Grandmaternal smoke exposure reduces female fertility in a murine model, with great-grandmaternal smoke exposure unlikely to have an effect.

STUDY QUESTION: What effect does multigenerational (F2) and transgenerational (F3) cigarette smoke exposure have on female fertility in mice? SUMMARY ANSWER: Cigarette smoking has a multigenerational effect on female fertility. WHAT IS KNOWN ALREADY: It has been well established that cigarette smoking decreases female fertility. Furthermore, a growing body of evidence suggests that smoking during pregnancy decreases the fertility of daughters and increases cancer and asthma incidence in grandchildren and great-grandchildren. STUDY DESIGN, SIZE, DURATION: Six-week-old C57BL/6 female mice were exposed nasally to cigarette smoke or room air (controls) for 5 weeks prior to being housed with males. Females continued to be exposed to smoke throughout pregnancy and lactation until pups were weaned. A subset of F1 female pups born to these smoke and non-smoke exposed females were bred to create the F2 grandmaternal exposed generation (multigenerational). Finally, a subset of F2 females were bred to create the F3 great-grandmaternal exposed generation (transgenerational). The reproductive health of F2 and F3 females was examined at 8 weeks and 9 months. PARTICIPANTS/MATERIALS, SETTING, METHODS: Ovarian and oocyte quality was examined in smoke exposed and control animals. A small-scale fertility trial was performed before ovarian changes were examined using ovarian histology and immunofluorescence and/or immunoblotting analysis of markers of apoptosis (TUNEL) and proliferation (proliferating cell nuclear antigen (PCNA) and anti-Mullerian hormone (AMH)). Oocyte quality was examined using immunocytochemistry to analyze the metaphase II spindle and ploidy status. Parthenogenetic activation of oocytes was used to investigate meiosis II timing and preimplantation embryo development. Finally, diestrus hormone serum levels (FSH and LH) were quantified. MAIN RESULTS AND THE ROLE OF CHANCE: F2 smoke exposed females had no detectable change in ovarian follicle quality at 8 weeks, although by 9 months ovarian somatic cell proliferation was reduced (P = 0.0197) compared with non-smoke exposed control. Further investigation revealed changes between control and smoke exposed F2 oocyte quality, including altered meiosis II timing at 8 weeks (P = 0.0337) and decreased spindle pole to pole length at 9 months (P = 0.0109). However, no change in preimplantation embryo development was observed following parthenogenetic activation. The most noticeable effect of cigarette smoke exposure was related to the subfertility of F2 females; F2 smoke exposed females displayed significantly increased time to conception (P = 0.0042) and significantly increased lag time between pregnancies (P = 0.0274) compared with non-smoke exposed F2 females. Conversely, F3 smoke exposed females displayed negligible oocyte and follicle changes up to 9 months of age, and normal preimplantation embryo development. LARGE SCALE DATA: None. LIMITATIONS, REASONS FOR CAUTION: This study focused solely on a mouse model of cigarette smoke exposure to simulate human exposure. WIDER IMPLICATIONS OF THE FINDINGS: Our results demonstrate that grandmaternal cigarette smoke exposure reduces female fertility in mice, highlighting the clinical need to promote cessation of cigarette smoking in pregnant women. STUDY FUNDING/COMPETING INTEREST(S): This study was funded by the Australian Research Council, National Health and Medical Research Council, Hunter Medical Research Institute, Newcastle Permanent Building Society Charitable Trust, and the University of Newcastle Priory Research Centers in Chemical Biology, Healthy Lungs and Grow Up Well. The authors declare no conflict of interest.

Subfertility in androgen-insensitive female mice is rescued by transgenic FSH.

Androgens synergise with FSH in female reproduction but the nature of their interaction in ovarian function and fertility is not clear. In the present study, we investigated this interaction, notably whether higher endogenous FSH can overcome defective androgen actions in androgen receptor (AR)-knockout (ARKO) mice. We generated and investigated the reproductive function of mutant mice exhibiting AR resistance with or without expression of human transgenic FSH (Tg-FSH). On the background of inactivated AR signalling, which alone resulted in irregular oestrous cycles and reduced pups per litter, ovulation rates and antral follicle health, Tg-FSH expression restored follicle health, ovulation rates and litter size to wild-type levels. However, Tg-FSH was only able to partially rectify the abnormal oestrous cycles observed in ARKO females. Hence, elevated endogenous FSH rescued the intraovarian defects, and partially rescued the extraovarian defects due to androgen insensitivity. In addition, the observed increase in litter size in Tg-FSH females was not observed in the presence of AR signalling inactivation. In summary, the findings of the present study reveal that FSH can rescue impaired female fertility and ovarian function due to androgen insensitivity in female ARKO mice by maintaining follicle health and ovulation rates, and thereby optimal female fertility.

Role of androgens in normal and pathological ovarian function.

Androgens mediate their actions via the androgen receptor (AR), a member of the nuclear receptor superfamily. AR-mediated androgen action is essential in male reproductive development and function; however, only in the last decade has the suspected but unproven role for AR-mediated actions in female reproduction been firmly established. Deciphering the specific roles and precise pathways by which AR-mediated actions regulate ovarian function has been hindered by confusion on how to interpret results from pharmacological studies using androgens that can be converted into oestrogens, which exert actions via the oestrogen receptors. The generation and analysis of global and cell-specific female Ar knockout mouse models have deduced a role for AR-mediated actions in regulating ovarian function, maintaining female fertility, and have begun to unravel the mechanisms by which AR-mediated androgen actions regulate follicle health, development and ovulation. Furthermore, observational findings from human studies and animal models provide substantial evidence to support a role for AR-mediated effects not only in normal ovarian function but also in the development of the frequent ovarian pathological disorder, polycystic ovarian syndrome (PCOS). This review focuses on combining the findings from observational studies in humans, pharmacological studies and animal models to reveal the roles of AR-mediated actions in normal and pathological ovarian function. Together these findings will enable us to begin understanding the important roles of AR actions in the regulation of female fertility and ovarian ageing, as well as providing insights into the role of AR actions in the androgen-associated reproductive disorder PCOS.

In vitro human skin absorption of linalool: Effects of vehicle composition, evaporation and occlusion on permeation and distribution.

In vitro human skin permeation and distribution of the fragrance material linalool (3,7-dimethyl-1,6-octadien-3-ol, CAS No. 78-70-6) following application in a range of single and mixed vehicles was determined, under unoccluded and occluded conditions, using human epidermal membranes. Vehicles were (70/30 v/v) ethanol[EtOH]/water, dipropyleneglycol [DPG], diethyl phthalate [DEP], (25/75 v/v) EtOH/DEP, (25/75 v/v) EtOH/DPG and petrolatum. Worst case absorbed dose values (% applied dose) for linalool under unoccluded conditions varied from 1.84% (DPG) to 4.08% (EtOH/water) and under occluded conditions from 5.9% (DEP) to 14.7% (EtOH/water). Occlusion always increased absorption but the magnitude of the effect varied with the vehicle from 2 to 6-fold. This study demonstrated that in vitro human skin permeation of linalool varied quite widely between test vehicles and that the magnitude of the effect of occlusion was also vehicle dependent. This was particularly significant in view of the reported variations in biological responses using different vehicles (Lalko et al., 2004; Politano et al., 2006).

Androgens and ovarian function: translation from basic discovery research to clinical impact.

In the last decade, it has been revealed that androgens play a direct and important role in regulating female reproductive function. Androgens mediate their actions via the androgen receptor (AR), and global and cell-specific Ar-knockout mouse models have confirmed that AR-mediated androgen actions play a role in regulating female fertility and follicle health, development and ovulation. This knowledge, along with the clinical data reporting a beneficial effect of androgens or androgen-modulating agents in augmenting in vitro fertilization (IVF) stimulation in women termed poor responders, has supported the adoption of this concept in many IVF clinics worldwide. On the other hand, substantial evidence from human and animal studies now supports the hypothesis that androgens in excess, acting via the AR, play a key role in the origins of polycystic ovary syndrome (PCOS). The identification of the target sites of these AR actions and the molecular mechanisms involved in underpinning the development of PCOS is essential to provide the knowledge required for the future development of novel, mechanism-based therapies for the treatment of PCOS. This review will summarize the basic scientific discoveries that have enhanced our knowledge of the roles of androgens in female reproductive function, discuss the impact these findings have had in the clinic and how a greater understanding of the role androgens play in female physiology may shape the future development of effective strategies to improve IVF outcomes in poor responders and the amelioration of symptoms in patients with PCOS.

Role of androgens in the ovary.

It has been well established for decades that androgens, namely testosterone (T) plays an important role in female reproductive physiology as the precursor for oestradiol (E2). However, in the last decade a direct role for androgens, acting via the androgen receptor (AR), in female reproductive function has been confirmed. Deciphering the specific roles of androgens in ovarian function has been hindered as complete androgen resistant females cannot be generated by natural breeding. In addition, androgens can be converted into estrogens which has caused confusion when interpreting findings from pharmacological studies, as observed effects could have been mediated via the AR or estrogen receptor. The creation and analysis of genetic mouse models with global and cell-specific disruption of the Ar gene, the sole mediator of pure androgenic action, has now allowed the elucidation of a role for AR-mediated androgen actions in the regulation of normal and pathological ovarian function. This review aims to summarize findings from clinical, animal, pharmacological and novel genetic AR mouse models to provide an understanding of the important roles androgens play in the ovary, as well as providing insights into the human implications of these roles.

Evidence from animal models on the pathogenesis of PCOS.

Polycystic ovarian syndrome (PCOS) is the most common endocrine condition in women, and is characterized by reproductive, endocrine and metabolic features. However, there is no simple unequivocal diagnostic test for PCOS, its etiology remains unknown and there is no cure. Hence, the management of PCOS is suboptimal as it relies on the ad hoc empirical management of its symptoms only. Decisive studies are required to unravel the origins of PCOS, but due to ethical and logistical reasons these are not possible in humans. Experimental animal models for PCOS have been established which have enhanced our understanding of the mechanisms underlying PCOS and propose novel mechanism-based therapies to treat the condition. This review examines the findings from various animal models to reveal the current knowledge of the mechanisms underpinning the development of PCOS, and also provides insights into the implications from these studies for improved clinical management of this disorder.

Multigenerational obesity-induced perturbations in oocyte-secreted factor signalling can be ameliorated by exercise and nicotinamide mononucleotide.

STUDY QUESTION: Can maternal and offspring high-fat diet (HFD)-induced changes in mRNA expression levels in mice be ameliorated by interventions in female offspring? SUMMARY ANSWER: Our results indicate that exercise and nicotinamide mononucleotide (NMN) can ameliorate the negative effects of maternal and post-weaning HFD in female offspring. WHAT IS KNOWN ALREADY: Maternal and post-weaning HFD can perturb offspring developmental trajectories. As rates of maternal obesity are rising globally, there is a need for effective treatments in offspring to ameliorate the negative effects from a maternal obesogenic environment. Modulation of the nicotinamide adenine dinucleotide (NAD+) pathway by exercise and the NAD+ precursor NMN has previously been shown to reduce the effects of obesity. STUDY DESIGN SIZE DURATION: This study consisted of a multigenerational study using C57Bl6 mice. Mice were fed a control (chow) or HFD ad libitum throughout mating, pregnancy and lactation (n = 13-25). Female offspring (n = 72) were then also supplied either a chow or HFD post-weaning. At 9 weeks of age offspring from HFD dams were subjected to exercise on a treadmill for 9 weeks or at 16 weeks of age administered NMN (i.p.) for 2.5 weeks. At 18.5 weeks mice were euthanized and ovaries and cumulus-oocyte complexes (COC) were collected to examine the possibility of ameliorating the negative effects of maternal and post-weaning HFD. PARTICIPANTS/MATERIALS SETTING METHODS: Ovary and COC mRNA expression was analysed using RT-qPCR. An initial screen of candidate genes was developed to test which molecular pathways may be involved in generating adverse reproductive system effects. For histological analysis, ovarian tissue was fixed in paraformaldehyde and embedded in paraffin and stained with haematoxylin and eosin. The numbers of primordial, primary, secondary and antral follicles were counted. MAIN RESULTS AND THE ROLE OF CHANCE: In the offspring's COC, maternal obesity increased both growth differentiation factor 9 (Gdf9: 2-fold; P < 0.05, HFD versus chow) and bone morphogenetic protein 15 (Bmp15: 4-fold; P < 0.05, HFD versus chow) mRNA expression levels while exercise and NMN interventions did not regulate Gdf9 and Bmp15 in the same manner. In whole ovary, maternal diet programmed a 25-50% reduction in FSH receptor and sirtuin-3 mRNA expression levels in daughter ovaries (P < 0.05, HFD versus chow). There was a significant interaction between HFD and intervention on the proportion of large preantral and preovulatory follicles (P < 0.05). However, the increase in preovulatory follicles did not translate to increased oocyte yield. NMN administration resulted in reduced body weight in HFD-fed individuals. LIMITATIONS REASONS FOR CAUTION: It is unclear if the changes in oocyte mRNA expression levels reported here will impact oocyte quality and fertility in offspring. Offspring ovulation rate or fecundity could not be studied here and fertility trials are required to determine if the changes in gene expression do reduce fertility. WIDER IMPLICATIONS OF THE FINDINGS: Our results demonstrate that maternal and offspring HFD perturbs key signalling pathways that are known to regulate fertility in mice, highlighting the importance of interventions in helping to prevent the declining rates of fertility in the context of the current obesity epidemic. STUDY FUNDING/COMPETING INTERESTS: This work was supported by grants and fellowships from the National Health and Medical Research Council to R.B.G. (APP1023210, APP1062762, APP1117538) and to M.J.M. and D.A.S. (APP1044295). DAS is a consultant to and inventor on patents licenced to Ovascience, Metrobiotech and GlaxoSmithKline. The other authors declare that there is no conflict of interest.

Female mice haploinsufficient for an inactivated androgen receptor (AR) exhibit age-dependent defects that resemble the AR null phenotype of dysfunctional late follicle development, ovulation, and fertility.

The role of classical genomic androgen receptor (AR) mediated actions in female reproductive physiology remains unclear. Female mice homozygous for an in-frame deletion of exon 3 of the Ar (AR(-/-)) were subfertile, exhibiting delayed production of their first litter (AR(+/+) = 22 d vs. AR(-/-) = 61 d, P < 0.05) and producing 60% fewer pups/litter (AR(+/+): 8.1 +/- 0.4 vs. AR(-/-): 3.2 +/- 0.9, P < 0.01). Heterozygous females (AR(+/-)) exhibited an age-dependent 55% reduction (P < 0.01) in pups per litter, evident from 6 months of age (P < 0.05), compared with AR(+/+), indicating a significant gene dosage effect on female fertility. Ovulation was defective with a significant reduction in corpora lutea numbers (48-79%, P < 0.01) in 10- to 12- and 26-wk-old AR(+/-) and AR(-/-) females and a 57% reduction in oocytes recovered from naturally mated AR(-/-) females (AR(+/+): 9.8 +/- 1.0 vs. AR(-/-): 4.2 +/- 1.2, P < 0.01); however, early embryo development to the two-cell stage was unaltered. The delay in first litter, reduction in natural ovulation rate, and aromatase expression in AR(+/-) and AR(-/-) ovaries, coupled with the restored ovulation rate by gonadotropin hyperstimulation in AR(-/-) females, suggest aberrant gonadotropin regulation. A 2.7-fold increase (AR(+/+): 35.4 +/- 13.4 vs. AR(-/-): 93.9 +/- 6.1, P < 0.01) in morphologically unhealthy antral follicles demonstrated deficiencies in late follicular development, although growing follicle populations and growth rates were unaltered. This novel model reveals that classical genomic AR action is critical for normal ovarian function, although not for follicle depletion and that haploinsufficiency for an inactivated AR may contribute to a premature reduction in female fecundity.

Percutaneous penetration of diethanolamine through human skin in vitro: application from cosmetic vehicles.

Concern has been raised over the safety of diethanolamine (DEA) which may be present as a minor component of alkanolamide ingredients of cosmetic formulations. Skin penetration data were therefore generated for a range of typical formulations under in-use conditions. Seven rinse-off formulations (A-E, G and H), a leave-on emulsion (F), representing prototype cosmetic formulations and containing representative levels of DEA were prepared. Target levels of DEA were attained by inclusion of DEA as either (14)C-DEA or a combination of (14)C-DEA and unlabeled DEA. Skin permeation and distribution were evaluated using human skin in vitro, static diffusion cells and phosphate buffered saline (pH 7.4) as the receptor phase. At least 12 replicate epidermal membranes were prepared from a minimum of four donors for each test group. Receptor phase samples were taken at appropriate time intervals. At the end of the test period, radioactivity remaining on the skin surface and on the diffusion cell donor cap was determined before the skin samples were tape-stripped. The remaining tissue was solubilized and radioactivity determined. Permeation was very low from all vehicles applied under in-use conditions (range 1-48 ng/cm(2) over 24 h). Comparison was also made between permeation and distribution of DEA from an infinite dose of a simple aqueous solution and the leave-on formulation (F) through paired samples of fresh and frozen full thickness skin from the same donors. When applied as an infinite dose in aqueous solution DEA permeation at 24 h was greater through frozen than through fresh skin. From the leave-on formulation, permeation was similar and very low for both fresh and frozen skin. Recovery of DEA after application of the aqueous solution to fresh human skin and subsequent aqueous and organic extraction of the epidermal and dermal tissue indicated that the majority (>98%) of DEA was in the aqueous extract, suggesting that DEA was in the free state and not associated with the lipid fraction. These data provide a basis for the estimation of the potential systemic exposure and safety margins for DEA in representative cosmetic formulations.

Veterinary drug delivery: potential for skin penetration enhancement.

A range of topical products are used in veterinary medicine. The efficacy of many of these products has been enhanced by the addition of penetration enhancers. Evolution has led to not only a highly specialized skin in animals and humans, but also one whose anatomical structure and skin permeability differ between the various species. The skin provides an excellent barrier against the ingress of environmental contaminants, toxins, and microorganisms while performing a homeostatic role to permit terrestrial life. Over the past few years, major advances have been made in the field of transdermal drug delivery. An increasing number of drugs are being added to the list of therapeutic agents that can be delivered via the skin to the systemic circulation where clinically effective concentrations are reached. The therapeutic benefits of topically applied veterinary products is achieved in spite of the inherent protective functions of the stratum corneum (SC), one of which is to exclude foreign substances from entering the body. Much of the recent success in this field is attributable to the rapidly expanding knowledge of the SC barrier structure and function. The bilayer domains of the intercellular lipid matrices within the SC form an excellent penetration barrier, which must be breached if poorly penetrating drugs are to be administered at an appropriate rate. One generalized approach to overcoming the barrier properties of the skin for drugs and biomolecules is the incorporation of suitable vehicles or other chemical compounds into a transdermal delivery system. Indeed, the incorporation of such compounds has become more prevalent and is a growing trend in transdermal drug delivery. Substances that help promote drug diffusion through the SC and epidermis are referred to as penetration enhancers, accelerants, adjuvants, or sorption promoters. It is interesting to note that many pour-on and spot-on formulations used in veterinary medicine contain inert ingredients (e.g., alcohols, amides, ethers, glycols, and hydrocarbon oils) that will act as penetration enhancers. These substances have the potential to reduce the capacity for drug binding and interact with some components of the skin, thereby improving drug transport. However, their inclusion in veterinary products with a high-absorbed dose may result in adverse dermatological reactions (e.g., toxicological irritations) and concerns about tissue residues. These are important considerations when formulating a veterinary transdermal product when such compounds are added, either intentionally or otherwise, for their penetration enhancement ability.

Physicochemical characterization of the human nail: I. Pressure sealed apparatus for measuring nail plate permeabilities.

Diffusion characteristics of the nail plate are necessary in providing the baselines for rational topical management of nail infections. In order to develop such baselines a unique stainless steel diffusion cell has been designed. The cell permits the exposure of 0.38 cm2 of nail plate to a bathing medium which is stirred by small motors mounted above the cell. The diffusion of water, methanol and ethanol at constant temperature (37 degrees C), has been examined over periods up to 4 h. Average permeability coefficients of water, methanol and ethanol were determined as 16.5 +/- 5.9 X 10(-3) cm hr-1, 5.6 X 10(-3) cm hr-1 and 5.8 +/- 3.1 X 10(-3) cm hr-1 respectively. Moreover rates of diffusion across the nail were inversely proportional to nail thickness. Based on methanol data, nail plate barrier property appears stable for long periods of aqueous immersion.

Photophysics of phenyleneethynylene metal-organic oligomers. Probing the lowest excited state by time-resolved IR spectroscopy.

The long-lived excited state in a series of metal-organic phenyleneethynylene oligomers is probed by UV-visible and infrared transient absorption spectroscopy.

Comparison of the transdermal delivery of estradiol from two gel formulations.

OBJECTIVE: Conventional oral oestrogen replacement therapy can relieve postmenopausal symptoms but is associated with undesirable side-effects which can be minimised by avoiding the fluctuating hormonal blood levels resulting from oral therapy and eliminating hepatic first-pass metabolism by the use of the transdermal route. The two commercially available transdermal gel formulations differ in composition and application recommendations. Sandrena Gel contains 0.1% (w/w) and Oestrogel 0.06% (w/w) estradiol and recommended dosages are 0.5-1.5 g over 200-400 cm2 (Sandrena Gel) and 2.5 g gel over 720 cm2 (Oestrogel). In transdermal therapy the formulation composition may have a significant effect on drug delivery and we have therefore compared the permeation of estradiol from these formulations across human skin in vitro. METHODS: The in vitro percutaneous penetration of estradiol from the formulations through epidermal membranes prepared from excised female human thing skin was assessed over a 24 h period using static type Franz diffusion cells. RESULTS: Permeation of the active was similar from each formulation representing (at 24 h) 18.2 +/- 3.5% of the applied dose from Sandrena Gel and 17.4 +/- 4.8% of the applied dose from Oestrogel. These percentages equate to cumulative skin permeations of 0.65 +/- 0.15 microgram/cm2 and 0.45 +/- 0.15 microgram/cm2 respectively. CONCLUSION: The results suggest that the two formulations are bioequivalent at the recommended dose levels.

Percutaneous penetration of dimethylnitrosamine through human skin in vitro: application from cosmetic vehicles.

Human skin penetration of N-dimethylnitrosamine (DMN) from three vehicles has been determined in vitro. When applied as an infinite dose in isopropyl myristate (IPM, 1 microgram/microliter) the average total absorption over 48 hr was 2.6 +/- 1.2% of the applied dose (all data presented are expressed as means +/- standard errors). When applied as a finite dose in a representative oil-in-water emulsion vehicle the average total absorption over 48 hr was 4.0 +/- 0.3% of the applied dose. When applied as a finite dose in a representative shampoo vehicle for 10 min followed by rinsing (i.e. to represent in-use exposure conditions) the average total absorption over 48 hr was 1.1 +/- 0.1% of the applied dose. Approximately 72% of the DMN in the applied shampoo vehicle was removed by rinsing. There was considerable evaporative loss of DMN from the IPM and oil-in-water emulsion vehicles, such that absorption was complete within 3 hr of application. The overall data indicate that DMN can penetrate the skin rapidly but that in practice the amount actually available for penetration is significantly reduced by high permeant volatility. In contrast, application of N-nitrosodiethanolamine (NDELA) at a concentration of 1 microgram/microliter as an infinite dose generated an average total absorption over 48 hr of 23.6 +/- 6.4%, representing a total flux of 103.9 +/- 28.4 micrograms/cm2. In the case of NDELA, no evaporative loss was evident.

Percutaneous penetration of octyl salicylate from representative sunscreen formulations through human skin in vitro.

The human skin penetration of [14C]octyl salicylate from two representative sunscreen vehicles was determined in vitro. 3H-sucrose was incorporated into all formulations and provided a marker for membrane integrity. When applied as a finite dose in an oil-in-water emulsion vehicle containing 5% (w/w) octyl salicylate, the average total absorption of 14C over 48 hr was 0.65+/-0.16% of the applied dose (representing a total amount permeated of 1.58+/-0.36 microg/cm2). When applied as an infinite dose in the oil-in-water emulsion vehicle the average total absorption of 14C over 48 hr was 0.47+/-0.22% of the applied dose (representing a total amount permeated of 27.54+/-13.91 microg/cm2). When applied as a finite dose in a representative hydroalcoholic formulation containing 5% (w/w) octyl salicylate, the average total absorption of 14C over 48 hr was 0.59+/-0.09% of the applied dose (representing a total amount permeated of 1.58+/-0.25 microg/cm2). When applied as an infinite dose in the hydroalcoholic formulation the average total absorption of 14C over 48 hr was 0.23+/-0.05% of the applied dose (representing a total amount permeated of 11.28+/-2.55 microg/cm2). The penetration of [14C]salicylic acid [applied at a concentration of 2.7% (w/w), in the oil-in-water emulsion] was also determined. When applied as a finite dose the average total absorption of 14C over 48 hr was 1.14+/-0.23% of the applied dose (representing a total amount permeated of 1.65+/-0.39 microg/cm2). These results suggest that the in vitro human skin permeation of octyl salicylate is relatively low. The amounts of octyl salicylate and salicylic acid permeated when applied in similar vehicles were remarkably similar over 48 hr (1.58 microg/cm2 and 1.65 microg/cm2, respectively). This suggests the possibility that the 14C label appearing in the receptor fluid may, in both cases, represent salicylic acid. If this is the case, then it is possible that the amount of octyl salicylate permeating through the skin is much less than that suggested by the data obtained here. This supposition is, however, entirely speculative and has yet to be confirmed experimentally.

Percutaneous penetration of N-nitroso-N-methyldodecylamine through human skin in vitro: application from cosmetic vehicles.

The human skin penetration of N-nitroso-N-methyldodecylamine (NDOMA) from isopropyl myristate (IPM) and two vehicles representative of cosmetic/personal care formulations was determined in vitro. When applied as an infinite dose in IPM (1 microgram/microliter) the average total absorption over 48 hr was 0.10 +/- 0.01% of the applied dose (all data are expressed as means +/- SE). When applied as a finite dose in a representative oil-in-water emulsion formulation the average total absorption over 48 hr was 4.66 +/- 0.76% of the applied dose. When applied as a finite dose in a representative shampoo formulation for 10 min, followed by rinsing (to represent in-use exposure conditions), the average total absorption over 48 hr was 0.75 +/- 0.17% of the applied dose. Approximately 72% of the NDOMA in the applied shampoo formulation was removed by rinsing. The overall data indicated that NDOMA could penetrate the skin but that penetration was low. The rate and extent of absorption, however, could be affected by differences in the vehicle of application, time of exposure and whether the formulation is (and the conditions are designed to mimic) a rinse-off or leave-on product.

Non-ionic surfactant effects on hairless mouse skin permeability characteristics.

The influence of a range of polyethoxylated non-ionic surfactants upon the transport of methyl nicotinate across hairless mouse skin in-vitro was investigated using standard two-compartment diffusion cells. Those surfactants having a linear alkyl chain greater than C8 and an ethylene oxide chain length of less than E14 caused significant increases in the flux of methyl nicotinate. Surfactants having branched or aromatic moieties in the hydrophobic portion were ineffective. Maximum enhancement of flux was obtained using polyoxyethylene (10) lauryl ether (Brij 36T). Two possible modes of surfactant action are proposed. Initially the surfactant may penetrate into the intercellular regions of the stratum corneum, increase fluidity and eventually solubilise and extract lipid components. Secondly, penetration of the surfactant into the intracellular matrix followed by interaction and binding with the keratin filaments may result in a disruption of order within the corneocyte. The structural specificity required for the latter mechanism may explain, to some extent, the maximum activity obtained with the C12 surfactant.

Facilitated transport of sodium salicylate across an artificial lipid membrane by Azone.

The ability of Azone (1-dodecylazacylcoheptan-2-one), a recently developed penetration enhancer, to facilitate the transport of sodium salicylate across an artificial lipid membrane has been investigated using the rotating diffusion cell, a well defined model for percutaneous absorption. Azone was found to be capable of enhancing the transport of the salicylate anion across an isopropyl myristate membrane, by using a pH gradient as the chemical driving force. The results indicate that Azone may be capable of forming ion pairs with anionic drugs.

Penetration of the human nail plate: the effects of vehicle pH on the permeation of miconazole.

In order to assess the relative permeability of the nail plate to ionized and unionized drugs the permeation of miconazole at varying pH has been followed as a function of time. The pH was adjusted from 3.1 to 8.2 to obtain between 5 and 100% dissociation of the drug. No significant difference in the rate of permeation was demonstrated. The data suggests that the ionic form of miconazole dissolves as easily in the nail plate as the free base and, therefore, topical bioavailability can be enhanced by decreasing the formulation pH thereby increasing drug solubility.