Serum metabolic profiling in pregnant Holstein cows 3 weeks prior to parturition using two-dimensional gas chromatography time-of-flight mass spectrometry.

This preliminary study explored potential serum biomarkers for predicting the onset of milk fever (MF), a bovine parturient disease with hypocalcemia. We conducted two-dimensional gas chromatography mass spectrometry-based metabolomics in 8 and 17 pregnant Holstein cows that did and did not develop MF 3 weeks later, respectively. In principal component analysis (PCA) applied to a dataset containing 1498 metabolites, serum metabolites exhibited highly similar chemical profiles between cows with and without MF. PCA with a limited dataset of metabolites containing fatty acids, which had significantly different values between the groups and/or correlation coefficients of >0.5 for the serum calcium concentration, distinguished the two groups. These suggest the possibility of developing serum biomarkers for predicting bovine MF.

Sprayed Urine Emits a Pungent Odor due to its Increased Adhesion to Vertical Objects via Urinary Proteins Rather Than to Changes in its Volatile Chemical Profile in Domestic Cats.

Spraying urine on vertical objects by raising the tail is a commonly observed functional behavior for chemical communication in Felidae species, including domestic cats (Felis silvestris catus). The sprayed urine is recognized as a chemical signal for territorial ownership of their habitats. Previous studies reported that sprayed urine emits a more pungent odor than urine excreted from a squatting position. However, little is known about how sprayed urine acts as a strong scent mark in the environment. Here, we showed that sprayed urine originates only from bladder urine without any secretions, such as anal sac secretions, but it can effectively emit volatile organic compounds (VOCs) when smeared on vertical objects due to its strong adhesion. Chemical profiles of VOCs and odor qualities were similar between fresh sprayed urine and bladder urine sampled immediately after spraying from the same individuals. Meanwhile, feline-specific proteinuria arising from excretion of a carboxylesterase that produces a precursor of cat-specific odorants resulted in reduced surface tension of the urine and increased adhesion to vertical surfaces, which kept sprayed urine on the surfaces and led to the emission of large amounts of VOCs. In conclusion, proteinuria contributes to the emission of a strong odor through its enhanced adhesion to vertical objects without other secretions containing malodorous substances. These findings improve our understanding of the mechanism of scent marking via the spraying of urine for chemical communication in cats.

Development of the gas chromatography/mass spectrometry-based aroma designer capable of modifying volatile chemical compositions in complex odors.

Many volatile organic compounds (VOCs) are used to produce various commercial products with aromas mimicking natural products. The VOCs responsible for aromas have been identified from many natural products. The current major strategy is to analyze chemical compositions and aroma qualities of individual VOCs using gas chromatography/mass spectrometry (GC/MS) and GC-olfactometry. However, such analyses cannot determine whether candidate VOCs contribute to the characteristic aroma in mixtures of many VOCs. In this study, we developed a GC/MS-based VOC collection/omission system that can modify the VOC compositions of samples easily and rapidly. The system is composed of GC/MS with a switching unit that can change gas flow routes between MS and a VOC collection device. We first applied this system to prepare gas samples for omission tests, and the aroma qualities of VOC mixtures with and without some VOCs were evaluated by panelists. If aroma qualities were different between the 2 samples, the omitted VOCs were likely key odorants. By collecting VOCs in a gas bag attached to the collection device and transferring some VOCs to MS, specific VOCs could be omitted easily from the VOC mixture. The system could prepare omission samples without chemical identification, preparation of each VOC, and laborious techniques for mixing VOCs, thus overcoming the limitations of previous methods of sample preparation. Finally, the system was used to prepare artificial aromas by replacing VOC compositions between different samples for screening of key odorants. In conclusion, the system developed here can improve aroma research by identifying key odorants from natural products.

Assessing the safety and suitability of using silver vine as an olfactory enrichment for cats.

Olfactory enrichment is a strategy that can improve welfare among animals managed in captivity, such as household domestic cats. Catnip (Nepeta cataria) and silver vine (Actinidia polygama) that produce iridoids are used as olfactory enrichments for cats, but little is known about the safety or the best plant resources to use that maximize positive cat responses. We report physiological effects and suitable harvest and drying methods for using silver vine as olfactory enrichment. Continuous exposure of cats to silver vine showed no hallmarks of addictive behavior, while blood indicators of stress and hepatic or renal injury showed no increase in cats stimulated with it. Drying the leaves changed the iridoid profile, enhancing the feline response. In conclusion, dried silver vine leaves are the most suitable resource for developing olfactory enrichment that maximizes feline typical response, which would not result in dependence, stress, or toxicity to the liver or kidneys in cats.

Cold stress induces malformed tomato fruits by breaking the feedback loops of stem cell regulation in floral meristem.

Fruit malformation is a major constrain in fruit production worldwide resulting in substantial economic losses. The farmers for decades noticed that the chilling temperature before blooming often caused malformed fruits. However, the molecular mechanism underlying this phenomenon is unclear. Here we examined the fruit development in response to cold stress in tomato, and demonstrated that short-term cold stress increased the callose accumulation in both shoot apical and floral meristems, resulting in the symplastic isolation and altered intercellular movement of WUS. In contrast to the rapidly restored SlWUS transcription during the recovery from cold stress, the callose removal was delayed due to obstructed plasmodesmata. The delayed reinstatement of cell-to-cell transport of SlWUS prevented the activation of SlCLV3 and TAG1, causing the interrupted feedback inhibition of SlWUS expression, leading to the expanded stem cell population and malformed fruits. We further showed that the callose dynamics in response to short-term cold stress presumably exploits the mechanism of bud dormancy during the seasonal growth, involving two antagonistic hormones, abscisic acid and gibberellin. Our results provide a novel insight into the cold stress regulated malformation of fruit.

Domestic cat damage to plant leaves containing iridoids enhances chemical repellency to pests.

Catnip (Nepeta cataria) and silver vine (Actinidia polygama) produce iridoids with arthropod-repellent effects. Cats rub and roll against these plants, transferring iridoids to their fur that repels mosquitoes. Cats also lick and chew plant leaves during this response, although the benefit of this additional behavior has remained unknown. Here, we show that feline leaf damage substantially increases iridoid emission from both plants while also diversifying iridoids in silver vine. Cats show an equivalent duration of response to the complex cocktail of iridoids in damaged silver vine and to the much higher level of a single iridoid produced by damaged catnip. The more complex iridoid cocktail produced when silver vine is licked and chewed by cats increases mosquito repellency at low concentration. In conclusion, feline leaf damage contributes by releasing more mosquito-repellent iridoids. Feline olfactory and behavioral sensitivity is fine-tuned to plant-specific iridoid production for maximizing the mosquito repellency gained.

The characteristic response of domestic cats to plant iridoids allows them to gain chemical defense against mosquitoes.

Domestic cats and other felids rub their faces and heads against catnip (Nepeta cataria) and silver vine (Actinidia polygama) and roll on the ground as a characteristic response. While this response is well known, its biological function and underlying mechanism remain undetermined. Here, we uncover the neurophysiological mechanism and functional outcome of this feline response. We found that the iridoid nepetalactol is the major component of silver vine that elicits this potent response in cats and other felids. Nepetalactol increased plasma ß-endorphin levels in cats, while pharmacological inhibition of µ-opioid receptors suppressed the classic rubbing response. Rubbing behavior transfers nepetalactol onto the faces and heads of respondents where it repels the mosquito, Aedes albopictus Thus, self-anointing behavior helps to protect cats against mosquito bites. The characteristic response of cats to nepetalactol via the µ-opioid system provides an important example of chemical pest defense using plant metabolites in nonhuman mammals.

Detection of urinary luteinizing hormone in Japanese black cows after administration of gonadotropin-releasing hormone.

The blood luteinizing hormone (LH) surge in cows is well studied. However, little is known about urinary LH in cows. This study examined urinary LH concentrations after administration of gonadotropin-releasing hormone (GnRH) in six Japanese black cows to induce LH secretion from the pituitary gland into the bloodstream. Abrupt rises in plasma and urinary LH were observed after GnRH administration. Plasma and urinary LH peaked at 2 and 5 hr, respectively. A positive correlation was observed between plasma LH concentrations and urinary LH amounts. Ovulation was confirmed in the cows after 48 hr of GnRH administration. These data strongly suggest that urinary LH is derived from plasma LH, which triggers ovulation in cows.

LC-ESI-MS/MS quantification of carnosine, anserine, and balenine in meat samples.

The histidine-containing imidazole dipeptide carnosine and its methylated analogs anserine and balenine are present at high concentrations in vertebrate tissues. Although the physiological functions of the imidazole dipeptides have not been elucidated yet, it has been suggested that they play significant biological roles in animals. Despite increasing interest, few studies have challenged the quantifications of carnosine, anserine, and balenine by a single HPLC run because they have similar retention times. In this study, we developed a method to quantify these imidazole dipeptides in meat samples using an LC-ESI-MS/MS triple-quadrupole mass spectrometer. We improved the liquid chromatographic separation of the imidazole dipeptides by applying a mix-mode column, which provides both normal phase and ion exchange separations, and developed multiple reaction-monitoring of the transitions for quantification of m/z 227 → 110 for carnosine, m/z 241 → 126 for anserine, m/z 241 → 124 for balenine, and m/z 269 → 110 for L-histidyl-L-leucine (internal standard). The established method met all pre-defined validation criteria. Intra- and inter-day accuracy and precision were ±10.0% and ≤14.8%, respectively. The ranges of quantifications were 14.7 ng/mL to 1.5 mg/mL for carnosine, 15.6 ng/mL to 1.6 mg/mL for anserine, and 15.6 ng/mL to 1.6 mg/mL for balenine. In conclusion, the validated method was successfully applied to the quantification of imidazole dipeptides in biological samples without derivatization.