Effective prevention of stress‐induced sweating and axillary malodour formation in teenagers

Emotional sweating and malodour production represent a relevant challenge to today’s antiperspirant (AP) and deodorant products as stress in everyday life increases continuously. The aim of this study was to investigate stress‐induced sweating in teenagers who are known to experience various stressful situations, e.g. exams at school or job interviews. To induce emotional sweating in 20 female and 20 male adolescents (16–18 years of age), we applied the Trier Social Stress Test (TSST), considered today to be the most reliable and standardized stress protocol. In this study, we demonstrate that the TSST induces high amounts of sweat and strong axillary malodour in the tested age group. Notably, male teenagers showed significantly higher stress‐induced odour scores than female subjects, although no gender differences were detected concerning other physiological stress markers. Testing of a novel deodorant/AP product developed to specifically address the needs of adolescent consumers revealed excellent deodorant and AP efficacy under the challenging conditions of the TSST.     

Effect of antiperspirants on whole body sweat rate and thermoregulation

It is well established that the evaporation of sweat from the human body surface is the main mechanism by which heat balance is maintained following a rise in body core temperature. Since the introduction of the first brand name antiperspirant in the United States during the early 1900s, antiperspirant products designed to control underarm wetness have grown to represent one of the largest cosmetic categories in most global markets. However, although axillary sweating only constitutes less than 1% of whole body sweat rate, consumers, particularly in hot countries, have begun to articulate the concern that antiperspirants may interfere with the body's natural cooling process. To investigate this, we undertook carefully designed experiments that measured the effects of axillary antiperspirant application on whole body sweat rate and body core temperature, following a regimen of exercise-induced heat stress in a hot environment in human volunteers. Our data show clearly that although antiperspirant prevents sweat production in the axillary area, this does not impact the ability of the body to thermoregulate following a rise in body core temperature. Thus, recent consumer questioning over this aspect of antiperspirant use appears to be unwarranted.     

Body malodours and their topical treatment agents

Body malodour, including foot odour, suppresses social interaction by diminishing self-confidence and accelerating damage to the wearer's clothes and shoes. Most treatment agents, including aluminium anti-perspirant salts, inhibit the growth of malodourous bacteria. These metallic salts also reduce sweat by blocking the excretory ducts of sweat glands, minimizing the water source that supports bacterial growth. However, there are some drawback effects that limit the use of aluminium anti-perspirant salts. In addition, over-the-counter anti-perspirant and deodourant products may not be sufficiently effective for heavy sweaters, and strong malodour producers. Body odour treatment agents are rarely mentioned in the literature compared with other cosmetic ingredients. This review briefly summarizes the relationship among sweat, skin bacteria, and body odour; describes how odourous acids, thiols, and steroids are formed; and discusses the active ingredients, including metallic salts and herbs, that are used to treat body odour. A new class of ingredients that function by regulating the release of malodourants will also be described. These ingredients do not alter the balance of the skin flora.     

New antiaxillary odour deodorant made with antimicrobial Ag-zeolite (silver-exchanged zeolite)

The causative substances for axillary osmidrosis, which are often found in apocrine sweat, are the decomposed/denatured products of short-chain fatty acid and other biological metabolite compounds produced by axillary-resident bacteria. Conventional underarm deodorants suppress the process of odour production mostly by the following mechanism: (1) suppression of perspiration, (2) reduction in numbers of resident bacteria, (3) deodorization and (4) masking. The most important and effective method to reduce odour is to suppress the growth of resident bacteria with antimicrobials, which have several drawbacks, especially in their safety aspect. To solve these problems, we focused on Ag-zeolite (silver-exchanged zeolite) that hold stable Ag, an inorganic bactericidal agent, in its structure, and therefore, poses less risk in safety. Its bactericidal effect on skin-resident bacteria was found to be excellent and comparable with that of triclosan, a most frequently used organic antimicrobial in this product category. The dose-response study of Ag-zeolite powder spray (0-40 w/w%) using 39 volunteers revealed that 5-40 w/w% Ag-zeolite could show a sufficient antimicrobial effect against skin-resident bacteria. The comparison study using 0.2 w/w% triclosan as the control and 10 w/w% Ag-zeolite indicated that: (1) one application of the powder spray containing 10 w/w% Ag-zeolite could show a sufficient antimicrobial effect against the resident bacteria and its effect continued for 24 h, (2) a powder spray containing 0.2 w/w% triclosan was unable to show a sufficient antimicrobial effect, and (3) no adverse event was observed. These studies show that Ag-zeolite has a superior antimicrobial ability that is rarely found in conventional antimicrobials used in deodorant products and a strong antiaxillary odour deodorant ability because of its long-lasting effect. During clinical study, patch tests with humans and other clinical studies of this product showed no adverse events related to the treatment with the Ag-zeolite product.     

Molecular basis of human body odour formation: insights deduced from corynebacterial genome sequences

During the past few decades, there has been an increased interest in the essential role of commensal skin bacteria in human body odour formation. It is now generally accepted that skin bacteria cause body odour by biotransformation of sweat components secreted in the human axillae. Especially, aerobic corynebacteria have been shown to contribute strongly to axillary malodour, whereas other human skin residents seem to have little influence. Analysis of odoriferous sweat components has shown that the major odour-causing substances in human sweat include steroid derivatives, short volatile branched-chain fatty acids and sulphanylalkanols. In this mini-review, we describe the molecular basis of the four most extensively studied routes of human body odour formation, while focusing on the underlying enzymatic processes. Considering the previously reported role of β-oxidation in odour formation, we analysed the genetic repertoire of eight Corynebacterium species concerning fatty acid metabolism. We particularly focused on the metabolic abilities of the lipophilic axillary isolate Corynebacterium jeikeium K411.     

Manipulation of body odour alters men's self-confidence and judgements of their visual attractiveness by women

Human body odour is important in modulating self-perception and interactions between individuals. Artificial fragrances have been used for thousands of years to manipulate personal odour, but the nature and extent of influences on person perception are relatively unexplored. Here we test the effects of a double-blind manipulation of personal odour on self-confidence and behaviour. We gave to male participants either an aerosol spray containing a formulation of fragrance and antimicrobial agents or an otherwise identical spray that lacked these active ingredients. Over several days, we found effects between treatment groups on psychometric self-confidence and self-perceived attractiveness. Furthermore, although there was no difference between groups in mean attractiveness ratings of men's photographs by a female panel, the same women judged men using the active spray as more attractive in video-clips, suggesting a behavioural difference between the groups. Attractiveness of an individual male's non-verbal behaviour, independent of structural facial features, was predicted by the men's self-reported proclivity towards the provided deodorant. Our results demonstrate the pervasive influence of personal odour on self-perception, and how this can extend to impressions on others even when these impressions are formed in the absence of odour cues.     

Design and application of a screening and training protocol for odour testers in the field of personal care products

The assessment of odours and in particular of human axillary odour is an integral part of the research and development of deodorant and anti-perspirant products. One method to perform odour assessment is the odour evaluation that is carried out by experts, designated as odour testers or sniffers. Product development decisions are therefore based on human assessment. As for every scientific measurement, the influencing factors need to be standardized or regularly calibrated as effectively as possible for reasons of quality assurance. We therefore developed a screening and training concept aiming to examine the general suitability of odour testers by determining the individual odour sensitivity for relevant odours. This newly developed method is based on the national and international standards and guidelines EN 13725:2003, VDI 3882 sheet 1 and ASTM-1207. Suitable odour testers are subsequently trained to correlate their individual odour intensity perception with an intensity calibration scale in order to achieve reproducible results. Training sessions held on a regular basis help to achieve a greater homology in the response of an existing panel. Our established screening and training protocol has already been successfully put into practice and is also subject to permanent improvement with regard to practical requirements.     

Iron sequestration on skin: a new route to improved deodorancy

Axillary malodour is caused by the biotransformation of non-odorous precursors present in apocrine sweat and sebum by the axillary microflora. To counter this, underarm products typically contain high levels of bactericides. However, after an initial decrease in bacterial numbers, the surviving cells grow, producing a concomitant rise in axillary odour. A sustained deodorant effect might be achieved without recourse to bactericidal action if this bacterial growth could be inhibited for extended periods. The current study attempted to inhibit axillary bacterial growth by nutrient deprivation, primarily that of iron (Fe(III)). In vitro analyses identified iron (Fe(III)) as the trace metal whose deprivation had the most profound effect on bacterial growth. Further in vitro investigations with Fe-chelating agents demonstrated that a number of compounds with high binding constants for Fe(III) showed optimal activity. One candidate molecule, diethylenetriaminepentaacetic acid (DTPA), was capable of effectively inhibiting bacterial growth in vitro and on the skin of the lower back. Some bacterial species could additionally utilize iron bound to the iron carrier protein transferrin present in eccrine sweat. This was minimized by the use of an agent, butylated hydroxytoluene (BHT), capable of liberating iron from transferrin via reduction of transferrin-bound ferric ions, allowing subsequent sequestration of Fe(II). Deodorant efficacy evaluation of the combination of DTPA and BHT showed deodorancy benefits over and above that afforded by DTPA alone. This mixture of DTPA and BHT supplemented to a standard ethanolic deodorant, used on 50 people for 2 weeks, was highly effective in limiting bacterial growth in the axilla. Total aerobic bacteria in the axillae were reduced from a mean of log 5.75 (+/-0.73) to log 4.50 (+/-0.90) colony-forming units (cfu) cm(-2) (n = 27, P < 0.01) compared with a non-fortified standard ethanolic deodorant. This was reflected in significant decreases in axillary malodour production, as determined by malodour assessments (P < 0.01). The profile of the axillary microflora was maintained, and all populations were rapidly returned to preuse levels after cessation of product use. This new deodorant technology was benchmarked against leading antimicrobial-based deodorant systems. In three separate deodorant efficacy evaluations, the combination of DTPA and BHT was tested against Triethyl citrate, Triclosan and Farnesol in standard unfragranced ethanolic formulations. The combination of DTPA and BHT showed highly significant deodorancy benefits over and above all these antimicrobial-based deodorant technologies. The combination of an efficient iron chelator with an agent capable of liberating iron from transferrin offers significant benefits in terms of bacterial growth inhibition on the skin and provides a new route to axillary deodorancy.     

Reduced barrier efficiency in axillary stratum corneum

The skin of the axilla is cosmetically important with millions of consumers daily applying antiperspirant/deodorant products. Despite this, we know virtually nothing about axillary skin or how antiperspirant (AP) use impacts upon it. To characterize the axillary stratum corneum and determine whether this is a unique skin type, we have looked at stratum corneum composition and function, particularly its barrier properties, and compared it with other body sites. Transepidermal water loss (TEWL) and corneosurfametry (CSM) revealed a reduced barrier function in the axilla. HPTLC analysis of the stratum corneum lipids demonstrated statistically elevated levels of fatty acids, ceramides, and particularly cholesterol in the axilla. Both ceramide and cholesterol did not appear to change with depth, indicating that they were predominantly of stratum corneum origin. On the other hand, at least some of the fatty acid had a sebaceous origin. We hypothesized that the reduced barrier function might be owing to the changes in the crucial ceramide : cholesterol ratio. To address this, we used a combination of attenuated total reflectance-Fourier-transformed infrared spectroscopy (ATR-FTIR) with cyanoacrylate sampling. These results demonstrated more ordered lipid-lamellae phase behaviour in the axilla, suggesting that the elevated cholesterol might form crystal microdomains within the lipid lamellae, allowing an increase in water flux. Since an exaggerated application of antiperspirant had no effect upon the axilla barrier properties, it is concluded that this region of skin physiologically has a reduced barrier function.     

Erroneous gender differences in axillary skin surface/sweat pH

Assessing accurately the pH of axillary eccrine sweat is of vital importance in the antiperspirant industry. Eccrine sweat pH is a critical parameter in determining the effectiveness of antiperspirants; antiperspirant salts dissolve in sweat and diffuse into the sweat glands, where the resultant acidic solution hydrolyses in more alkaline sweat forming an amorphous metal hydroxide gel, thereby restricting the flow of eccrine sweat. Comparison of the skin surface and sweat pH of males and females reported in the literature shows that, although consistent male/female differences have been observed on the forearm, determination of significant gender-based pH differences across other sites are less conclusive. Studies on the back and infra-mammary regions exhibited significant gender differences in skin surface pH, whereas those on the forehead, cheek, neck and inguinal area showed no such difference. With regard to the axilla specifically, four studies have been reported, three showing no significant difference in axillary skin surface pH and one indicating that females have an eccrine sweat pH of 7 and males have a sweat pH of 5.6. This paper describes a series of carefully controlled studies aimed at assessing potential gender differences in eccrine sweat and skin surface pH following exposure to a variety of temperature, humidity and time conditions. The results highlight the importance of controlling precisely the time of investigation, site of measurement and, most importantly, the necessity to pre-equilibrate samples in 40 mmHg carbon dioxide (equivalent to arterial CO(2) tension (pCO2)) before determining sweat pH. When these parameters are controlled no gender differences in axillary sweat or skin surface pH are observed. Large differences in eccrine sweat and skin surface pH are found, however, between the vault (hairy region) and fossa (non-hairy region) of the axilla.     

The evaluation of underarm deodorants

Double-blind cross-over trials in which trained assessors evaluated axillary malodour on a 0 to 10 scale showed that ethanol at 60% and 99% w/w significantly reduced odour for up to 24 h. Reduction in odour was increased by the addition of non-volatile antimicrobial ingredients such as chlorhexidine, Irgasan DP 300 (Triclosan, Ciba Geigy) or aluminium chlorhydrate.
Volunteer panellists were able to identify statistically significant deodorant effects when they rated their own axillary odour on a 0 to 10 scale.
An odour absorber, zinc ricinoleate (Grillocin, Grillo-Werke, A. G.) was used to treat existing malodour and gave reduced perception of that malodour for up to 24 h.
A close correlation was found between panel odour scores and the bacterial status of the axilla as assessed by the replipad technique when biocidal systems were studied. Irgasan DP 300, a bacteriostat, did not perform so well in skin flora sampling procedures, but was nevertheless effective as a deodorant ingredient. Thus in vivo microbiological techniques, although often useful as predictors of deodorancy, do not replace the carefully controlled clinical trial with direct assessment of odour.
L'evaluation de l'odeur des aisselles     

Circadian rhythms in axillary skin surface pH

Sweat and skin surface pH are critical parameters in determining the performance of antiperspirants. The mechanism of action, the so-called 'plug theory' first proposed by Reller and Luedders, involves the expression of eccrine sweat onto the surface of the skin into which the solid antiperspirant salts, typically an aluminium chlorohydrate or zirconium aluminium glycine, dissolve. The resultant acidic 'solution' then diffuses with time into the sweat glands, where it hydrolyses in more alkaline sweat and forms an amorphous metal hydroxide agglomerate that physically plugs the ducts some 20-100 mum into the glands. It is therefore important to understand whether diurnal variations in skin surface pH exist in the axilla, as these may influence strongly the time of day at which antiperspirant should be applied in order to yield maximal protection. Clinical studies demonstrate a significant fall in axillary skin surface pH between the morning (pH = 5.87 +/- 0.23) and the evening (pH = 5.49 +/- 0.23). This diurnal variation in skin surface pH suggests that antiperspirant efficacy will be optimal when products are applied in the morning. In addition, the data suggest a circadian rhythm in axillary skin barrier function, indicating that chronopharmacology, the timing of administration of medication, could be used to optimize treatment of axillary hyperhidrotics using topical administration of anticholinergic drugs.     

A new microbiological approach to the assessment of underarm deodorants

A general survey of odour generation in the axillae is given, with particular reference to the role of topically applied antimicrobial agents in odour suppression.
A new apparatus is described which will allow a rapid and reliable quantification of in vivo antimicrobial action and which may be usefully employed in optimization of individual systems to achieve effective deodorancy.
Findings are presented on the antimicrobial efficacy of deodorant compositions and correlated with practical olfactory studies.
Nouvelle approche microbiologique dans l'évaluation des déodorants axillaires     

Retention of axillary odour on apparel fabrics

Clothing worn in close proximity to the human axilla can retain and emanate human body odour even remaining odorous long after removal from the body. Intensity of odour is affected by the fibre type from which the garment is made. Headspace analysis of axillary volatile compounds released from three interlock fabrics (cotton, wool, polyester) following wear were measured using an online monitoring instrument, proton transfer reaction-mass spectrometry (PTR-MS). Compounds likely to be short-chain carboxylic acids increased in the headspace above the polyester fabrics after 7 days. This increase was not evident above either the wool or cotton fabrics. The intensity of axillary odour emanating from these fabrics was inversely related to fibre hygroscopicity. The relationship between a textile fibre/fabric's ability to retain and emanate odour is likely to be related to the metabolic versatility of resident microbial strains and/or the chemical and physical morphology of the fibre and its ability to absorb volatile compounds and precursors to odour.     

The suppressive effect of apricot kernel extract on 5alpha-Androst-16-en-3-one generated by microbial metabolism

Body odours are generated from dead skin cells and secreted materials, such as sweat and sebum, through the metabolism of microorganisms living on the skin. Volatile steroids, key compounds in body odours, are also generated through the metabolism of microorganisms. These volatile steroids strengthen the intensity of the overall body malodour and are sensed differently by males and females. Females are more sensitive than males to volatile steroids, especially 5alpha-androst-16-en-3-one (androstenone). To regulate body odours that are especially unpleasant for women, we devised an androstenone-generation model using the metabolism of Corynebacterium xerosis, which is one of the bacteria living on the axillary skin. Using this model, we studied the suppressive effect of plant extracts on the generation of androstenone. We found that apricot kernel extract (AKE) had the most positive effect among the plant extracts to which we applied the model. However, although AKE did suppress androstenone generation, it did not show any bactericidal effect. Using the cell-free system, AKE also suppressed the generation of androstenone. In conclusion, we found that AKE suppressed the generation of androstenone, which is especially unpleasant for women, and the mechanism was not bactericidal but metabolic inhibition. The results of these studies provide new understanding of the regulation of androstenone, which, in turn, should lead to the development of novel deodorant systems.     

A new approach to assessing sweat production

Since 1916 when Stillians (1) first reported that excessive axillary sweating could be reduced by the application of 25% aqueous aluminium chloride, sales of antiperspirant products have increased dramatically. In spite of many studies, the mechanism of action of these compounds has not been clearly elucidated (2). In most cases, investigations have been performed on animal foot pads, excised human biopsy specimens and skin samples in vivo (3). A model which eliminated problems encountered in whole body investigations (for example, influence of blood flow) would be preferable. One such method, introduced in part by Sato (3), has been investigated. Preliminary investigations suggest that this method could become useful in the evaluation of the mode of action of antiperspirants.
Sweat production was induced using carbachol and methacholine in isolated rat plantar sweat glands. These were maintained in a suitable environment (modified Krebs-Ringer bicarbonate at 38°C containing 30% w/v fresh rat serum). The output of sweat was measured every 5 minutes for 85 minutes and the maximal final volume obtained with any one gland was 23 nanolitres.
Sweat production ceased on administration of appropriate concentrations of atropine or the withdrawal of calcium ions from the bathing fluid.
Une nouvelle approche pour évaluer la transpiration     

Characterization of the microflora of the human axilla

It is widely accepted that axillary malodour is attributable to the microbial biotransformation of odourless, natural secretions into volatile odorous products. Consequently, there is a need to understand the microbial ecology of the axilla in order that deodorant products, which control microbial action in this region, can be developed in the appropriate manner. A detailed characterization of the axillary microflora of a group of human volunteers has been performed. The axillary microflora is composed of four principal groups of bacteria (staphylococci, aerobic coryneforms, micrococci and propionibacteria), and the yeast genus Malassezia. Results indicated that the axillary microflora was dominated by either staphylococcal or aerobic coryneform species. Comparisons between axillary bacterial numbers and levels of axillary odour demonstrated the greatest association between odour levels and the presence of aerobic coryneforms in the under-arm. As the taxonomy of cutaneous aerobic coryneforms is poorly understood, a further study was conducted to characterize selected axillary aerobic coryneform isolates. Using the molecular technique of 16S rDNA sequencing, selected genomic sequences of a number of axillary aerobic coryneform isolates were obtained. Comparisons with sequence databases indicated the likely presence of a range of Corynebacterium species on axillary skin, although the majority of isolates were most similar to either Corynebacterium G-2 CDC G5840 or C. mucifaciens DMMZ 2278. Although for a panel of individuals differences in the carriage of Corynebacterium species were noted, similar species were carried by a number of panellists. All isolates examined in this limited evaluation failed to demonstrate the capability to metabolize long-chain fatty acids (LCFAs) to shorter chain, more volatile products. The application of this modern molecular phylogenetic technique has increased understanding of the diversity of aerobic coryneform carriage in the axilla, and on human skin. The application of this technique in other studies to assess the ethnic differences in cutaneous bacterial ecology, or the effects on the microflora of specific product use, will assist in the future development of novel deodorant systems.     

Prebiotic Cosmetics: An Alternative to Antibacterial Products

The concept of prebiotics is well known from food products, where several ingredients are used to stimulate the beneficial gut microflora. However, prebiotic substances can in principle be applied to balance almost any microbial community to achieve advantageous effects. Since many cosmetic products target skin bacteria as a cause for undesirable conditions like inflammation or body odour, prebiotic actives can help to cope with these problems in a very effective way. To ensure a focused and successful mode of action we demonstrated that it is essential to analyse the microflora and the interactions between different bacterial species carefully. These analyses were made using a molecular method, fluorescence in situ hybridization (FISH), allowing a precise determination of a bacterial community without the drawbacks of classical culture methods. In a further step it was then possible to identify new actives that inhibit harmful or unwanted bacterial species while protecting the beneficial flora. Using a quick and reliable in vitro screening method we found several plant extracts (e.g. Ginseng or Black currant) which inhibit the inflammation‐causing bacterium Propionibacterium acnes, but do not affect beneficial species like coagulase‐negative staphylococci. In addition, it could also be demonstrated that a prebiotic product line for treatment of inflamed (or acne prone) skin showed the predicted effects on the microflora composition in a first in vivo trial, i.e. decrease of Propionibacterium acnes at a constant level of total bacteria. After 3 weeks, the microflora of 91% of the volunteers had been rebalanced in this way. Furthermore, these formulations have been shown to exhibit excellent skin compatibility compared to an antibacterial product. Thus, prebiotic substances have the potential to provide a gentle and sustainable alternative to undirected antibacterial ingredients which can cause irritation, and frequently only show short time effects. Paper presented at the IFSCC Conference 2005, Florence, Italy.     

Analysis of human male armpit sweat after fenugreek ingestion: Instrumental and sensory optimisation of the extraction method

In this study, we intend to develop a simple and selective procedure to extract compounds involved in the strong odour which appears after fenugreek ingestion (this odour is responsible of the so-called pseudo-maple syrup urine disease). Two procedures, solvent extraction and headspace solid-phase microextraction (HS-SPME), were employed to extract compounds from armpits sweat samples collected from two volunteer subjects. The HS-SPME extraction parameters were first optimised and then applied to extracts of armpit sweat collected from subjects after fenugreek ingestion. The sensory evaluation of the different extracts was carried by eight trained assessors; the HS-SPME and solvent extracts were, respectively, smelled by direct gas chromatography–olfactometry and direct olfaction. The results of sensory evaluation indicate that HS-SPME with a divinylbenzene/carboxen/polydimethylsiloxane (DVB/CAR/PDMS) fibre, of 50/30 μm film thickness and 2 cm length, gives a global odour close to that of sweat after ingestion of fenugreek infusion.     

Analysis of human male armpit sweat after fenugreek ingestion: Characterisation of odour active compounds by gas chromatography coupled to mass spectrometry and olfactometry

In this study, the strong “maple-syrup” odour which appears after fenugreek ingestion was investigated. Headspace solid-phase microextraction (HS-SPME) was applied to extract volatile odourant compounds from human male armpit sweat samples. Two male volunteers were considered who have similar diet; they had to ingest fenugreek infusion over the same period of time. The HS-SPME extracts obtained were then analysed by gas chromatography coupled either to mass spectrometry (GC–MS) or flame ionisation detection and olfactometry (GC–O). In that latter case, a panel of eight assessors was used, and the detection frequency methodology was applied. A total of 44 compounds could be identified in sweat samples, with a wide range of chemical structures, some of them being reported for the first time in human armpit sweat. Eight compounds appearing only after fenugreek ingestion could be identified: 2,5-dimethylpyrazine, β-pinene, 3-octen-2-one, camphor, terpinen-4-ol, 4-isopropyl-benzaldehyde, neryl acetate and β-caryophyllene. Due to their odourant notes, such compounds should be responsible for the strong “maple-syrup” odour present in sweat after fenugreek ingestion. GC–O confirmed the role of some odourant compounds in the “maple-syrup” odour of sweat, especially 2,5-dimethylpyrazine which was the best perceived odour. Among these eight compounds, some of them were previously reported in the fenugreek seeds, namely β-pinene, 3-octen-2-one and camphor.