androst-16-en-3-one and indole

The occurrence of boar taint in meat from uncastrated males may significantly affect the economics of pork production. The aim of this study was to analyse associations of four single nucleotide polymorphisms (SNPs) in the porcine CYP2E1 gene with skatole, indole, and androstenone levels in the Czech Large White-Czech Landrace commercial crossbreds. The SNPs were: g.2412C>T, c.1422C>T, c.1423G>A and c.*14G>T. Skatole, indole and androstenone levels were estimated by HPLC, and genotypes at the SNPs were determined by PCR-RFLP. SNPs c.1423G>A and c.*14G>T were in complete linkage disequilibrium. In boars, all SNPs were associated with the indole levels (P<0.05; P<0.01). There also were differences in the skatole levels in different genotypes, but these were not significant. No associations with androstenone levels were found. The associations of the SNPs with indole compounds should be studied in other commercial populations of boars to verify the favourable alleles and genotypes, with the prospect for their application in marker-assisted selection.

Topics: Adipose Tissue; Androstenes; Animals; Chromosome Mapping; Crosses, Genetic; Cytochrome P-450 CYP2E1; Female; Genetic Association Studies; Indoles; Linkage Disequilibrium; Male; Polymorphism, Single Nucleotide; Red Meat; Skatole; Sus scrofa

There is a commitment by the European pig sector to ban surgical castration of male piglets in the European Union in 2018. One alternative to castration is to raise entire male pigs, with an increased risk of boar taint. A field study was performed to: (1) evaluate inter- and intra-farm variation in boar taint prevalence, (2) investigate factors measured at slaughter influencing boar taint and (3) evaluate the relationship between sensorial scoring by a trained panel and the concentration of boar taint components. From 34 farms, neck fat samples were collected from all entire male pigs in at least two slaughter batches per farm (78 batches; 9167 animals). In addition to olfactory boar taint analysis, data were also collected on fresh skin lesions (score 0 to 3) at the slaughter line, slaughter weight, lean meat percentage, duration of transport, time spent in lairage, total delivery duration, day length, shortening of days and outdoor mean temperature. Using the hot iron method, neck fat samples were scored (eight-point scale) for boar taint. Average boar taint prevalence (score ≥3) was 5.6±2.5% and the mean difference between the maximum and minimum prevalence per farm was 4.3±3.2%. Androstenone (AND), skatole (SKA) and indole concentrations were measured for a subset (n=254) of the samples. According to binomial univariate mixed models, entire male pigs with a higher skin lesion score had higher odds of having boar taint (P=0.031), as did fatter entire male pigs (P<0.001). In the binomial multivariate mixed model lean meat percentage (P<0.001) and outdoor mean temperature (P=0.005) remained as only significant factors. Based on our results, we can conclude that these statistically significant at least partially influence the prevalence of boar taint. According to the binomial univariate mixed models SKA concentration in liquid fat seems a better predictor for boar taint than AND. There were no significant synergetic effects between boar taint compounds.

Topics: Abattoirs; Adipose Tissue; Androstenes; Animal Husbandry; Animals; Belgium; Indoles; Male; Meat; Skatole; Smell; Sus scrofa

Tannins have long been considered 'anti-nutritional' factors in monogastric nutrition, shown to reduce feed intake and palatability. However, recent studies revealed that compared with condensed tannins, hydrolysable tannins (HT) appear to have far less impact on growth performance, but may be inhibitory to the total activity of caecal bacteria. This in turn could reduce microbial synthesis of skatole and indole in the hindgut of entire male pigs (EM). Thus, the objective of this study was to determine the impact of a group of dietary HT on growth performance, carcass traits and boar taint compounds of group housed EM. For the study, 36 Swiss Large White boars were assigned within litter to three treatment groups. Boars were offered ad libitum one of three finisher diets supplemented with 0 (C), 15 (T15) or 30 g/kg (T30) of HT from day 105 to 165 of age. Growth performance, carcass characteristics, boar taint compounds in the adipose tissue and cytochrome P450 (CYP) isoenzymes CYP2E1, CYP1A2 and CYP2A19 gene expression in the liver was assessed. Compared with C, feed efficiency but not daily gain and daily feed intake was lower (P<0.05) in T15 and T30 boars. Except for the percentage carcass weight loss during cooling, which tended (P<0.10) to be greater in T30 than C and T15, carcass characteristics were not affected by the diets. In line with the numerically lower androstenone level, bulbourethral and salivary glands of T30 boars were lighter (P<0.05) than of T15 with intermediate values for C. Indole level was lower (P<0.05) in the adipose tissue of T30 than C pigs with intermediate levels in T15. Skatole levels tended (P<0.10) to be lower in T30 and C than T15 pigs. Hepatic gene expression of CYP isoenzymes did not differ between-treatment groups, but was negatively correlated (P<0.05) with androstenone (CYP2E1 and CYP1A2), skatole (CYP2E1, CYP2A) and indole (CYP2A) level. In line with the numerically highest androstenone and skatole concentrations, boar taint odour but not flavour was detected by the panellists in loins from T15 compared with loins from C and T30 boars. These results provide evidence that HT affected metabolism of indolic compounds and androstenone and that they affected the development of accessory sex glands. However, the effects were too small to be detected by sensory evaluation.

Topics: Adipose Tissue; Androstenes; Animals; Bulbourethral Glands; Cytochrome P-450 Enzyme System; Diet; Hydrolyzable Tannins; Indoles; Male; Odorants; Phenotype; Red Meat; Salivary Glands; Skatole; Swine

Boar taint is an off-odour that entails negative consumer reactions. In this study two extraction and UHPLC-HRMS analysis methods, valuable for evaluation of consumer acceptance towards boar meat, were developed for quantification of indole, skatole, and androstenone in different meat products. Sample pretreatment consisted of extraction with methanol and a homogenising step (cooked ham, minced meat, tenderloin, bacon, cutlets, blade loin, uncooked ham) or a melting step (salami sausage and liver paste). Both methods were validated according to CD 2002/657/EC and ISO 17025 guidelines. Good performance characteristics were obtained. Good linearity (R(2) ⩾ 0.99) and no lack of fit was observed (95% confidence interval; F-test, p > 0.05). Also good recovery (89-110%) and satisfactory precision: repeatability (RSD ⩽ 14.9%) and within-laboratory reproducibility (RSD ⩽ 17.2%) were obtained. Analysis of cooked ham and salami sausage samples proved the applicability of both methods for routine analysis.

Topics: Androstenes; Animals; Chromatography, High Pressure Liquid; Indoles; Limit of Detection; Male; Mass Spectrometry; Meat Products; Red Meat; Skatole; Swine

This exploratory study investigated the variability of fatty acid composition in entire male pigs with extremely divergent levels of boar taint compounds. Fatty acids were quantified in back fat samples from 20 selected carcasses of Pietrain*F1 sired boars (average carcass weight 84 kg) with extremely low (LL) or extremely high (HH) levels of androstenone, skatole, and indole. Concentrations of polyunsaturated fatty acids (PUFA) were significantly (p<0.05) increased in LL boars (23.4%) compared to HH boars (19.7%). This was mainly due to increased levels of linoleic acid (C18:2 n-6) and α-linolenic acid (C18:3 n-3). Correspondingly, unsaturated fatty acids (SFA) were significantly lower (p<0.05) in LL boars (35.2%) compared to HH boars (37.7%). The findings are discussed with respect to potential effects on flavor formation in boar fat and meat. Further research is needed to study the gender specificity and the interplay of the synthesis and the metabolism of steroids, lipids, and the clearance of skatole in pigs.

Topics: alpha-Linolenic Acid; Androstenes; Animals; Breeding; Fatty Acids, Unsaturated; Humans; Indoles; Linoleic Acid; Male; Meat; Odorants; Skatole; Subcutaneous Fat; Swine; Taste

Boar taint in carcasses may vary between farms and abattoirs, although the underlying mechanisms are not yet fully understood. In the present study, 169 boars from three farms were split into two groups and slaughtered at two abattoirs. Duration of transport and the time between arrival at the abattoir and unloading (pre-unloading time) were recorded. During slaughter, blood, feces, and urine were collected to measure testosterone and cortisol levels. Carcasses were classified according to the number of skin lesions, and fat samples were taken to determine skatole, indole and androstenone levels. Androstenone in fat and testosterone in blood, feces, and urine were mainly influenced by the duration of transport. Skatole and indole concentrations were increased by both pre-unloading time and duration of transport, but were also related to skin lesions. Thus it is concluded that androstenone and skatole concentrations in fat are significantly modified by pre-slaughter conditions.

Topics: Abattoirs; Adipose Tissue; Androstenes; Animals; Humans; Hydrocortisone; Indoles; Male; Meat; Odorants; Skatole; Skin; Stress, Physiological; Swine; Testosterone; Transportation

Boar taint is an off-odour occurring while heating meat or fat from boars. A method detecting the three compounds (androstenone, skatole and indole) simultaneously in blood would offer substantial advantages since it would allow monitoring the impact of rearing strategies. Therefore, a UHPLC-HR-Orbitrap-MS analysis method is optimized and validated for the quantification of these compounds in plasma or serum. Sample pre-treatment involved an extraction with diethylether followed by a centrifugal filtration (30 kDa). Limits of detection and quantification varied between 0.5 and 1 μg L(-1) and 2 and 3 μg L(-1) for the three compounds, respectively. Besides, an excellent repeatability (RSD < 7.6%), within-laboratory reproducibility (RSD<10.5%), recovery (87-97%) and linearity (R(2)>0.99) were recorded. Correlations between serum/plasma and fat levels of the boar taint compounds were positive for skatole (r(serum) = 0.39 and r(plasma) = 0.84) and androstenone (r(serum) = 0.73-0.78 and r(plasma) = 0.32-0.80).

Topics: Androstenes; Animals; Chromatography, High Pressure Liquid; Indoles; Male; Mass Spectrometry; Meat; Odorants; Skatole; Swine

An association study between polymorphisms of six genes and boar taint related compounds androstenone, skatole and indole was performed in a boar population (n=370). Significant association (P<0.05) was detected for SNP of FMO5 (g.494A>G) with all boar taint compounds, SNP of CYP21 (g.3911T>C) with skatole and indole, and SNP of ESR1 (g.672C>T) with androstenone and indole. mRNA expression of CYP21 and ESR1 was higher in CAB (castrated boar) compared to non-castrated boars; whereas, the expression of FMO5 and ESR1 was higher in LBT (low boar taint) compared to HBT (high boar taint) in liver tissue. FMO5, CYP21 and ESR1 proteins were less detectable in HBT compared with LBT and CAB in liver tissues. These findings suggest that FMO5, CYP21 and ESR1 gene variants might have effects on the boar taint compounds.

Topics: Androstenes; Animals; Estrogen Receptor alpha; Food Quality; Genotyping Techniques; Indoles; Liver; Male; Meat; Oxygenases; Phenotype; Polymorphism, Single Nucleotide; RNA, Messenger; Skatole; Steroid 21-Hydroxylase; Sulfotransferases; Swine

Societal pressure to ban surgical castration of male piglets is rising due to animal welfare concerns, thus other methods to prevent boar taint need to be explored. Genetic selection against boar taint appears to be a long-term sustainable alternative. However, as boar taint is linked to reproductive hormones, it is important to consider possible negative side effects such as delayed sexual maturity or changes in behaviour. We reported earlier that the melanocortin-4 receptor (MC4R) marker can be used to reduce boar taint levels in fat of boars. The objective of this study was to evaluate whether MC4R marker-assisted selection for lower boar taint prevalence affects plasma levels of boar taint compounds and testosterone; sexual maturity; behaviour; skin lesions; and lameness in boars and gilts. Using an intervention study with a 2×2 design, 264 boars and gilts differing on position 893 of the MC4R gene (AA v. GG) were compared. The MC4R polymorphism did not affect the plasma concentration of either androstenone or testosterone at different time points, whereas the concentration of skatole was significantly lower (P=0.003) and the concentration of indole tended to be lower (P=0.074) in GG compared with AA boars. A higher percentage of gilts of the GG genotype were in puberty at slaughter age compared with AA gilts (P<0.001). The age of the boars at sexual maturity (as indicated by the first positive preputial smear test) did not differ between AA and GG boars. In contrast, weight of GG boars at sexual maturity tended to be lower (P=0.065). During the period from 6 weeks of age to slaughter, boars and gilts of the GG genotype showed more playing behaviour (P=0.015) and less passive and feeding behaviour (P=0.003). They showed more skin lesions on their back and caudal area (P=0.022), and tended to show more skin lesions on their head and anterior area (P=0.093) compared with AA animals. In conclusion, the polymorphism in the MC4R gene can be used as a marker without negative effects on reproduction characteristics in boars and gilts. Genetic selection towards a lower prevalence of boar taint will lead to more active pigs with more skin lesions. Management strategies may therefore be necessary to reduce skin lesions in the selected animals.

Topics: Androstenes; Animal Welfare; Animals; Body Weight; Female; Genotype; Indoles; Male; Polymorphism, Genetic; Receptor, Melanocortin, Type 4; Reproduction; Sexual Maturation; Skatole; Swine; Testosterone

This work outlines an optimized gas chromatrography-mass spectrometry (GC-MS) based protocol for screening of the presence of the three boar-taint-producing compounds indole, skatole (3-methylindole), and androstenone (5α-androst-16-en-3-one) in porcine fat. The study shows that an accuracy suitable for sample screening can be achieved even when speed and ease of sample handling is prioritized and without the use of internal standards. The method provides levels of detection of 82 ng/g for indole, 97 ng/g for skatole, and 623 ng/g for androstenone (in a 2 g natural backfat matrix). A least-squares approach of predicting the sample analyte mass from instrument response is shown to have prediction errors (root-mean-square error) of 96 ng/g for indole, 94 ng/g for skatole, and 331.3 ng/g for androstenone. The method is intended for discrimination of samples in risk groups, and a scheme is proposed to turn the GC-MS response of the boar-taint analytes into risk classes.

Topics: Androstenes; Animals; Fats; Food Contamination; Gas Chromatography-Mass Spectrometry; Indoles; Molecular Structure; Skatole; Swine

A reduction in boar taint, an unpleasant odor arising in pork from some intact males, is desirable if routine castration of piglets needs to be stopped. Commercial slaughter pigs are typically crosses between sire lines mainly selected for finishing traits and dam lines mainly selected for reproduction traits. Previous studies suggest the possibility of reducing boar taint in sire lines due to favorable genetic correlations between boar taint and finishing traits. However, there are indications of unfavorable genetic associations between boar taint and female reproduction traits, but a lack of genetic correlation estimates remain a major roadblock in reducing boar taint in dam lines. This study was conducted to estimate genetic correlations between boar taint traits and female reproduction traits, investigate differences in these genetic relationships among sire and dam lines, and evaluate possible consequences of selection against boar taint in dam lines. The data consisted of 32,549 reproduction records from a Landrace dam line, 23,874 records from a Yorkshire dam line, and 3,745 records from a Pietrain sire line. Androstenone, skatole, and indole were measured on 1,896 carcasses, and human nose scores were recorded on 7,742 carcass samples. In general, the level of boar taint was significantly greater (P < 0.05) in the two dam lines than in the sire line. A majority of genetic correlations of boar taint compounds with reproduction traits were either low or nonsignificant, except for those of skatole and indole, with age at first insemination in dam lines that were -0.32 and -0.46, respectively. Genetic correlations also differed (P < 0.05) between sire and dam lines. The consequences of selection against boar taint in dam lines were evaluated, using selection indexes based on reproduction traits only, boar taint traits only, and both boar taint and reproduction traits. Selection on an index of only reproduction traits increased the number of carcasses with boar taint from 4 to 7.3% in 5 generations. Selection on a combined index reduced carcasses with boar taint from 4 to <0.1% in 19 generations at the cost of a 10% less economic gain in reproduction traits. In markets for intact males, overall economic gain is 78% greater than with selection for reproduction only. Breeding programs for boar taint in commercial production should include boar taint in breeding goals of dam lines as the levels of boar taint and the risk of a further increase are greater.

Topics: Androstenes; Animals; Breeding; Chromatography, High Pressure Liquid; Chromatography, Liquid; Female; Indoles; Male; Mass Spectrometry; Meat; Odorants; Quantitative Trait, Heritable; Selection, Genetic; Skatole; Sus scrofa

The steroidal pig pheromones androstenone (5α-androst-16-en-3-one), 3α-androstenol (5α-androst-16-en-3α-ol), and 3β-androstenol (5α-androst-16-en-3β-ol) as well as the heterocyclic aromatic amines skatole and indole, originating from microbial degradation of tryptophan in the intestine of pigs, are frequently recognized as the major compounds responsible for boar taint. A new procedure, applying stable isotope dilution analysis (SIDA) and headspace solid-phase microextraction-gas chromatography/mass spectrometry (HS-SPME-GC/MS) for the simultaneous quantitation of these boar taint compounds in pig fat was developed and validated. The deuterated compounds androstenone-d(3), 3β-androstenol-d(3), skatole-d(3), and indole-d(6) were synthesized and successfully employed as internal standards for SIDA. The new procedure is characterized by a fast, simple, and economic sample preparation: methanolic extraction of the melted fat followed by a freezing and an evaporation step allows for extraction and enrichment of all five analytes. Additional time-consuming cleanup steps were not necessary, as HS-SPME sampling overcomes fat-associated injector and column contamination. The method has been validated by determining intra- and interday precision and accuracy as well as the limit of detection (LOD) and limit of quantitation (LOQ). Additionally, a cross-validation for androstenone, skatole, and indole was carried out comparing the results of 25 back fat samples obtained simultaneously by the new SIDA-HS-SPME-GC/MS procedure with those obtained in separate GC/MS and high-performance liquid chromatography fluorescence detection (HPLC-FD) measurements. The cross-validation revealed comparable results and confirms the feasibility of the new SIDA-HS-SPME-GC/MS procedure.

Topics: Adipose Tissue; Androstenes; Androstenols; Animals; Calibration; Deuterium; Gas Chromatography-Mass Spectrometry; Indoles; Isotope Labeling; Skatole; Solid Phase Microextraction; Swine

CYP2A6 is one of the enzymes involved in the hepatic metabolism of a naturally produced compound, skatole, in the pig. Low CYP2A6 activity has been linked to excessive accumulation of skatole in pig adipose tissue and development of the phenomenon "boar taint." CYP2A6 activity varies between male and female animals, suggesting the involvement of sex hormones in regulation of the enzyme activity and/or expression. The present study investigated whether pig hepatic CYP2A6 protein expression is regulated by the testicular steroids testosterone, androstenone, or estrone sulfate using primary cultured hepatocytes as a model system. The study has also examined whether CYP2A6 expression can be modulated by the boar taint compounds skatole and indole. The research has established that androstenone inhibits CYP2A6 protein expression at the concentration of 1, 10, and 100 nM by 55, 37, and 44%, respectively. In contrast to androstenone, skatole and indole (final concentrations, 1, 10, and 100 nM) had a stimulatory effect on CYP2A6 expression. The effect of indole was more pronounced than that of skatole (maximum induction by 145 and 70%, respectively). Estrone sulfate and testosterone did not have a significant effect on CYP2A6 protein level. This is, as far as we know, the first communication to report the regulation of pig hepatic CYP2A6 expression by steroids and boar taint compounds. The hormonal modulation of CYP2A6 expression might contribute to gender-related differences in pig hepatic CYP2A6 activity and skatole accumulation in pig adipose tissue.

Topics: Androstenes; Animals; Aryl Hydrocarbon Hydroxylases; Blotting, Western; Cells, Cultured; Cytochrome P-450 CYP2A6; Dose-Response Relationship, Drug; Down-Regulation; Electrophoresis, Polyacrylamide Gel; Estrone; Hepatocytes; Indoles; Male; Mixed Function Oxygenases; Skatole; Swine; Testis; Testosterone; Testosterone Congeners; Up-Regulation

Boar taint in entire male pigs remains a problem for fresh pork production. Since castration of pigs will be prohibited in the future on animal welfare reasons, attempts are made to detect boar taint pre and post mortem. Post mortem techniques focus on simultaneous quantification of the three boar taint substances by one simple and reliable method. In this study a liquid chromatographic multiple mass spectrometric (LC-MS(n)) method has been developed and validated for the simultaneous determination of indole (2,3-benzopyrrole, ID), skatole (3-methylindole, SK) and androstenone (5alpha-androst-16-en-3-one, AEON) in pig fat samples. Sample preparation was kept as short as possible, since a single extraction method for structurally different indols and steroids was seeked after. Analytes were extracted from the fat matrix by methanol and clean-up consisted of freezing the extract in liquid nitrogen followed by a filtration step. The analytes were chromatographically separated on a Symmetry C(18) column. Recoveries for ID, SK and AEON, as calculated from fortified fat samples using 2-methylindole (2-MID) as internal standard, were 96, 91 and 104%, respectively. However, matrix interferences were encountered determining the androstenone levels in fat. Linearity, determined in fat samples, was in the range of 50-1600 microg kg(-1) for the indolic compounds and 125-2000 microg kg(-1) for the steroid AEON. The correlation coefficients (R(2)) of the calibration curves were 0.998 for ID, 0.997 for SK and 0.810 for AEON.

Topics: Androstenes; Animals; Calibration; Chromatography, Liquid; Indoles; Lipids; Mass Spectrometry; Reproducibility of Results; Skatole; Swine

A rapid high-performance reversed-phase liquid chromatographic method for the simultaneous quantitative determination of the main boar taint compounds androstenone, skatole (3-methylindole) and indole, in back fat from pigs has been developed. The compounds are extracted by a simple homogenisation of adipose tissue in methanol; interfering lipids are removed by precipitation after cooling and centrifugation. Androstenone is derivatized pre-column with dansylhydrazine (5 min at ambient temperature) using BF3 as catalyst. The compounds are separated on a 60 x 4.6 mm I.D., 3 microns Hypersil ODS column (Hewlett-Packard) using a step-gradient; total time for the separation is 15 min. Fluorescence is used for selective detection. The limit of quantitation for indole and skatole is 30 ng/g and for androstenone 200 ng/g back fat. The results for skatole obtained by the present method were compared with those of colorimetric method, while androstenone determinations were compared with two GC-MS methods and a RIA method. The correlations observed were in the range of 0.946-0.993. The average contents of androstenone and skatole were 640 +/- 700 and 78 +/- 113 ng/g (n = 1162 male pigs), respectively.

Topics: Adipose Tissue; Androstenes; Animals; Chromatography, High Pressure Liquid; Colorimetry; Gas Chromatography-Mass Spectrometry; Indoles; Male; Radioimmunoassay; Reproducibility of Results; Skatole; Spectrometry, Fluorescence; Swine