online 25 August 2014
understood. Despite the growing knowledge about hair metabolism, hair removal is mostly achieved by physical and chemical means. The present study was motivated by anecdotic findings from Turkmenistan describing an epilatory effect of topically applied licorice, an extract from the root of Glycyrrhiza glabra. One of the main compounds of licorice is glycyrrhizic acid, a triterpenoid saponin, responsible for the sweet taste. To date, clinical and experimental studies suggest a plethora of useful pharmacological properties of glycyrrhizic acid including antiinflammatory, antiviral, anticancer and immunomodulatory effects to name just a few (Asl & Hosseinzadeh, 2008). To this, the present study added a yet not known property, namely an epilatory effect after topical application.
Materials and methods The efficacy of glycyrrhizic acid was tested in a rat model. All experiments were approved by the local animal experimentation commission (Regierungspra¨sidium Darmstadt, No. F79/22) and performed in certified laboratories of the School of Medicine in Frankfurt/M. Female Wistar rats (weight 250 g) obtained from The Jackson Laboratory (Bar Harbor, ME), held under humidity- and temperature-controlled conditions were fed a standard diet and had free access to water. The epilatory effect was tested in different formulations containing increasing concentrations of glycyrrhizic acid (4%,
Glycyrrhizic acid causes hair removal
DOI: 10.3109/13880209.2014.884608
1363
Table 1. Methods for hair removal (modified from Shenenberger & Utrecht, 2002). Mode Mechanical
Treatment
Advantages
Disadvantages
Plucking/waxing (epilation)
Inexpensive, slow regrowth Easy to apply, inexpensive Inexpensive Potential permanent removal Long-lasting removal
Pain, folliculitis, pseudofolliculitis, burns, irritation, postinflammatory hyperpigmentation, scarring, time consuming Pseudofolliculitis barbae, skin irritation, fast regrowth, time consuming Skin irritation, contact eczema Pain, edema, erythema, scarring, keloid, pigment changes, not suitable for patients with pacemakers, repeated sessions, expensive Pain, edema, erythema, pigment changes, repeated sessions
Easy to apply
Only for androgen-driven hair growth, takes month to show benefit, adverse effects, long-term treatment required Pain, pseudofolliculitis barbae, acne
Shaving (depilation) Chemical Electrical
Thioglycolate Electrolysis
Photothermolysis
Laser and intense pulsed light (IPL) Antiandrogens and oral contraceptives Eflornithine cream
Hormones and inhibitors
Slow regrowth
Figure 1. Epilation by glycyrrhizic acid. Wistar rats were treated with solvent only (a) or with 15% glycyrrhizic acid for 3 d (b) or 6 d (c), respectively. Of note, the treatment causes distinct epilation without clinical signs of skin irritation. To observe long-term effects of glycyrrhizic acid on hair regrowth, animals were treated until complete loss of hair, taking about 6–12 d, then left untreated for 4 weeks, before starting a new treatment cycle. A treatment for seven cycles with solvent only (d) shows no alterations in the fur. In the presence of glycyrrhizic acid, treatments for five (e) and seven cycles (f) lead to impaired regrowth of new hair. Experiments shown were repeated three times with similar results.
8%, and 15%). At 15% glycyrrhizic acid, the epilatory effect was maximal (data not shown). Higher concentrations of glycyrrhizic acid were not soluble in the solvent used. The epilatory effect of glycyrrhizic acid shown in this study was tested in a formulation containing 15% glycyrrhizic acid ammonium salt (Sigma-Aldrich, Munich, Germany) and 10%
urea dissolved in 20% ethanol. The solution was warmed up to 80 C until all components were solubilized and then chilled in a water bath. The neck area of the animals was twice daily treated with 3 500 mL of the solution applied to a toothbrush. For control, served a similar solution devoid of glycyrrhizic acid. At indicated time intervals, representative
1364
J. Ivosevic-Zaper et al.
Pharm Biol, 2014; 52(10): 1362–1365
Figure 2. Glycyrrhizic acid induces modification of the hair cuticle. The surface structure of regrowing hair, plucked out after a treatment-cycle with glycyrrhizic acid, was examined by scanning electron microscopy. Control hair (a and b) of solvent-treated animals features a well-structured hair cuticle with typical surface scales. In contrast, treated hair (c and d) shows a distinctively smoothed surface.
photos were taken. Each group tested comprised five animals. The experiments were repeated three times. The epilatory effect was visually scored by the examiner and expressed in percent. In order to study the hair surface architecture, hairs of anaesthetized animals were plucked out and observed using a scanning electron microscope (SEM, Hitachi S-4500, Hitachi, Oberhausen, Germany).
Results and discussion The epilatory effect of 15% glycyrrhizic acid solution is shown in Figure 1. Already after treatment for 3 d a visible loss of hair of 20–30% was detected (Figure 1b). After 6 d, 90–95% of the treated area was free of hair (Figure 1c). Animals treated with solvent only showed no loss of hair (Figure 1a). Interestingly, the epilated skin showed no clinical signs of irritation. Correspondingly, histological sections maintained a normal profile (data not shown). To observe long-term effects on hair regeneration, animals were treated cyclically with glycyrrhizic acid and finally left untreated for 4 weeks (details are given in the figure legend). Of note, the regrowth of hair was impaired by 10–20% after 5 and 7 treatment cycles giving rise to a moth-eaten appearance (Figure 1e and f). Control animals were treated for seven cycles with solvent only showed a normal fur (Figure 1d). Interestingly, the treatment had an effect on the hair surface. SEM analysis revealed that the cuticle of regrowing hair in
treated animals is smoother with smaller scales than in the controls (Figure 2). There are a plethora of molecular factors involved in hair growth. In this context, the b-1 integrin serves as a key regulator, a transmembrane adhesion protein expressed on keratinocytes (Kloepper et al., 2008; Piwko-Czuchra et al., 2009). In mutant mice deficient for b-1 integrin, the hair follicle morphology is abnormal showing progressive hair loss (Brakebusch et al., 2000). It is up to future studies to investigate if glycyrrhizic acid is involved in the b-1 integrin metabolism. Moreover, b-1 integrin knock-out mice also show severe skin reactions such as blistering and disorganization of the interfollicular epidermis (Brakebusch et al., 2000) which did not occur in our experiments. It might be possible that the penetration route of topically applied glycyrrhizic acid preferably targets hair follicular cells leaving other skin cells unaffected. Another possible mechanism for the epilating effect might be found in the chemical structure of glycyrrhizic acid sharing some similarity with sex steroids. Experiments with prostate cancer cells showed that glycyrrhetinic acid, a metabolite of glycyrrhizic acid, offers a growth-inhibiting effect only in cells with an androgen-dependent phenotype (Hawthorne & Gallagher, 2008). Therefore, it can be speculated about a hormone-like effect of glycyrrhizic acid on hair follicle cells which are known to be sensitive to sex steroids (Azzi et al., 2005). Moreover, the thinning of regrowing hair might be due to an impaired adhesion between smoothed hair cuticle (Figure 2) and inner root sheath cells of
DOI: 10.3109/13880209.2014.884608
the follicular canal facilitating hair loss. Thus, further studies are needed to address the aforementioned aspects. For practical reasons, the experiments presented were performed in rats who, like other rodents, have a synchronized hair follicle cycle (Muller-Rover et al., 2001) allowing fast and reproducible assessment of epilatory compounds. It will be interesting to see whether glycyrrhizic acid offers similar effects on human hair growth. In case of positive results, this compound has the potential to amend the arsenal of methods for hair removal. Of particular interest is the absence of skin irritations which are present in most established methods (Table 1).
Declaration of interest The authors have no conflict of interest.
References Asl MN, Hosseinzadeh H. (2008). Review of pharmacological effects of Glycyrrhiza sp. and its bioactive compounds. Phytother Res 22: 709–24.
Glycyrrhizic acid causes hair removal
1365
Azzi L, El-Alfy M, Martel C, Labrie F. (2005). Gender differences in mouse skin morphology and specific effects of sex steroids and dehydroepiandrosterone. J Invest Dermatol 124:22–7. Brakebusch C, Grose R, Quondamamatteo F, et al. (2000). Skin and hair follicle integrity is crucially dependent on beta 1 integrin expression on keratinocytes. EMBO J 19:3990–4003. Hawthorne S, Gallagher S. (2008). Effects of glycyrrhetinic acid and liquorice extract on cell proliferation and prostate-specific antigen secretion in LNCaP prostate cancer cells. J Pharm Pharmacol 60: 661–6. Kloepper JE, Hendrix S, Bodo E, et al. (2008). Functional role of beta 1 integrin-mediated signalling in the human hair follicle. Exp Cell Res 314:498–508. Muller-Rover S, Handjiski B, van der Veen C, et al. (2001). A comprehensive guide for the accurate classification of murine hair follicles in distinct hair cycle stages. J Invest Dermatol 117:3–15. Olsen EA. (1999). Methods of hair removal. J Am Acad Dermatol 40: 143–55; quiz 156–7. Piwko-Czuchra A, Koegel H, Meyer H, et al. (2009). Beta 1 integrinmediated adhesion signalling is essential for epidermal progenitor cell expansion. PLoS One 4:e5488. Shenenberger DW, Utrecht LM. (2002). Removal of unwanted facial hair. Am Fam Physician 66:1907–11. Stenn KS, Paus R. (2001). Controls of hair follicle cycling. Physiol Rev 81:449–94.
Copyright of Pharmaceutical Biology is the property of Taylor & Francis Ltd and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use.
SHORT COMMUNICATION
Topically applied glycyrrhizic acid causes hair removal in rats Julijana Ivosevic-Zaper1*, Matthias Hofmann1*, Ayna Kakadjanova1, Eva Valesky1, Markus Meissner1, Ju¨rgen Bereiter-Hahn2, Roland Kaufmann1, August Bernd1, and Stefan Kippenberger1 1
Department of Dermatology, Venereology and Allergology, Johann Wolfgang Goethe University, Frankfurt/Main, Germany and 2Kinematic Cell Research Group, Johann Wolfgang Goethe University, Frankfurt/Main, Germany Abstract
Keywords
Context: Anecdotic reports from Turkmenistan suggest an epilatory effect of sweet licorice extract after topical application. Objective: This study examines hair removal after topical application of glycyrrhizic acid, the main compound of sweet licorice. Materials and methods: An aqueous solution containing 15% of the ammonium salt of glycyrrhizic acid, 10% urea, and 20% ethanol was topically applied two times per day on the neck areas of Wistar rats using a toothbrush. Results: After 3 d, 20–30% of the treated areas were free of hair. After treatment for 6–12 d, 90–95% of the hair was gone. Clinical as well as immunohistological examinations showed no signs of inflammation even after long-term treatment for more than 9 months. Interestingly, long-term treatment reduced the regrowth of hair of about 20%. Examination by scanning electron microscopy showed a smoothed hair cuticle that might facilitate detachment of the hair shaft from the follicular wall. Discussion and conclusion: Our findings suggest glycyrrhizic acid as an interesting molecule for treating hypertrichosis in humans.
Epilation, hirsutism, licorice
Introduction Particularly in modern societies, most women feel compelled to remove unwanted hair from legs and armpits. Recently, more males also desire to have hairless breasts and legs. Although, the transition between cosmetic and clinical conditions is somehow blurred, there is no doubt that patients with hirsutism or hairy congenital nevi suffer from social stigmatization and require efficient hair removal by physicians. The currently used methods for hair removal are often inefficient, expensive, and associated with adverse effects (Table 1). Hair growth is cyclic comprising periods of active growth (anagen), regression (catagen), and resting (telogen). In humans, the lengths of each period depend from the body site and can differ considerably from an anagen of 2 to 6 years for scalp hair to only few months for arms and thigh hair (Olsen, 1999). In the orchestration of the hair cycle, numerous growth factors, adhesion molecules, cytokines, hormones, neuropeptides and their cognate receptors as well as transcription factors and enzymes are involved (Stenn & Paus, 2001). However, the complete mechanism is still not fully *These authors contribute equally to the paper. Correspondence: Prof. Dr. Stefan Kippenberger, Department of Dermatology, Venereology and Allergology, Johann Wolfgang Goethe University, Theodor-Stern-Kai 7, D-60590 Frankfurt/Main, Germany. Tel: 49 69 6301 7734. Fax: +49 69 6301 6466. E-mail: kippenberger@ em.uni-frankfurt.de
History Received 20 November 2013 Accepted 14 January 2014 Published online 25 August 2014
understood. Despite the growing knowledge about hair metabolism, hair removal is mostly achieved by physical and chemical means. The present study was motivated by anecdotic findings from Turkmenistan describing an epilatory effect of topically applied licorice, an extract from the root of Glycyrrhiza glabra. One of the main compounds of licorice is glycyrrhizic acid, a triterpenoid saponin, responsible for the sweet taste. To date, clinical and experimental studies suggest a plethora of useful pharmacological properties of glycyrrhizic acid including antiinflammatory, antiviral, anticancer and immunomodulatory effects to name just a few (Asl & Hosseinzadeh, 2008). To this, the present study added a yet not known property, namely an epilatory effect after topical application.
Materials and methods The efficacy of glycyrrhizic acid was tested in a rat model. All experiments were approved by the local animal experimentation commission (Regierungspra¨sidium Darmstadt, No. F79/22) and performed in certified laboratories of the School of Medicine in Frankfurt/M. Female Wistar rats (weight 250 g) obtained from The Jackson Laboratory (Bar Harbor, ME), held under humidity- and temperature-controlled conditions were fed a standard diet and had free access to water. The epilatory effect was tested in different formulations containing increasing concentrations of glycyrrhizic acid (4%,
Glycyrrhizic acid causes hair removal
DOI: 10.3109/13880209.2014.884608
1363
Table 1. Methods for hair removal (modified from Shenenberger & Utrecht, 2002). Mode Mechanical
Treatment
Advantages
Disadvantages
Plucking/waxing (epilation)
Inexpensive, slow regrowth Easy to apply, inexpensive Inexpensive Potential permanent removal Long-lasting removal
Pain, folliculitis, pseudofolliculitis, burns, irritation, postinflammatory hyperpigmentation, scarring, time consuming Pseudofolliculitis barbae, skin irritation, fast regrowth, time consuming Skin irritation, contact eczema Pain, edema, erythema, scarring, keloid, pigment changes, not suitable for patients with pacemakers, repeated sessions, expensive Pain, edema, erythema, pigment changes, repeated sessions
Easy to apply
Only for androgen-driven hair growth, takes month to show benefit, adverse effects, long-term treatment required Pain, pseudofolliculitis barbae, acne
Shaving (depilation) Chemical Electrical
Thioglycolate Electrolysis
Photothermolysis
Laser and intense pulsed light (IPL) Antiandrogens and oral contraceptives Eflornithine cream
Hormones and inhibitors
Slow regrowth
Figure 1. Epilation by glycyrrhizic acid. Wistar rats were treated with solvent only (a) or with 15% glycyrrhizic acid for 3 d (b) or 6 d (c), respectively. Of note, the treatment causes distinct epilation without clinical signs of skin irritation. To observe long-term effects of glycyrrhizic acid on hair regrowth, animals were treated until complete loss of hair, taking about 6–12 d, then left untreated for 4 weeks, before starting a new treatment cycle. A treatment for seven cycles with solvent only (d) shows no alterations in the fur. In the presence of glycyrrhizic acid, treatments for five (e) and seven cycles (f) lead to impaired regrowth of new hair. Experiments shown were repeated three times with similar results.
8%, and 15%). At 15% glycyrrhizic acid, the epilatory effect was maximal (data not shown). Higher concentrations of glycyrrhizic acid were not soluble in the solvent used. The epilatory effect of glycyrrhizic acid shown in this study was tested in a formulation containing 15% glycyrrhizic acid ammonium salt (Sigma-Aldrich, Munich, Germany) and 10%
urea dissolved in 20% ethanol. The solution was warmed up to 80 C until all components were solubilized and then chilled in a water bath. The neck area of the animals was twice daily treated with 3 500 mL of the solution applied to a toothbrush. For control, served a similar solution devoid of glycyrrhizic acid. At indicated time intervals, representative
1364
J. Ivosevic-Zaper et al.
Pharm Biol, 2014; 52(10): 1362–1365
Figure 2. Glycyrrhizic acid induces modification of the hair cuticle. The surface structure of regrowing hair, plucked out after a treatment-cycle with glycyrrhizic acid, was examined by scanning electron microscopy. Control hair (a and b) of solvent-treated animals features a well-structured hair cuticle with typical surface scales. In contrast, treated hair (c and d) shows a distinctively smoothed surface.
photos were taken. Each group tested comprised five animals. The experiments were repeated three times. The epilatory effect was visually scored by the examiner and expressed in percent. In order to study the hair surface architecture, hairs of anaesthetized animals were plucked out and observed using a scanning electron microscope (SEM, Hitachi S-4500, Hitachi, Oberhausen, Germany).
Results and discussion The epilatory effect of 15% glycyrrhizic acid solution is shown in Figure 1. Already after treatment for 3 d a visible loss of hair of 20–30% was detected (Figure 1b). After 6 d, 90–95% of the treated area was free of hair (Figure 1c). Animals treated with solvent only showed no loss of hair (Figure 1a). Interestingly, the epilated skin showed no clinical signs of irritation. Correspondingly, histological sections maintained a normal profile (data not shown). To observe long-term effects on hair regeneration, animals were treated cyclically with glycyrrhizic acid and finally left untreated for 4 weeks (details are given in the figure legend). Of note, the regrowth of hair was impaired by 10–20% after 5 and 7 treatment cycles giving rise to a moth-eaten appearance (Figure 1e and f). Control animals were treated for seven cycles with solvent only showed a normal fur (Figure 1d). Interestingly, the treatment had an effect on the hair surface. SEM analysis revealed that the cuticle of regrowing hair in
treated animals is smoother with smaller scales than in the controls (Figure 2). There are a plethora of molecular factors involved in hair growth. In this context, the b-1 integrin serves as a key regulator, a transmembrane adhesion protein expressed on keratinocytes (Kloepper et al., 2008; Piwko-Czuchra et al., 2009). In mutant mice deficient for b-1 integrin, the hair follicle morphology is abnormal showing progressive hair loss (Brakebusch et al., 2000). It is up to future studies to investigate if glycyrrhizic acid is involved in the b-1 integrin metabolism. Moreover, b-1 integrin knock-out mice also show severe skin reactions such as blistering and disorganization of the interfollicular epidermis (Brakebusch et al., 2000) which did not occur in our experiments. It might be possible that the penetration route of topically applied glycyrrhizic acid preferably targets hair follicular cells leaving other skin cells unaffected. Another possible mechanism for the epilating effect might be found in the chemical structure of glycyrrhizic acid sharing some similarity with sex steroids. Experiments with prostate cancer cells showed that glycyrrhetinic acid, a metabolite of glycyrrhizic acid, offers a growth-inhibiting effect only in cells with an androgen-dependent phenotype (Hawthorne & Gallagher, 2008). Therefore, it can be speculated about a hormone-like effect of glycyrrhizic acid on hair follicle cells which are known to be sensitive to sex steroids (Azzi et al., 2005). Moreover, the thinning of regrowing hair might be due to an impaired adhesion between smoothed hair cuticle (Figure 2) and inner root sheath cells of
DOI: 10.3109/13880209.2014.884608
the follicular canal facilitating hair loss. Thus, further studies are needed to address the aforementioned aspects. For practical reasons, the experiments presented were performed in rats who, like other rodents, have a synchronized hair follicle cycle (Muller-Rover et al., 2001) allowing fast and reproducible assessment of epilatory compounds. It will be interesting to see whether glycyrrhizic acid offers similar effects on human hair growth. In case of positive results, this compound has the potential to amend the arsenal of methods for hair removal. Of particular interest is the absence of skin irritations which are present in most established methods (Table 1).
Declaration of interest The authors have no conflict of interest.
References Asl MN, Hosseinzadeh H. (2008). Review of pharmacological effects of Glycyrrhiza sp. and its bioactive compounds. Phytother Res 22: 709–24.
Glycyrrhizic acid causes hair removal
1365
Azzi L, El-Alfy M, Martel C, Labrie F. (2005). Gender differences in mouse skin morphology and specific effects of sex steroids and dehydroepiandrosterone. J Invest Dermatol 124:22–7. Brakebusch C, Grose R, Quondamamatteo F, et al. (2000). Skin and hair follicle integrity is crucially dependent on beta 1 integrin expression on keratinocytes. EMBO J 19:3990–4003. Hawthorne S, Gallagher S. (2008). Effects of glycyrrhetinic acid and liquorice extract on cell proliferation and prostate-specific antigen secretion in LNCaP prostate cancer cells. J Pharm Pharmacol 60: 661–6. Kloepper JE, Hendrix S, Bodo E, et al. (2008). Functional role of beta 1 integrin-mediated signalling in the human hair follicle. Exp Cell Res 314:498–508. Muller-Rover S, Handjiski B, van der Veen C, et al. (2001). A comprehensive guide for the accurate classification of murine hair follicles in distinct hair cycle stages. J Invest Dermatol 117:3–15. Olsen EA. (1999). Methods of hair removal. J Am Acad Dermatol 40: 143–55; quiz 156–7. Piwko-Czuchra A, Koegel H, Meyer H, et al. (2009). Beta 1 integrinmediated adhesion signalling is essential for epidermal progenitor cell expansion. PLoS One 4:e5488. Shenenberger DW, Utrecht LM. (2002). Removal of unwanted facial hair. Am Fam Physician 66:1907–11. Stenn KS, Paus R. (2001). Controls of hair follicle cycling. Physiol Rev 81:449–94.
Copyright of Pharmaceutical Biology is the property of Taylor & Francis Ltd and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use.
