In the United States (US), tianeptine is sold as a dietary supplement under the trade name Tianaa... more In the United States (US), tianeptine is sold as a dietary supplement under the trade name Tianaa™. Tianeptine is a synthetic drug prescribed by physicians as an antidepressant in parts of Europe, Asia and South America. The drug is not permitted for use by physicians in the US, be-cause it is a μ-opioid receptor agonist with a propensity for severe addiction. As incidence of Tianaa™ related opioid addiction across the southern US escalate, the current study aimed to quantify tianeptine in over the counter Tianaa™ White, Red, and Green products. The results of this investigation measured tianeptine levels between 3.1 to 10.9 mg per 531 mg capsules. Tianaa™ White capsules consistently contained the least tianeptine, while Green had the most. Close inspection of Tianaa™ products showed that capsule mass varied by as much as 16% from label claim, and the amount of tianeptine per capsule varied by as much as 35 % from the aver-age measured amount for each product. Tianaa™ Red contains kava leaf extract, which led to the identification of four kavapyrone components by mass spectrometry. The data presented pro-vide insight into tianeptine quantity and capsule mass variation for Tianaa™ supplements sold to customers naive to the risk of addiction.
According to the CDC approximately 76.8 million American adults are obese; a medical condition th... more According to the CDC approximately 76.8 million American adults are obese; a medical condition that costs the United States $147 billion dollars annually. One of the factors thought to be contributing to this growing epidemic is the over consumption of trans fats in today’s formulated foods. According to the FDA’s definition, trans- fatty acids are unsaturated fatty acids that have one or more of their double bonds changed from the cis- to trans- orientation. Hydrogenated oils are unsaturated oils that are stabilized to improve the shelf life of foods, but this process can sometimes produce trans- confirmations. The goal of this study is to investigate the trans fat content of commercial french fries before and after final frying in fresh, medium and heavily used soybean oil. To determine trans fat content in commercial fries before and after final frying, fries were crushed and placed in hexanes. After the extraction, methyl ester derivatives were synthesized by reaction with potassium hydroxide and methanol. The extracts were separated and analyzed by gas chromatography-mass spectrometry and high pressure liquid chromatography
Plants of the Veratrum genus have been used throughout history for their emetic properties, rheum... more Plants of the Veratrum genus have been used throughout history for their emetic properties, rheumatism, and for the treatment of high blood pressure. However, inadvertent consumption of these plants, which resemble wild ramps, induces life-threatening side effects attributable to an abundance of steroidal alkaloids. Several of the steroidal alkaloids from Veratrum spp. have been investigated for their ability to antagonize the Hedgehog (Hh) signaling pathway, a key pathway for embryonic development and cell proliferation. Uncontrolled activation of this pathway is linked to the development of various cancers; most notably, basal cell carcinoma and acute myeloid leukemia. Additional investigation of Veratrum spp. may lead to the identification of novel alkaloids with the potential to serve as chemotherapeutics. V. parviflorum is a relatively uncommon species of Veratrum that resides in the southeastern regions of North America. The phytochemical profile of this plant remains largely ...
Plants belonging to the genus Veratrum have been used throughout history for their medicinal prop... more Plants belonging to the genus Veratrum have been used throughout history for their medicinal properties. During the nineteenth and twentieth centuries, phytochemical investigations revealed a host of steroidal alkaloids in Veratrum species, some of which are potent bioactives. This review discusses Veratrum species that grow in North America with a focus on the medicinal history of these plants and the steroidal alkaloids they contain. While significant reviews have been devoted to singularly describing the plant species within the genus Veratrum (botany), the staggering breadth of alkaloids isolated from these and related plants (phytochemistry), and the intricacies of how the various alkaloids act on their biological targets (physiology and biochemistry), this review will straddle the margins of the aforementioned disciplines in an attempt to provide a unified, coherent picture of the Veratrum plants of North America and the medicinal uses of their bioactive steroidal alkaloids
Veratrum californicum is a rich source of steroidal alkaloids such as cyclopamine, a known inhibi... more Veratrum californicum is a rich source of steroidal alkaloids such as cyclopamine, a known inhibitor of the Hedgehog (Hh) signaling pathway. Here we provide a detailed analysis of the alkaloid composition of V. californicum by plant part through quantitative analysis of cyclopamine, veratramine, muldamine and isorubijervine in the leaf, stem and root/rhizome of the plant. To determine whether additional alkaloids in the extracts contribute to Hh signaling inhibition, the concentrations of these four alkaloids present in extracts were replicated using commercially available standards, followed by comparison of extracts to alkaloid standard mixtures for inhibition of Hh signaling using Shh-Light II cells. Alkaloid combinations enhanced Hh signaling pathway antagonism compared to cyclopamine alone, and significant differences were observed in the Hh pathway inhibition between the stem and root/rhizome extracts and their corresponding alkaloid standard mixtures, indicating that addition...
Veratrum spp. grow throughout the world and are especially prevalent in high mountain meadows of ... more Veratrum spp. grow throughout the world and are especially prevalent in high mountain meadows of North America. All parts of Veratrum plants have been used for the treatment of ailments including injuries, hypertension, and rheumatic pain since as far back as the 1600s. Of the 17–45 Veratrum spp., Veratrum californicum alkaloids have been proven to possess favorable medicinal properties associated with inhibition of hedgehog (Hh) pathway signaling. Aberrant Hh signaling leads to proliferation of over 20 cancers, including basal cell carcinoma, prostate and colon among others. Six of the most well-studied V. californicum alkaloids are cyclopamine (1), veratramine (2), isorubijervine (3), muldamine (4), cycloposine (5), and veratrosine (6). Recent inspection of the ethanolic extract from V. californicum root and rhizome via liquid chromatography–mass spectrometry has detected up to five additional alkaloids that are proposed to be verazine (7), etioline (8), tetrahydrojervine (9), dih...
KTM is a rationally designed alpha-conotoxin predicted to have optimal binding affinity for the r... more KTM is a rationally designed alpha-conotoxin predicted to have optimal binding affinity for the rat α3β2 nicotinic acetylcholine receptor isoform, a homology model for the human α6α4β2β3 receptor isoform implicated in Parkinson’s Disease. Validation of computational accuracy will help adjust computational parameters to give more accurate predictions of receptor binding, which are critical to receptor understanding and effective drug development for neurodegenerative diseases such as Parkinson’s. The NMR structure of KTM is currently being solved in order to validate computational results. Current progress indicates that the NMR structure follows the predicted structure well, but is not highly constrained
According to the CDC approximately 76.8 million American adults are obese; a medical condition th... more According to the CDC approximately 76.8 million American adults are obese; a medical condition that costs the United States $147 billion dollars annually. One of the factors thought to be contributing to this growing epidemic is the over consumption of trans fats in today’s formulated foods. According to the FDA’s definition, trans- fatty acids are unsaturated fatty acids that have one or more of their double bonds changed from the cis- to trans- orientation. Hydrogenated oils are unsaturated oils that are stabilized to improve the shelf life of foods, but this process can sometimes produce trans- confirmations. The goal of this study is to investigate the trans fat content of commercial french fries before and after final frying in fresh, medium and heavily used soybean oil. To determine trans fat content in commercial fries before and after final frying, fries were crushed and placed in hexanes. After the extraction, methyl ester derivatives were synthesized by reaction with potassium hydroxide and methanol. The extracts were separated and analyzed by gas chromatography-mass spectrometry and high pressure liquid chromatography
Veratrum spp. grow throughout the world and are especially prevalent in high mountain meadows of ... more Veratrum spp. grow throughout the world and are especially prevalent in high mountain meadows of North America. All parts of Veratrum plants have been used for the treatment of ailments including injuries, hypertension, and rheumatic pain since as far back as the 1600s. Of the 17–45 Veratrum spp., Veratrum californicum alkaloids have been proven to possess favorable medicinal properties associated with inhibition of hedgehog (Hh) pathway signaling. Aberrant Hh signaling leads to proliferation of over 20 cancers, including basal cell carcinoma, prostate and colon among others. Six of the most well-studied V. californicum alkaloids are cyclopamine (1), veratramine (2), isorubijervine (3), muldamine (4), cycloposine (5), and veratrosine (6). Recent inspection of the ethanolic extract from V. californicum root and rhizome via liquid chromatography–mass spectrometry has detected up to five additional alkaloids that are proposed to be verazine (7), etioline (8), tetrahydrojervine (9), dih...
A pheochromocytoma of the rat adrenal medulla derived (a.k.a. PC12) cell-based assay for dopamine... more A pheochromocytoma of the rat adrenal medulla derived (a.k.a. PC12) cell-based assay for dopamine measurement by luminescence detection was customized for the qualitative evaluation of agonists and antagonists of nicotinic acetylcholine receptors (nAChRs). The assay mechanism begins with ligand binding to transmembrane nAChRs, altering ion flow into the cell and inducing dopamine release from the cell. Following release, dopamine is oxidized by monoamine oxidase generating hydrogen peroxide that catalyzes a chemiluminescence reaction involving luminol and horseradish peroxidase, thus producing a detectable response. Results are presented for the action of nAChR agonists (acetylcholine, nicotine, and cytisine), and antagonists (α-conotoxins (α-CTxs) MII, ImI, LvIA, and PeIA) that demonstrate a luminescence response correlating to the increase or decrease of dopamine release. A survey of cell growth and treatment conditions, including nerve growth factor, nicotine, ethanol, and temper...
KTM is a 16 amino acid peptide with the sequence WCCSYPGCYWSSSKWC. Here, we present the nuclear m... more KTM is a 16 amino acid peptide with the sequence WCCSYPGCYWSSSKWC. Here, we present the nuclear magnetic resonance (NMR) structure and bioactivity of this rationally designed α-conotoxin (α-CTx) that demonstrates potent inhibition of rat α3β2-nicotinic acetylcholine receptors (rα3β2-nAChRs). Two bioassays were used to test the efficacy of KTM. First, a qualitative PC12 cell-based assay confirmed that KTM acts as a nAChR antagonist. Second, bioactivity evaluation by two-electrode voltage clamp electrophysiology was used to measure the inhibition of rα3β2-nAChRs by KTM (IC50 = 0.19 ± 0.02 nM), and inhibition of the same nAChR isoform by α-CTx MII (IC50 = 0.35 ± 0.8 nM). The three-dimensional structure of KTM was determined by NMR spectroscopy, and the final set of 20 structures derived from 32 distance restraints, four dihedral angle constraints, and two disulfide bond constraints overlapped with a mean global backbone root-mean-square deviation (RMSD) of 1.7 ± 0.5 Å. The structure of...
Four steroidal alkaloids extracted from the roots and rhizomes of Veratrum californicum were sepa... more Four steroidal alkaloids extracted from the roots and rhizomes of Veratrum californicum were separated by high performance liquid chromatography (HPLC) and identified using high resolution electrospray ionization time of flight tandem mass spectrometry (ESI-TOF-MS/MS) as veratrosine, cycloposine, veratramine, and cyclopamine. This paper compares ethanol and benzene as extraction solvents, HPLC solvent conditions leading to good separation of steroidal alkaloids, and MS/MS fragmentation patterns for the four steroidal alkaloids which have been released to the public database MassBank.jp. The reported Soxhlet extraction method nearly triples the recovery of steroidal alkaloids from V. californicum.
Nicotinic acetylcholine receptors (nAChRs) are found throughout the mammalian body and have been ... more Nicotinic acetylcholine receptors (nAChRs) are found throughout the mammalian body and have been studied extensively because of their implication in a myriad of diseases. α-Conotoxins (α-CTxs) are peptide neurotoxins found in the venom of marine snails of genus Conus. α-CTxs are potent and selective antagonists for a variety of nAChR isoforms. Over the past 40 years, α-CTxs have proven to be valuable molecular probes capable of differentiating between closely related nAChR subtypes and have contributed greatly to understanding the physiological role of nAChRs in the mammalian nervous system. Here, we review the amino acid composition and structure of several α-CTxs that selectively target nAChR isoforms and explore strategies and outcomes for introducing mutations in native α-CTxs to direct selectivity and enhance binding affinity for specific nAChRs. This review will focus on structure-activity relationship studies involving native α-CTxs that have been rationally mutated and molec...
is a rich source of steroidal alkaloids such as cyclopamine, a known inhibitor of the Hedgehog (H... more is a rich source of steroidal alkaloids such as cyclopamine, a known inhibitor of the Hedgehog (Hh) signaling pathway. Here we provide a detailed analysis of the alkaloid composition of by plant part through quantitative analysis of cyclopamine, veratramine, muldamine and isorubijervine in the leaf, stem and root/rhizome of the plant. To determine whether additional alkaloids in the extracts contribute to Hh signaling inhibition, the concentrations of these four alkaloids present in extracts were replicated using commercially available standards, followed by comparison of extracts to alkaloid standard mixtures for inhibition of Hh signaling using Shh-Light II cells. Alkaloid combinations enhanced Hh signaling pathway antagonism compared to cyclopamine alone, and significant differences were observed in the Hh pathway inhibition between the stem and root/rhizome extracts and their corresponding alkaloid standard mixtures, indicating that additional alkaloids present in these extracts...
Conventional de novo drug design is costly and time consuming, making it accessible to only the b... more Conventional de novo drug design is costly and time consuming, making it accessible to only the best resourced research organizations. An emergent approach to new drug development is drug repurposing, in which compounds that have already gone through some level of clinical testing are examined for efficacy against diseases divergent than their original application. Repurposing of existing drugs circumvents the time and considerable cost of early stages of drug development, and can be accelerated by using software to screen existing chemical databases to identify suitable drug candidates. Small-molecule Peptide-Influenced Drug Repurposing (SPIDR) was developed to identify small molecule drugs that target a specific receptor by exploring the conformational binding space of peptide ligands. SPIDR was tested using the potent and selective 16-amino acid peptide α-conotoxin MII ligand and the αβ-nicotinic acetylcholine receptor (nAChR) isoform. SPIDR incorporates a genetic algorithm-based...
Clinical toxicology (Philadelphia, Pa.), Jan 28, 2018
Steroidal alkaloids are found in plants of the genus Veratrum. Their toxicity manifests as gastro... more Steroidal alkaloids are found in plants of the genus Veratrum. Their toxicity manifests as gastrointestinal symptoms followed by a Bezold-Jarisch reflex: hypopnea, hypotension, and bradycardia. Some Veratrum steroidal alkaloids are also teratogens interfering with the hedgehog-2 signaling pathway, which causes cyclopsia and holoprosencephaly. We present a case of accidental poisoning from Veratrum parviflorum mistaken for the edible Allium tricoccum (ramps, wild leek). A 27-year-old man and his 25-year-old wife presented to the emergency department with nausea, vomiting, hypotension, and bradycardia after foraging and ingesting plants that they believed to be a local native species of wild leek. We collected and analyzed the implicated fresh plant material and both patients' serum/plasma. We used liquid chromatography-mass spectroscopy and high-resolution electrospray ionization time of flight tandem mass spectrometry to extract and characterize steroidal alkaloids from the fora...
In the United States (US), tianeptine is sold as a dietary supplement under the trade name Tianaa... more In the United States (US), tianeptine is sold as a dietary supplement under the trade name Tianaa™. Tianeptine is a synthetic drug prescribed by physicians as an antidepressant in parts of Europe, Asia and South America. The drug is not permitted for use by physicians in the US, be-cause it is a μ-opioid receptor agonist with a propensity for severe addiction. As incidence of Tianaa™ related opioid addiction across the southern US escalate, the current study aimed to quantify tianeptine in over the counter Tianaa™ White, Red, and Green products. The results of this investigation measured tianeptine levels between 3.1 to 10.9 mg per 531 mg capsules. Tianaa™ White capsules consistently contained the least tianeptine, while Green had the most. Close inspection of Tianaa™ products showed that capsule mass varied by as much as 16% from label claim, and the amount of tianeptine per capsule varied by as much as 35 % from the aver-age measured amount for each product. Tianaa™ Red contains kava leaf extract, which led to the identification of four kavapyrone components by mass spectrometry. The data presented pro-vide insight into tianeptine quantity and capsule mass variation for Tianaa™ supplements sold to customers naive to the risk of addiction.
According to the CDC approximately 76.8 million American adults are obese; a medical condition th... more According to the CDC approximately 76.8 million American adults are obese; a medical condition that costs the United States $147 billion dollars annually. One of the factors thought to be contributing to this growing epidemic is the over consumption of trans fats in today’s formulated foods. According to the FDA’s definition, trans- fatty acids are unsaturated fatty acids that have one or more of their double bonds changed from the cis- to trans- orientation. Hydrogenated oils are unsaturated oils that are stabilized to improve the shelf life of foods, but this process can sometimes produce trans- confirmations. The goal of this study is to investigate the trans fat content of commercial french fries before and after final frying in fresh, medium and heavily used soybean oil. To determine trans fat content in commercial fries before and after final frying, fries were crushed and placed in hexanes. After the extraction, methyl ester derivatives were synthesized by reaction with potassium hydroxide and methanol. The extracts were separated and analyzed by gas chromatography-mass spectrometry and high pressure liquid chromatography
Plants of the Veratrum genus have been used throughout history for their emetic properties, rheum... more Plants of the Veratrum genus have been used throughout history for their emetic properties, rheumatism, and for the treatment of high blood pressure. However, inadvertent consumption of these plants, which resemble wild ramps, induces life-threatening side effects attributable to an abundance of steroidal alkaloids. Several of the steroidal alkaloids from Veratrum spp. have been investigated for their ability to antagonize the Hedgehog (Hh) signaling pathway, a key pathway for embryonic development and cell proliferation. Uncontrolled activation of this pathway is linked to the development of various cancers; most notably, basal cell carcinoma and acute myeloid leukemia. Additional investigation of Veratrum spp. may lead to the identification of novel alkaloids with the potential to serve as chemotherapeutics. V. parviflorum is a relatively uncommon species of Veratrum that resides in the southeastern regions of North America. The phytochemical profile of this plant remains largely ...
Plants belonging to the genus Veratrum have been used throughout history for their medicinal prop... more Plants belonging to the genus Veratrum have been used throughout history for their medicinal properties. During the nineteenth and twentieth centuries, phytochemical investigations revealed a host of steroidal alkaloids in Veratrum species, some of which are potent bioactives. This review discusses Veratrum species that grow in North America with a focus on the medicinal history of these plants and the steroidal alkaloids they contain. While significant reviews have been devoted to singularly describing the plant species within the genus Veratrum (botany), the staggering breadth of alkaloids isolated from these and related plants (phytochemistry), and the intricacies of how the various alkaloids act on their biological targets (physiology and biochemistry), this review will straddle the margins of the aforementioned disciplines in an attempt to provide a unified, coherent picture of the Veratrum plants of North America and the medicinal uses of their bioactive steroidal alkaloids
Veratrum californicum is a rich source of steroidal alkaloids such as cyclopamine, a known inhibi... more Veratrum californicum is a rich source of steroidal alkaloids such as cyclopamine, a known inhibitor of the Hedgehog (Hh) signaling pathway. Here we provide a detailed analysis of the alkaloid composition of V. californicum by plant part through quantitative analysis of cyclopamine, veratramine, muldamine and isorubijervine in the leaf, stem and root/rhizome of the plant. To determine whether additional alkaloids in the extracts contribute to Hh signaling inhibition, the concentrations of these four alkaloids present in extracts were replicated using commercially available standards, followed by comparison of extracts to alkaloid standard mixtures for inhibition of Hh signaling using Shh-Light II cells. Alkaloid combinations enhanced Hh signaling pathway antagonism compared to cyclopamine alone, and significant differences were observed in the Hh pathway inhibition between the stem and root/rhizome extracts and their corresponding alkaloid standard mixtures, indicating that addition...
Veratrum spp. grow throughout the world and are especially prevalent in high mountain meadows of ... more Veratrum spp. grow throughout the world and are especially prevalent in high mountain meadows of North America. All parts of Veratrum plants have been used for the treatment of ailments including injuries, hypertension, and rheumatic pain since as far back as the 1600s. Of the 17–45 Veratrum spp., Veratrum californicum alkaloids have been proven to possess favorable medicinal properties associated with inhibition of hedgehog (Hh) pathway signaling. Aberrant Hh signaling leads to proliferation of over 20 cancers, including basal cell carcinoma, prostate and colon among others. Six of the most well-studied V. californicum alkaloids are cyclopamine (1), veratramine (2), isorubijervine (3), muldamine (4), cycloposine (5), and veratrosine (6). Recent inspection of the ethanolic extract from V. californicum root and rhizome via liquid chromatography–mass spectrometry has detected up to five additional alkaloids that are proposed to be verazine (7), etioline (8), tetrahydrojervine (9), dih...
KTM is a rationally designed alpha-conotoxin predicted to have optimal binding affinity for the r... more KTM is a rationally designed alpha-conotoxin predicted to have optimal binding affinity for the rat α3β2 nicotinic acetylcholine receptor isoform, a homology model for the human α6α4β2β3 receptor isoform implicated in Parkinson’s Disease. Validation of computational accuracy will help adjust computational parameters to give more accurate predictions of receptor binding, which are critical to receptor understanding and effective drug development for neurodegenerative diseases such as Parkinson’s. The NMR structure of KTM is currently being solved in order to validate computational results. Current progress indicates that the NMR structure follows the predicted structure well, but is not highly constrained
According to the CDC approximately 76.8 million American adults are obese; a medical condition th... more According to the CDC approximately 76.8 million American adults are obese; a medical condition that costs the United States $147 billion dollars annually. One of the factors thought to be contributing to this growing epidemic is the over consumption of trans fats in today’s formulated foods. According to the FDA’s definition, trans- fatty acids are unsaturated fatty acids that have one or more of their double bonds changed from the cis- to trans- orientation. Hydrogenated oils are unsaturated oils that are stabilized to improve the shelf life of foods, but this process can sometimes produce trans- confirmations. The goal of this study is to investigate the trans fat content of commercial french fries before and after final frying in fresh, medium and heavily used soybean oil. To determine trans fat content in commercial fries before and after final frying, fries were crushed and placed in hexanes. After the extraction, methyl ester derivatives were synthesized by reaction with potassium hydroxide and methanol. The extracts were separated and analyzed by gas chromatography-mass spectrometry and high pressure liquid chromatography
Veratrum spp. grow throughout the world and are especially prevalent in high mountain meadows of ... more Veratrum spp. grow throughout the world and are especially prevalent in high mountain meadows of North America. All parts of Veratrum plants have been used for the treatment of ailments including injuries, hypertension, and rheumatic pain since as far back as the 1600s. Of the 17–45 Veratrum spp., Veratrum californicum alkaloids have been proven to possess favorable medicinal properties associated with inhibition of hedgehog (Hh) pathway signaling. Aberrant Hh signaling leads to proliferation of over 20 cancers, including basal cell carcinoma, prostate and colon among others. Six of the most well-studied V. californicum alkaloids are cyclopamine (1), veratramine (2), isorubijervine (3), muldamine (4), cycloposine (5), and veratrosine (6). Recent inspection of the ethanolic extract from V. californicum root and rhizome via liquid chromatography–mass spectrometry has detected up to five additional alkaloids that are proposed to be verazine (7), etioline (8), tetrahydrojervine (9), dih...
A pheochromocytoma of the rat adrenal medulla derived (a.k.a. PC12) cell-based assay for dopamine... more A pheochromocytoma of the rat adrenal medulla derived (a.k.a. PC12) cell-based assay for dopamine measurement by luminescence detection was customized for the qualitative evaluation of agonists and antagonists of nicotinic acetylcholine receptors (nAChRs). The assay mechanism begins with ligand binding to transmembrane nAChRs, altering ion flow into the cell and inducing dopamine release from the cell. Following release, dopamine is oxidized by monoamine oxidase generating hydrogen peroxide that catalyzes a chemiluminescence reaction involving luminol and horseradish peroxidase, thus producing a detectable response. Results are presented for the action of nAChR agonists (acetylcholine, nicotine, and cytisine), and antagonists (α-conotoxins (α-CTxs) MII, ImI, LvIA, and PeIA) that demonstrate a luminescence response correlating to the increase or decrease of dopamine release. A survey of cell growth and treatment conditions, including nerve growth factor, nicotine, ethanol, and temper...
KTM is a 16 amino acid peptide with the sequence WCCSYPGCYWSSSKWC. Here, we present the nuclear m... more KTM is a 16 amino acid peptide with the sequence WCCSYPGCYWSSSKWC. Here, we present the nuclear magnetic resonance (NMR) structure and bioactivity of this rationally designed α-conotoxin (α-CTx) that demonstrates potent inhibition of rat α3β2-nicotinic acetylcholine receptors (rα3β2-nAChRs). Two bioassays were used to test the efficacy of KTM. First, a qualitative PC12 cell-based assay confirmed that KTM acts as a nAChR antagonist. Second, bioactivity evaluation by two-electrode voltage clamp electrophysiology was used to measure the inhibition of rα3β2-nAChRs by KTM (IC50 = 0.19 ± 0.02 nM), and inhibition of the same nAChR isoform by α-CTx MII (IC50 = 0.35 ± 0.8 nM). The three-dimensional structure of KTM was determined by NMR spectroscopy, and the final set of 20 structures derived from 32 distance restraints, four dihedral angle constraints, and two disulfide bond constraints overlapped with a mean global backbone root-mean-square deviation (RMSD) of 1.7 ± 0.5 Å. The structure of...
Four steroidal alkaloids extracted from the roots and rhizomes of Veratrum californicum were sepa... more Four steroidal alkaloids extracted from the roots and rhizomes of Veratrum californicum were separated by high performance liquid chromatography (HPLC) and identified using high resolution electrospray ionization time of flight tandem mass spectrometry (ESI-TOF-MS/MS) as veratrosine, cycloposine, veratramine, and cyclopamine. This paper compares ethanol and benzene as extraction solvents, HPLC solvent conditions leading to good separation of steroidal alkaloids, and MS/MS fragmentation patterns for the four steroidal alkaloids which have been released to the public database MassBank.jp. The reported Soxhlet extraction method nearly triples the recovery of steroidal alkaloids from V. californicum.
Nicotinic acetylcholine receptors (nAChRs) are found throughout the mammalian body and have been ... more Nicotinic acetylcholine receptors (nAChRs) are found throughout the mammalian body and have been studied extensively because of their implication in a myriad of diseases. α-Conotoxins (α-CTxs) are peptide neurotoxins found in the venom of marine snails of genus Conus. α-CTxs are potent and selective antagonists for a variety of nAChR isoforms. Over the past 40 years, α-CTxs have proven to be valuable molecular probes capable of differentiating between closely related nAChR subtypes and have contributed greatly to understanding the physiological role of nAChRs in the mammalian nervous system. Here, we review the amino acid composition and structure of several α-CTxs that selectively target nAChR isoforms and explore strategies and outcomes for introducing mutations in native α-CTxs to direct selectivity and enhance binding affinity for specific nAChRs. This review will focus on structure-activity relationship studies involving native α-CTxs that have been rationally mutated and molec...
is a rich source of steroidal alkaloids such as cyclopamine, a known inhibitor of the Hedgehog (H... more is a rich source of steroidal alkaloids such as cyclopamine, a known inhibitor of the Hedgehog (Hh) signaling pathway. Here we provide a detailed analysis of the alkaloid composition of by plant part through quantitative analysis of cyclopamine, veratramine, muldamine and isorubijervine in the leaf, stem and root/rhizome of the plant. To determine whether additional alkaloids in the extracts contribute to Hh signaling inhibition, the concentrations of these four alkaloids present in extracts were replicated using commercially available standards, followed by comparison of extracts to alkaloid standard mixtures for inhibition of Hh signaling using Shh-Light II cells. Alkaloid combinations enhanced Hh signaling pathway antagonism compared to cyclopamine alone, and significant differences were observed in the Hh pathway inhibition between the stem and root/rhizome extracts and their corresponding alkaloid standard mixtures, indicating that additional alkaloids present in these extracts...
Conventional de novo drug design is costly and time consuming, making it accessible to only the b... more Conventional de novo drug design is costly and time consuming, making it accessible to only the best resourced research organizations. An emergent approach to new drug development is drug repurposing, in which compounds that have already gone through some level of clinical testing are examined for efficacy against diseases divergent than their original application. Repurposing of existing drugs circumvents the time and considerable cost of early stages of drug development, and can be accelerated by using software to screen existing chemical databases to identify suitable drug candidates. Small-molecule Peptide-Influenced Drug Repurposing (SPIDR) was developed to identify small molecule drugs that target a specific receptor by exploring the conformational binding space of peptide ligands. SPIDR was tested using the potent and selective 16-amino acid peptide α-conotoxin MII ligand and the αβ-nicotinic acetylcholine receptor (nAChR) isoform. SPIDR incorporates a genetic algorithm-based...
Clinical toxicology (Philadelphia, Pa.), Jan 28, 2018
Steroidal alkaloids are found in plants of the genus Veratrum. Their toxicity manifests as gastro... more Steroidal alkaloids are found in plants of the genus Veratrum. Their toxicity manifests as gastrointestinal symptoms followed by a Bezold-Jarisch reflex: hypopnea, hypotension, and bradycardia. Some Veratrum steroidal alkaloids are also teratogens interfering with the hedgehog-2 signaling pathway, which causes cyclopsia and holoprosencephaly. We present a case of accidental poisoning from Veratrum parviflorum mistaken for the edible Allium tricoccum (ramps, wild leek). A 27-year-old man and his 25-year-old wife presented to the emergency department with nausea, vomiting, hypotension, and bradycardia after foraging and ingesting plants that they believed to be a local native species of wild leek. We collected and analyzed the implicated fresh plant material and both patients' serum/plasma. We used liquid chromatography-mass spectroscopy and high-resolution electrospray ionization time of flight tandem mass spectrometry to extract and characterize steroidal alkaloids from the fora...
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Papers by Owen McDougal