Introduction

Females in many mammalian species undergo a period of time referred to as estrus, when they are most likely to conceive and are more receptive to copulation attempts by males (Miller, Tybur, & Jordan, 2007). During estrus, several primate species exhibit visual cues to their physiological status, like sexual swelling (Grammer, Fink, & Neave, 2005). Humans do not exhibit overt visual cues during ovulation. Thus, human ovulation is often termed concealed, which may provide some advantages to women, for example, increasing the appeal of monogamous pair-bonding thereby increasing male parental investment, increasing chances of successful cuckoldry, and decreasing the risk of infanticide (Grammer et al., 2005; Roberts et al., 2004).

Sexual Conflict and Parental Investment Theory explains the diverging and often conflicting evolutionary interests of males and females (Chapman, Arnqvist, Bangham, & Rowe, 2003). The disparity between male and female potentials for lifetime reproductive success is reflected in the mating stratagems adopted by each sex (Trivers, 1972). Sperm are produced throughout a lifetime, eggs, on the other hand, are finite. Eggs are also the more energetically costly sex cell. Any copulation event for a human female could result in 9 months of investment in a single offspring, thus females benefit from being selective in their mate choice. Male investment, on the other hand, can end with ejaculation. Males must compete for access to the limiting resource, in this case females, which results in some males monopolizing multiple females and other males being left out the mating game entirely. Females have a low risk of being left out the mating game, as mating with any female is low cost and high benefit for a male. Because the best interest of males and females are in conflict, with regard to reproductive success, an evolutionary “arms race” manifested as certain behavioral tendencies, such as mate guarding, and physiological adaptations, like nonadvertised ovulation, may be underway (Trivers, 1972).

Mate guarding and sexual jealousy occurs when a partner perceives a threat to their mating relationship (Buss, 2002). Buss (2002) described seven typical mate-guarding tactics: vigilance, violence, concealment, monopolization of time, sexual inducement, physical signs of possession, and possessive ornamentation. Some examples of these tactics include obsessive calling and checking the whereabouts of one’s partner, dissuading a partner from attending a social gathering where the opposite sex will be present, and even the custom of wedding rings that signify the wearer is already in a committed relationship.

Haselton and Gangestad (2006) showed that nonpill using pair-bonded women self-reported more flirtatious behavior and an increase in their partner’s mate guarding around ovulation. Near and during ovulation women are more likely to seek extrapair copulations. Women may choose to engage in a long-term relationship with men who are willing to invest their time and resources in an offspring but seek “sexy genes” during ovulation in an attempt to improve the overall genetic quality of their offspring while also cuckolding parentally investing men (Trivers, 1972). In self-report surveys, ovulating women do show an increased preference for more attractive men and an increased desirability for men besides their current partner. This result is more extreme when the woman’s primary partner does not possess “sexy traits” (Haselton & Gildersleeve, 2011). Gangestad, Garver-Apgar, Cousins, and Thronhill (2014) found that nonpill using women report greater self-assertiveness and engage in more behaviors aimed at resisting their partners mate guarding during their fertile phase. The women’s partners also self-reported more assertive behaviors toward their partner, while she was at high conception risk.

Increased mate-guarding behavior during ovulation may be related to something more than a response to changes in female behavior (i.e., increased flirtation and desire for extrapair copulation; Haselton & Gangestad, 2006). A large portion of research on sexual behavior concerns visual cues and behavioral signaling. Substantially less research has been devoted to olfactory influences as humans have traditionally been classified as microsmatic (poor smellers; Carlson, 2013). Porter et al. (2007) suggest that the perceived differences in human and other species olfactory capabilities might be due to the anatomical adaptation of bipedality and not a direct reflection of olfactory ability. The current study argues men may be able to detect odor changes related to the physiological status of women, and this may explain subsequent behavioral and physiological changes in men.

In a number of animals, including some primates, chemicals have been shown to advertise female ovulation and affect male mating behavior (Miller & Maner, 2011). These chemicals are referred to as pheromones. Pheromones are defined as chemicals released from an organisms’ body into the environment that result in certain responses, both physiological and behavioral, in conspecifics. Some well-known physiological responses in mammals to pheromones include the Lee-Boot effect, the Bruce effect, and the Whitten effect (Grammer et al., 2005).

Cummins et al. (2011) reported a striking example of intrasexual aggression induced via a pheromone produced by female squid. A protein, Loligo β-microsemino, synthesized by female squid causes male squid upon contact to rapidly change from a calm state to one of extreme aggression, resulting in the exhibition of fighting displays. In their natural environment, female squid produces this protein and inserts it into the outer portion of the egg capsule. Males are attracted to egg capsules visually and upon contact with Loligo β-microsemino, become aggressive. Male–male competition ensues and the female squid benefits from the acquisition of the victor’s sperm. Laboratory studies showed that the presence of Loligo β-microsemino without a female or egg capsules present was enough to trigger behavioral shift in males. Loligo β-microsemino is related, albeit distantly, to a protein represented in female mammalian reproductive secretions.

In some primate species, females produce vaginal secretions that may influence males behaviorally and hormonally. Michael and Keverne (1970) reported that estrogen stimulated some primates to produce vaginal secretions containing a chain of five volatile aliphatic acids: acetic, propanoic, methylpropanoic, butanoic, and methylbutanoic. This mixture of fatty acids, called copulins, could potentially be classified as a pheromone, since they are produced vaginally and trigger certain behavioral and physiological reactions in conspecifics (Grammer et al., 2005).

Copulins were first identified within the vaginal secretions of rhesus monkeys (Macaca mulatta; Michael & Zumpe, 1982). Copulins are registered by male rhesus monkeys through olfaction and affect mate choice. Michael and Zumpe (1982) placed four bilaterally ovariectomized females into a social group with one male. Testosterone was administered to two of the females causing them to be more receptive to approach and mounting attempts. One of the testosterone treated females was also treated with synthetic copulins, while the other was treated with ether. Female treatments were altered every trial to control for male mate-choice bias. The results indicated that copulins significantly increased mounting attempts and ejaculations. This suggests copulins act as a significant olfactory stimulus affecting male rhesus monkey mating behavior by allowing them to discriminate, which females in a social group are ovulating (Michael & Zumpe, 1982).

A similar study was conducted with stump-tailed macaques (Macaca arctoides). Female stump-tailed macaques lack sexual swellings, yet males have been shown to discriminatorily engage in copulatory behavior with females during the periovulatory phase. Vaginal secretions were collected throughout the five menstrual phases, and males were exposed to cotton swabs soaked in these secretions or salt water. Upon returning to the home cage, males exposed to secretions from the ovulatory phase showed increased exploratory and coercive sexual behaviors (Cerda-Molina, Hernandez-Lopez, Rojas-Maya, Murcia-Mejia, & Mondragon-Ceballos, 2006).

Matsumoto-Oda et al. (2003) identified a chain of six volatile fatty acids: mucus, acetic acid, propionic acid, butyric acid, iso-butyric acid, and valeric acid in the vaginal secretions of chimpanzees. Four captive female chimpanzees were observed throughout their cycles. One female exhibited significantly higher levels of acetic and propionic acids in her vaginal secretions. She was also the only female to be followed by a male and impregnated during the study. Levels of acetic and propionic acids were stable across all the female’s cycles. Only the concentration of iso-butyric acid fluctuated throughout the menstrual cycle. This finding suggests that iso-butyric acid may act as a signal or cue to reproductive status. The other acids, specifically acetic and propionic, may play a role in advertising overall genetic quality and fecundity.

Cross-species comparisons between humans and primates that exhibit sexual swelling or seasonal breeding patterns suggest that ovulatory cues have been lost in humans. However, women do display some cues related to their reproductive status such as behavioral shifts. Grammer, Renninger, and Fischer (2004) reported that ovulating nonpill using women at night clubs in Vienna danced more provocatively, showed more skin, and self-rated their outfit choices as sexier and bolder. They interpreted their results to imply women consciously display social signals with their clothing choices. Men report their perception of a woman as a partner is altered by her clothing choice. Women models showing more skin are reported as attractive for a sexual partner but less attractive as a long-term mate (Grammer, Renninger, & Fischer, 2004). Other research shows that near ovulation, women report an increased preference for men, other than their partner, that possess more symmetrical bodies and immunocompatible odors (Singh & Bronstad, 2001). Grammer, Fink, and Neave (2005) found that 13.8% of unprotected extrapair copulations happened during ovulation. According to parental investment and sexual selection, women should be seeking both good genes and good parental partners. Women may form pair bonds with more parentally investing partners and sneak extrapair copulations during ovulation with men exhibiting sexy genes. Sperm competition will improve the overall genetic quality of a woman’s offspring, as the “best” sperm will impregnate her egg (Grammer et al., 2004; Trivers, 1972). Similar research demonstrates that women not only exhibit behavioral shifts but also shift in physical appearance throughout their cycles.

Symmetry is selected for in many species as an indicator of overall genetic quality. For example, fluctuating asymmetry in plants is a product of sexual selection and involved in signaling to pollinators (MØller & Eriksson, 1994). Human mating stratagems may correlate with symmetry, in that higher symmetry, men are likely to develop short-term mating strategies and asymmetrical men are likely to develop long-term mating strategies (Haselton & Gangestad, 2006). The sexy genes discussed in previous paragraphs is a reference to symmetry, among other factors (Grammer et al., 2004; Trivers, 1972). Women exhibit fluctuating asymmetry in relation to the menstrual cycle (Roberts et al., 2004). Soft tissues (ears, fingers, and breast) become more symmetrical during ovulation (Manning, Scutt, Whitehouse, Leinster, & Walton, 1996). Other subtle shifts in a woman’s physical features include lighter facial skin pigmentation, improved waist-to-hip ratio, and fluctuations in vocal pitch (Miller & Maner, 2011; Van den Berghe & Frost, 1986). Men judge photos of a woman not using hormonal contraceptives as more attractive during the follicular phase and less attractive during the luteal phase, suggesting that men may use fluctuating asymmetry as a cue to reproductive status (Roberts et al., 2004).

Miller, Tybur, and Jordan (2007) found that nonpill using lap dancers earned US$350 in tips while ovulating compared to US$260 in the luteal phase and US$185 during menstruation. One hypothesis based on behavioral differences suggests the discrepancy in earnings is due to more flirtation and provocative dancing during ovulation. Another hypothesis suggests men are unconsciously detecting olfactory cues to the physiological status of the dancer (Haselton & Gangestad, 2006; Haselton & Gildersleeve, 2011).

Michael, Bonsall, and Kutner (1975) examined vaginal secretions throughout the menstrual cycle of 50 young women via a tampon method. Their findings show that humans produce the same copulins as nonhuman primates and that copulins increase in concentration significantly during high fertility phases and decrease during low fertility phases. Before entering into a discussion on chemical communication via copulins in humans, it is worth mentioning that copulins could be a byproduct of shared phylogeny with nonhuman primates and of little importance in modern society. This will be brought up again in the discussion of future research (see the Section Discussion of Men’s Behavior in Response to Copulin Exposure). Michael et al. (1975) found that women using oral contraceptives were clearly distinguishable from normal cycling women because the overall content of fatty acids in their vaginal secretions was low and there was no alteration in concentrations throughout their menstrual cycle. Acetic acid was the most dominant acid found in the participants’ vaginal secretions, which is congruent with chimpanzee vaginal secretion findings (Matsumoto-Oda et al., 2003). In the chimpanzee study described earlier, researchers suggested that acetic and propionic acid signal overall female genetic quality because of their relatively stable concentrations across the menstrual cycle. Iso-butyric acid, on the other hand, fluctuates throughout the menstrual cycle, and these fluctuations may be detected by males (Matsumoto-Oda et al., 2003). Acetic acid in human vaginal secretions is also relatively stable across the menstrual cycle and has a high threshold for olfaction. Thus, acetic acid, although the most dominate, contributes the least to vaginal odor. Waltman et al. (1973) reported iso-butyric acid, which typically fluctuates the most in human vaginal secretions across the menstrual cycle, decreases in production by postmenopausal women. Postmenopausal women do not ovulate and subsequently do not need to secrete an acid that plays a role signaling reproductive status. Thus, changes in men’s behavior and perceptions surrounding an ovulating woman may not be solely due to changes in her behavior and fluctuating asymmetry but also related to fluctuations in concentrations of copulins, specifically iso-butyric acid, in her vaginal secretions.

Many studies show men are able to distinguish nonovulating women from ovulating women via scent alone. In one example of a classic smelly T-shirt study, nonpill using women in both the luteal and follicular phases wore t-shirts to bed for three days in a row. Afterward, men judged the scent of these T-shirts. Results indicated that men preferred the scent of women in the follicular phase. They rated those T-shirts as the most pleasant and sexy smelling (Singh & Bronstad, 2001).

Miller and Maner (2010) took hormonal measurements after exposing men to T-shirts worn by ovulating and nonovulating women. Men exposed to T-shirts worn by ovulating women exhibited a significant increase in testosterone. Testosterone has been implicated in, among other things, aggressive behaviors like competiveness and dominance. These are traits that may be valued by women, especially during ovulation, and are related to intrasexual aggression (Kenrick et al., 2009; Miller & Maner, 2010). In the presence of an ovulating woman, we suggest that men may be primed by copulins to compete with each other and display aggression, similar to the effects of Loligo β-microsemino in squid (Cummins et al., 2011). In a series of follow-up studies, Miller and Maner (2011) had men complete an emotional perception task after being exposed to T-shirts worn by ovulating and nonovulating women. Men in the ovulating condition reported higher levels of sexual arousal and were more likely to complete a word stem to form a sexual word. They also rated the sexual interest of a female confederate significantly greater than men in the nonovulating condition. In a separately run study, Miller and Maner (2011) had participants interact with a confederate, while she was at a high-conception risk or low-conception risk. Men were accessed on how often they mimicked her body language and their risk taking in a black jack game the confederate watched. Men in the ovulating condition were more likely to mimic the confederate’s posture and make riskier decisions (Miller & Maner, 2011).

Doty, Ford, Preti, and Huggins (1975) found that on average men and women rate preovulatory and ovulatory secretions as weaker and less intense than secretions from other phases of the menstrual cycle. Cerda-Molina, Hernandez-Lopez, De la O, Chavira-Ramirez, and Mondragon-Ceballos (2013) provide evidence that men exhibit endocrine changes specifically to increased concentrations of chemicals in women’s axillary and vulvar secretions at ovulation. Axillary and vulvar scents from high fertility phases increased cortisol and testosterone levels with vulvar scents, creating a longer lasting changes. General sexual interest also increased. Luteal vulvar scents were rated as intense and unpleasant compared to the other scents. These studies as well as others presented above show that men are sensitive to high fertility and low fertility odors, but these studies do not isolate copulins as the potential chemical mechanism.

Jütte and Grammer (1997) exposed men to copulins, while having them rate the attractiveness of women’s faces. Hormonal measurements showed an increase in testosterone for men in the copulin condition. Men in the copulin condition also seemed inhibited in their ability to discriminate attractiveness, meaning they rated all of the women’s faces as significantly more attractive than controls and exhibited a decrease in variance of responses. A follow-up study replicated the hormonal findings and indicated men exposed to copulins were less cooperative in a tragedy of the commons paradigm than controls (Steinbach, Oberzaucher, & Grammer, 2012).

In our study, we causally tested whether copulin exposure increased men’s mate-guarding behavior, ratings of women’s attractiveness, and self-perceived sexual desirability. We sought to replicate Jütte and Grammer (1997) by exposing men to synthetic copulins while having them rate photographs of women’s faces. Prior research has focused on behavioral shifts and fluctuating asymmetry as subtle cues to human ovulation. Yet we know, olfaction and chemical signaling play an important role in social interactions and mating behavior in diverse animal taxa. The relevance of this in primates, specifically humans, has been discounted, as human sensory evolution has been characterized by increased optical primacy. However, a profusion of recent studies challenges this view. Our study will add to this growing body of research and try to answer the question: What functions does olfaction play, if any, in human mating psychology?