A most aggressive bear: Safari videos document sloth bear defense against tiger predation.

Sloth bears are non-carnivorous yet they attack more people than any other bear. They often stand up and charge explosively if a person mistakenly gets too close. Here, we argue that their aggression toward humans is an extension of their behavior toward tigers, which are their only natural predator. Interactions between sloth bears and tigers have not previously been studied because scientists have rarely observed such events. We collected and examined 43 videos or photo documentations of sloth bear-tiger interactions posted on the internet or social media from 2011 to 2023, mainly by tourists visiting tiger parks in India. We observed that sloth bears were most likely to stand up and charge if they first became aware of the tiger at close range (<3 m away). This aggressive-defensive strategy, intended to dissuade the tiger from attacking, appeared to be successful, in that 86% of interactions ended with no contact, whereas four (9%) culminated in the bear's death. We propose that a myrmecophagous diet led to this species' aggressive behavior: (1) their long, blunt front claws, well adapted for digging termites and ants, hamper their ability to climb trees for escape, and (2) they walk with their head down focused on scents underground, and make considerable noise digging and blowing soil, enabling tigers to approach quite closely without being detected. Sloth bears have coexisted with tigers or other (now extinct) large felid predators for their entire evolutionary history. Whereas their aggressive behavior has served them well for millions of years, more recently, people's fear of and retaliation against sloth bears represents a major threat to their survival. Understanding how sloth bears react to tigers provides guidance for reducing attacks on humans, thereby contributing to sloth bear conservation. Our investigation was made possible by passive citizen scientists, who unknowingly collected valuable data.

Sloth bear attacks: regional differences and safety messaging.

Sloth bears behave aggressively toward humans when threatened and are among the most dangerous wildlife in India. Safety messaging for those who live in sloth bear country must be accurate to be effective, and messaging may need to be modified to account for regional differences in human-bear relationships. The timing of sloth bear attacks on the Deccan Plateau of Karnataka, both by season and by time of day, deviated enough from those reported in other areas such that it warranted further investigation. We compared data from eight studies of human-sloth bear conflict from across the Indian subcontinent and explored possibilities as to why differences exist. Seasonally all studies reported that human-sloth bear conflict was highest when human activity in the forest was greatest, though the season of highest human activity varied significantly by region (χ2 = 5921, df = 5, P < 0.001). The time of day that the majority of attacks occurred also varied significantly by region (χ2 = 666, df = 5, P < 0.001), though human activity was relatively consistent. We speculated that the rate of day attacks on the Deccan Plateau was lower due to the reduced probability of encountering a sleeping bear as they are concealed and secure in shallow caves. Additionally, the rate of attacks was significantly higher at night on the Deccan Plateau because people often to work into nighttime. We concluded that slight differences, or different emphasis, to bear safety messaging may be necessary on a regional basis to keep the messaging accurate and effective.

Development of Ecom50 and retention index models for nontargeted metabolomics: identification of 1,3-dicyclohexylurea in human serum by HPLC/mass spectrometry.

The goal of many metabolomic studies is to identify the molecular structure of endogenous molecules that are differentially expressed among sampled or treatment groups. The identified compounds can then be used to gain an understanding of disease mechanisms. Unfortunately, despite recent advances in a variety of analytical techniques, small molecule (<1000 Da) identification remains difficult. Rarely can a chemical structure be determined from experimental "features" such as retention time, exact mass, and collision induced dissociation spectra. Thus, without knowing structure, biological significance remains obscure. In this study, we explore an identification method in which the measured exact mass of an unknown is used to query available chemical databases to compile a list of candidate compounds. Predictions are made for the candidates using models of experimental features that have been measured for the unknown. The predicted values are used to filter the candidate list by eliminating compounds with predicted values substantially different from the unknown. The intent is to reduce the list of candidates to a reasonable number that can be obtained and measured for confirmation. To facilitate this exploration, we measured data and created models for two experimental features; MS Ecom50 (the energy in electronvolts required to fragment 50% of a selected precursor ion) and HPLC retention index. Using a data set of 52 compounds, Ecom50 models were developed based on both Molconn and CODESSA structural descriptors. These models gave r² values of 0.89 to 0.94 depending on the number of inputs, the modeling algorithm chosen, and whether neutral or protonated structures were used. The retention index model was developed with 400 compounds using a back-propagation artificial neural network and 33 Molconn structure descriptors. External validation gave a v² = 0.87 and standard error of 38 retention index units. As a test of the validity of the filtering approach, the Ecom50 and retention index models, along with exact mass and collision induced dissociation spectra matching, were used to identify 1,3-dicyclohexylurea in human plasma. This compound was not previously known to exist in human biofluids and its elemental formula was identical to 315 other candidate compounds downloaded from PubChem. These results suggest that the use of Ecom50 and retention index predictive models can improve nontargeted metabolite structure identification using HPLC/MS derived structural features.

Calculated carbon-hydrogen bond dissociation enthalpies for predicting oxidative susceptibility of drug substance molecules.

The carbon-hydrogen bond dissociation enthalpy (BDE) concept is evaluated as a potential computed indicator of stability of pharmaceutical drug substance candidates - specifically for oxidative stability of these molecules. Computational methods are discussed. Accuracy and validity of the methods are evaluated. BDEs are computed for several well-known molecules, for which stability and degradant identification information is known. Anecdotal correlations are noted between the lowest BDE energies of familiar molecules (sertraline, ezlopitant and related structures, ziprasidone, trovafloxacin, and varenicline), the sites of oxidative lability on these molecules and the identities of oxidative degradants. A low BDE may correlate in general with a reactive site on a molecule, not just an oxidatively susceptible one.

Method for determining the average degree of substitution of o-vanillin derivatized porcine somatotropin.

Electrospray mass spectral observation directly on a sample of a derivatized protein, such as porcine somatotropin (pST), affords a method for evaluating the degree of substitution of this protein. Derivatization of the lysine residues and the terminal amino residue here by formation of a Schiff base with a small aromatic aldehyde (in this case, o-vanillin) affords stabilization of the protein so that it may be used in a controlled release veterinary pharmaceutical formulation. This method permits direct observation of substitutions, optimization of manufacturing procedures for producing a commercial product, and permits quality evaluation of material.

Identification of low-level degradants from low dose tablets.

A multifaceted approach was successfully used to identify three of four unknown degradants in degraded low dose tablets. Accelerated solvent extraction (ASE) was found to be an invaluable tool in this multifaceted approach. ASE was capable of extracting four individual degradants of an active pharmaceutical component from 10 tablets into 15 mL of solvent with approximately 100% recovery for each degradant. Using ASE instead of manual extraction led to the extraction and isolation of the degradants in 1 day instead of 7 days. One of the degradants was extracted by ASE, isolated by semi-prep HPLC, and identified by LC-MS and NMR spectroscopy. The structures of two of the remaining three degradants were confirmed by synthesis of authentic samples, while the fourth degradant is yet to be identified.

Pharmaceutical impurity identification: a case study using a multidisciplinary approach.

A multidisciplinary team approach to identify pharmaceutical impurities is presented in this article. It includes a representative example of the methodology. The first step is to analyze the sample by LC-MS. If the structure of the unknown impurity cannot be conclusively determined by LC-MS, LC-NMR is employed. If the sample is unsuitable for LC-NMR, the impurity needs to be isolated for conventional NMR characterization. Although the technique of choice for isolation is preparative HPLC, enrichment is often necessary to improve preparative efficiency. One such technique is solid-phase extraction. For complete verification, synthesis may be necessary to compare spectroscopic characteristics to those observed in the original sample. Although not widely practiced, an effective means of getting valuable structural information is to conduct a degradation study on the purified impurity itself. This systematic strategy was successfully applied to the identification of an impurity in the active pharmaceutical ingredient 1-(1,2,3,5,6,7-hexahydro-s-indacen-4-yl)-3-[4-(1-hydroxy-1-methyl-ethyl)-furan-2-sulphonylurea. Identification required the use of all of the previously mentioned techniques. The instability of the impurity under acidic chromatographic conditions presented an additional challenge to purification and identification. However, we turned this acidic instability to an advantage, conducting a degradation study of the impurity, which provided extensive and useful information about its structure. The following discussion describes how the information gained from each analytical technique was brought together in a complementary fashion to elucidate a final structure.

Soft ionization mass spectrometry of chromanols produces radical cation molecular ions.

Several chromanol drug substance candidates exhibit unconventional behavior under the soft ionization conditions of fast atom bombardment and electrospray ionization in the mass spectrometer. Under FAB, these compounds produce radical cation molecular ions rather than protonated molecular ions. Similarly, under acidic mobile phase conditions in an electrospray LC-MS experiment, they produce radial cation molecular ions. Upon changing to a neutral, ammonium acetate-containing mobile phase, the molecular ion species is an ammonium adduct. The two example compounds behave conventionally under negative ion detection, both being free carboxylic acids and forming abundant [M - H](-). Examination of structural analogs indicates that the chromanol, methoxyl and chroman compounds behave this way. Oxidation to a chromanone causes formation of a conventional [M + H](+). Oxidation to a chromene produces even more complex behavior-namely a mixture of [M - H](+), M(+') and [M + H](+). We propose that, for these compounds, elimination of a valence electron to form the radical cation is the more energetically favored reaction than attachment of a proton.