The toxicities of A30P and A53T α-synuclein fibrils can be uniquely altered by the length and saturation of fatty acids in phosphatidylserine.

Progressive degeneration of dopaminergic neurons in the midbrain, hypothalamus, and thalamus is a hallmark of Parkinson's disease (PD). Neuronal death is linked to the abrupt aggregation of α-synuclein (α-syn), a small protein that regulates vesicle trafficking in synaptic clefts. Studies of families with a history of PD revealed several mutations in α-syn including A30P and A53T that are linked to the early onset of this pathology. Numerous pieces of evidence indicate that lipids can alter the rate of protein aggregation, as well as modify the secondary structure and toxicity of amyloid oligomers and fibrils. However, the role of lipids in the stability of α-syn mutants remains unclear. In this study, we investigate the effect of phosphatidylserine (PS), an anionic lipid that plays an important role in the recognition of apoptotic cells by macrophages, in the stability of WT, A30P, and A53T α-syn. We found PS with different lengths and saturation of fatty acids accelerated the rate of WT and A30P aggregation. At the same time, the opposite effect was observed for most PS on A53T. We also found that PS with different lengths and saturation of fatty acids change the secondary structure and toxicities of WT, A30P, and A53T fibrils. These results indicate that lipids can play an important role in the onset and spread of familial PD.

Secondary structure and toxicity of lysozyme fibrils are determined by the length and unsaturation of phosphatidic acid.

A progressive aggregation of misfolded proteins is a hallmark of numerous pathologies including diabetes Type 2, Alzheimer's disease, and Parkinson's disease. As a result, highly toxic protein aggregates, which are known as amyloid fibrils, are formed. A growing body of evidence suggests that phospholipids can uniquely alter the secondary structure and toxicity of amyloid aggregates. However, the role of phosphatidic acid (PA), a unique lipid that is responsible for cell signaling and activation of lipid-gated ion channels, in the aggregation of amyloidogenic proteins remains unclear. In this study, we investigate the role of the length and degree of unsaturation of fatty acids (FAs) in PA in the structure and toxicity of lysozyme fibrils formed in the presence of this lipid. We found that both the length and saturation of FAs in PA uniquely altered the secondary structure of lysozyme fibrils. However, these structural differences in PA caused very little if any changes in the morphology of lysozyme fibrils. We also utilized cell toxicity assays to determine the extent to which the length and degree of unsaturation of FAs in PA altered the toxicity of lysozyme fibrils. We found that amyloid fibrils formed in the presence of PA with C18:0 FAs exerted significantly higher cell toxicity compared to the aggregates formed in the presence of PA with C16:0 and C18:1 FAs. These results demonstrated that PA can be an important player in the onset and spread of amyloidogenic diseases.

Elucidating the mechanisms of α-Synuclein-lipid interactions using site-directed mutagenesis.

α-Synuclein (α-syn) is a small protein that is involved in cell vesicle trafficking in neuronal synapses. A progressive aggregation of this protein is the expected molecular cause of Parkinson's disease, a disease that affects millions of people around the world. A growing body of evidence indicates that phospholipids can strongly accelerate α-syn aggregation and alter the toxicity of α-syn oligomers and fibrils formed in the presence of lipid vesicles. This effect is attributed to the presence of high copies of lysines in the N-terminus of the protein. In this study, we performed site-directed mutagenesis and replaced one out of two lysines at each of the five sites located in the α-syn N-terminus. Using several biophysical and cellular approaches, we investigated the extent to which six negatively charged fatty acids (FAs) could alter the aggregation properties of K10A, K23A, K32A, K43A, and K58A α-syn. We found that FAs uniquely modified the aggregation properties of K43A, K58A, and WT α-syn, as well as changed morphology of amyloid fibrils formed by these mutants. At the same time, FAs failed to cause substantial changes in the aggregation rates of K10A, K23A, and K32A α-syn, as well as alter the morphology and toxicity of the corresponding amyloid fibrils. Based on these results, we can conclude that K10, K23, and K32 amino acid residues play a critical role in protein-lipid interactions since their replacement on non-polar alanines strongly suppressed α-syn-lipid interactions.

Elucidating the Role of Lipids in the Aggregation of Amyloidogenic Proteins.

The abrupt aggregation of misfolded proteins is linked to the onset and spread of amyloidogenic diseases, including diabetes type 2, systemic amyloidosis, and Alzheimer's (AD) and Parkinson's diseases (PD). Although the exact cause of these pathological processes is unknown, a growing body of evidence suggests that amyloid diseases are triggered by misfolded or unfolded proteins, forming highly toxic oligomers. These transient species exhibit high structural and morphological heterogeneity. Protein oligomers can also propagate into ß-sheet-rich filaments that braid and coil with other filaments to form amyloid fibrils and supramolecular structures with both flat and twisted morphologies. Microscopic examination of protein deposits formed in the brains of both AD and PD patients revealed the presence of fragments of lipid membranes. Furthermore, nanoscale infrared analysis of ex vivo extracted fibrils revealed the presence of lipids in their structure (Zhaliazka, K.; Kurouski, D. Protein Sci. 2023, 32, e4598). These findings demonstrated that lipid bilayers could play an important role in the aggregation of misfolded proteins.Experimental findings summarized in this Account show that (i) lipids uniquely change the aggregation rate of amyloidogenic proteins. In this case, the observed changes in the rates directly depend on the net charge of the lipid and the length and saturation of lipid fatty acids (FAs). For instance, zwitterionic phosphatidylcholine (PC) with 14:0 FAs inhibited the aggregation of insulin, lysozyme, and α-synuclein (α-Syn), whereas anionic phosphatidylserine with the same FAs dramatically accelerated the aggregation rate of these proteins (Dou, T., et al. J. Phys. Chem. Lett. 2021, 12, 4407. Matveyenka, M., et al. FASEB J. 2022, 36, e22543. Rizevsky, S., et al. J. Phys. Chem. Lett. 2022, 13, 2467). Furthermore, (ii) lipids uniquely alter the secondary structure and morphology of protein oligomers and fibrils formed in their presence. Utilization of nano-infrared spectroscopy revealed that such aggregates, as well as ex vivo extracted fibrils, possessed lipids in their structure. These findings are significant because (iii) lipids uniquely alter the toxicity of amyloid oligomers and fibrils formed in their presence. Specifically, PC lowered the toxicity of insulin and lysozyme oligomers, whereas α-Syn oligomers formed in the presence of this phospholipid were found to be significantly more toxic to rat dopaminergic cells compared to α-Syn oligomers grown in the lipid-free environment. Thus, the toxicity of protein oligomers and fibrils is directly determined by the chemical structure of the lipid and the secondary structure of amyloidogenic proteins (Dou, T., et al. J. Phys. Chem. Lett. 2021, 12, 4407. Matveyenka, M., et al. FASEB J. 2022, 36, e22543. Rizevsky, S., et al. J. Phys. Chem. Lett. 2022, 13, 2467). Experimental results discussed in this Account also suggest that amyloidogenic diseases could be caused by pathological changes in the lipid composition of both plasma and organelle membranes, which, in turn, may trigger protein aggregation that results in the formation of highly toxic oligomers and fibrils. Finally, the Account discusses the effects of polyunsaturated FAs on the aggregation properties of amyloidogenic proteins. Experimental findings reported by the author's laboratory revealed that polyunsaturated FAs drastically accelerated the aggregation rate of both insulin and α-Syn as well as strongly changed the secondary structure of amyloid fibrils formed in their presence.

Unsaturated fatty acids uniquely alter aggregation rate of α-synuclein and insulin and change the secondary structure and toxicity of amyloid aggregates formed in their presence.

Docosahexaenoic (DHA) and arachidonic acids (ARA) are omega-3 and omega-6 long-chain polyunsaturated fatty acids (LCPUFAs). These molecules constitute a substantial portion of phospholipids in plasma membranes. Therefore, both DHA and ARA are essential diet components. Once consumed, DHA and ARA can interact with a large variety of biomolecules, including proteins such as insulin and α-synuclein (α-Syn). Under pathological conditions known as injection amyloidosis and Parkinson's disease, these proteins aggregate forming amyloid oligomers and fibrils, toxic species that exert high cell toxicity. In this study, we investigate the role of DHA and ARA in the aggregation properties of α-Syn and insulin. We found that the presence of both DHA and ARA at the equimolar concentrations strongly accelerated aggregation rates of α-Syn and insulin. Furthermore, LCPUFAs substantially altered the secondary structure of protein aggregates, whereas no noticeable changes in the fibril morphology were observed. Nanoscale Infrared analysis of α-Syn and insulin fibrils grown in the presence of both DHA and ARA revealed the presence of LCPUFAs in these aggregates. We also found that such LCPUFAs-rich α-Syn and insulin fibrils exerted significantly greater toxicities compared to the aggregates grown in the LCPUFAs-free environment. These findings show that interactions between amyloid-associated proteins and LCPUFAs can be the underlying molecular cause of neurodegenerative diseases.

Macrophages and Natural Killers Degrade α-Synuclein Aggregates.

Amyloid oligomers and fibrils are protein aggregates that exert a high cell toxicity. Efficient degradation of these protein aggregates can minimize the spread and progression of neurodegeneration. In this study, we investigate the properties of natural killer (NK) cells and macrophages in the degradation of α-synuclein (α-Syn) aggregates grown in a lipid-free environment and in the presence of phosphatidylserine and cholesterol (PS/Cho), which are lipids that are directly associated with the onset and progression of Parkinson's disease. We found that both types of α-Syn aggregates were endocytosed by neurons, which caused strong damage to cell endosomes. Our results also indicated that PS/Cho vesicles drastically increased the toxicity of α-Syn fibrils formed in their presence compared to the toxicity of α-Syn aggregates grown in a lipid-free environment. Both NK cells and macrophages were able to degrade α-Syn and α-Syn/Cho monomers, oligomers, and fibrils. Quantitative analysis of protein degradation showed that macrophages demonstrated substantially more efficient internalization and degradation of amyloid aggregates in comparison to NK cells. We also found that amyloid aggregates induced the proliferation of macrophages and NK cells and significantly changed the expression of their cytokines and chemokines.

Large Unilamellar Vesicles of Phosphatidic Acid Reduce the Toxicity of α-Synuclein Fibrils.

Parkinson's disease (PD) is a severe pathology that is caused by a progressive degeneration of dopaminergic neurons in substantia nigra pars compacta as well as other areas in the brain. These neurodegeneration processes are linked to the abrupt aggregation of α-synuclein (α-syn), a small protein that is abundant at presynaptic nerve termini, where it regulates cell vesicle trafficking. Due to the direct interactions of α-syn with cell membranes, a substantial amount of work was done over the past decade to understand the role of lipids in α-syn aggregation. However, the role of phosphatidic acid (PA), a negatively charged phospholipid with a small polar head, remains unclear. In this study, we examined the effect of PA large unilamellar vesicles (LUVs) on α-syn aggregation. We found that PA LUVs with 16:0, 18:0, and 18:1 FAs drastically reduced the toxicity of α-syn fibrils if were present in a 1:1 molar ratio with the protein. Our results also showed that the presence of these vehicles changed the rate of α-syn aggregation and altered the morphology and secondary structure of α-syn fibrils. These results indicate that PA LUVs can be used as a potential therapeutic strategy to reduce the toxicity of α-syn fibrils formed upon PD.

Raman-based diagnostics of drought, heat and light-induced stresses in three different varieties of hemp.

MAIN CONCLUSION: Hand-held Raman spectroscopy can be used for highly accurate differentiation between drought, heat and light-triggered stresses in hemp. The differentiation is based on the changes in the biochemistry of plants caused by such stresses. Hemp farming is a rapidly growing industry. This dioecious plant is primarily cultivated for its fibers, seeds, and cannabinoid-rich oils. The yield of these materials can be drastically lowered by many abiotic stresses, such as drought, heat and light. It becomes critically important to develop robust and reliable approaches that can be used to diagnose such abiotic stresses in hemp. In this study, we investigate the accuracy of Raman spectroscopy, an emerging tool within crop monitoring, in the confirmatory identification of drought, heat, and light-induced stresses in three varieties of hemp. Our results showed that mono, double and triple stresses uniquely alter plant biochemistry that results in small spectroscopic changes detected in the Raman spectra acquired from the hemp leaves. These changes could be used for the 80-100% accurate identification of individual abiotic stresses and their combinations in plants. These results demonstrate that a hand-held Raman spectrometer can be used for highly accurate, non-invasive, non-destructive, and label-free diagnostics of hemp stresses directly in the greenhouse or in the field.

Lipids uniquely alter secondary structure and toxicity of lysozyme aggregates.

Abrupt aggregation of misfolded proteins is a hallmark of the large group of amyloid pathologies that include diabetes type 2, Alzheimer and Parkinson's diseases. Protein aggregation yields oligomers and fibrils, ß-sheet-rich structures that exert cell toxicity. Microscopic examination of amyloid deposits reveals the presence of lipids membranes, which suggests that lipids can be involved in the process of pathogenic protein assembly. In this study, we show that lipids can uniquely alter the aggregation rates of lysozyme, a protein that is associated with systemic amyloidosis. Specifically, cardiolipin (CL), ceramide (CER), and sphingomyelin (SM) accelerate, phosphatidylcholine (PC) strongly inhibits, whereas phosphatidylserine (PS) has no effect on the rate of protein aggregation. Furthermore, lipids uniquely alter the secondary structure of lysozyme aggregates. Furthermore, we found that lysozyme aggregates grown in the presence of CL, CER, SM, PS, and CL:PC mixtures exert significantly lower production of reactive oxygen species and mitochondrial dysfunction compared to lysozyme:PC aggregates and lysozyme fibrils grown in the lipid-free environment. These findings suggest that a change in the lipid composition of cell membranes, which is taken place upon neurodegeneration, may trigger the formation of toxic protein species that otherwise would not be formed.

Can Light Alter the Yield of Plasmon-Driven Reactions on Gold and Gold-Palladium Nanoplates?

Noble-metal nanostructures, as well as their bimetallic analogues, catalyze a broad spectrum of plasmon-driven reactions. Catalytic properties of such nanostructures arise from light-generated surface plasmon resonances that decay forming transient hot electrons and holes. Hot carriers with "slower" dissipation rates accumulate on nanostructures generating an electrostatic potential. In this study, we examine whether light intensity can alter the electrostatic potential of mono- and bimetallic nanostructures changing yields of plasmon-driven reactions. Using tip-enhanced Raman spectroscopy (TERS), we quantified the yield of plasmon-driven transformations of 4-nitrobenzenethiol (4-NBT) and 3-mercaptobenzoic acid (3-MBA) on gold and gold-palladium nanoplates (AuNPs and Au@PdNPs, respectively). We found that on AuNPs 3-MBA decarboxylated forming thiophenol (TP), whereas 4-NBT was reduced to DMAB. The yield of both TP and DMAB gradually increased with increasing light intensity. On Au@PdNPs, 3-MBA could be reduced to 3-mercaptophenylmethanol (3-MPM), the yield of which was also directly dependent on the light intensity.

Non-Destructive Identification of Dyes on Fabric Using Near-Infrared Raman Spectroscopy.

Fabric is a commonly found piece of physical evidence at most crime scenes. Forensic analysis of fabric is typically performed via microscopic examination. This subjective approach is primarily based on pattern recognition and, therefore, is often inconclusive. Most of the fabric material found at crime scenes is colored. One may expect that a confirmatory identification of dyes can be used to enhance the reliability of the forensic analysis of fabric. In this study, we investigated the potential of near-infrared Raman spectroscopy (NIRS) in the confirmatory, non-invasive, and non-destructive identification of 15 different dyes on cotton. We found that NIRS was able to resolve the vibrational fingerprints of all 15 colorants. Using partial-squared discriminant analysis (PLS-DA), we showed that NIRS enabled ~100% accurate identification of dyes based on their vibrational signatures. These findings open a new avenue for the robust and reliable forensic analysis of dyes on fabric directly at crime scenes. Main conclusion: a hand-held Raman spectrometer and partial least square discriminant analysis (PLS-DA) approaches enable highly accurate identification of dyes on fabric.

Solid-to-Liposome Conformational Transition of Phosphatidylcholine and Phosphatidylserine Probed by Atomic Force Microscopy, Infrared Spectroscopy, and Density Functional Theory Calculations.

Liposomes are emerging therapeutic formulations for site-specific delivery of chemotherapeutic drugs. The efficiency and selectivity of drug delivery by these carriers largely rely on their surface properties, shape, and size. There is a growing demand for analytical approaches that can be used for structural and morphological characterization of liposomes at the single-vesicle level. AFM-IR is a modern optical nanoscopic technique that combines the advantages of scanning probe microscopy and infrared spectroscopy. Our findings show that AFM-IR can be used to probe conformational changes in phospholipids that take place upon their assembly into liposomes. Such conclusions can be made based on the corresponding changes in intensities of the lipid vibrational bands as the molecules transition from a solid state into large unilamellar vesicles (LUVs). This spectroscopic analysis of LUV formation together with density functional theory calculations also reveals the extent to which the molecular conformation and local environment of the functional groups alter the AFM-IR spectra of phospholipids. Using melittin as a test protein, we also examined the extent to which LUVs can be used for protein internalization. We found that melittin enters LUVs nearly instantaneously, which protects it from possible structural modifications that are caused by a changing environment. This foundational work empowers AFM-IR analysis of liposomes and opens new avenues for determination of the molecular mechanisms of liposome-drug interactions.

Raman spectroscopy enables highly accurate differentiation between young male and female hemp plants.

MAIN CONCLUSION: Hand-held Raman spectroscopy can be used for highly accurate differentiation between young male and female hemp plants. This differentiation is based on significantly different concentration of lutein in these plants. Last year, a global market of only industrial hemp attained the value of USD 4.7 billion. It is by far the fastest growing market with projected growth of 22.5% between 2021 and 2026. Hemp (Cannabis sativa L.) is a dioecious species that has separate male and female plants. In hemp farming, female plants are strongly preferred because male plants do not produce sufficient amount of cannabinoids. Male plants are also eliminated to minimize a possibility of uncontrolled cross-fertilization of plants. Silver treatments can induce development of male flowers on genetically female plants in order to produce feminized seed. Resulting cannabinoid hemp production fields should contain 100% female plants. However, any unintended pollination from male plants can produce unwanted males in production fields. Therefore, there is a growing demand for a label-free, non-invasive, and confirmatory approach that can be used to differentiate between male and female plants before flowering. In this study, we examined the extent to which Raman spectroscopy, an emerging optical technique, can be used for the accurate differentiation between young male and female hemp plants. Our findings show that Raman spectroscopy enables differentiation between male and female plants with 90% and 94% accuracy on the level of young and mature plants, respectively. Such analysis is entirely non-invasive and non-destructive to plants and can be performed in seconds using a hand-held spectrometer. High-performance liquid chromatography (HPLC) analysis and collected Raman spectra demonstrate that this spectroscopic differentiation is based on significantly different concentrations of carotenoids in male vs female plants. These findings open up a new avenue for quality control of plants grown in both field and a greenhouse.

Plasmon-Driven Chemistry on Mono- and Bimetallic Nanostructures.

ConspectusHot carriers are highly energetic species that can perform a large spectrum of chemical reactions. They are generated on the surfaces of nanostructures via direct interband, phonon-assisted intraband, and geometry-assisted decay of localized surface plasmon resonances (LSPRs), which are coherent oscillations of conductive electrons. LSPRs can be induced on the surface of noble metal (Ag or Au) nanostructures by illuminating the surfaces with electromagnetic irradiation. These noble metals can be coupled with catalytic metals, such as Pt, Pd, and Ru, to develop bimetallic nanostructures with unique catalytic activities. The plasmon-driven catalysis on bimetallic nanostructures is light-driven, which essentially enables green chemistry in organic synthesis. During the past decade, surface-enhanced Raman spectroscopy (SERS) has been actively utilized to study the mechanisms of plasmon-driven reactions on mono- and bimetallic nanostructures. SERS has provided a wealth of knowledge about the mechanisms of numerous plasmon-driven redox, coupling, and scissoring reactions. However, the nanoscale catalytic properties of both mono- and bimetallic nanostructures as well as the underlying physical cause of their catalytic reactivity and selectivity remained unclear for decades.In this Account, we focus on the most recent findings reported by our and other research groups that shed light on the nanoscale properties of mono- and bimetallic nanostructures. This information was revealed by tip-enhanced Raman spectroscopy (TERS), a modern analytical technique that has single-molecule sensitivity and subnanometer spatial resolution. TERS findings have shown that plasmonic reactivity and the selectivity of bimetallic nanostructures are governed by the nature of the catalytic metal and the strength of the rectified electric field on their surfaces. TERS has also revealed that the catalytic properties of bimetallic nanostructures directly depend on the interplay between the catalytic and plasmonic metals. We anticipate that these findings will be used to tailor synthetic approaches that are used to fabricate novel nanostructures with desired catalytic properties. The experimental and theoretical results discussed in this Account will facilitate a better understanding of TERS and explain artifacts that could be encountered upon TERS imaging of a large variety of samples. Consequently, plasmon-driven chemistry should be considered as an essential part of near-field microscopy.

Non-Invasive and Confirmatory Differentiation of Hermaphrodite from Both Male and Female Cannabis Plants Using a Hand-Held Raman Spectrometer.

Cannabis (Cannabis sativa L.) is a dioecious plant that produces both male and female inflorescences. In nature, male and female plants can be found with nearly equal frequency, which determines species out-crossing. In cannabis farming, only female plants are preferred due to their high yield of cannabinoids. In addition to unfavorable male plants, commercial production of cannabis faces the appearance of hermaphroditic inflorescences, species displaying both pistillate flowers and anthers. Such plants can out-cross female plants, simultaneously producing undesired seeds. The problem of hermaphroditic cannabis triggered a search for analytical tools that can be used for their rapid detection and identification. In this study, we investigate the potential of Raman spectroscopy (RS), an emerging sensing technique that can be used to probe plant biochemistry. Our results show that the biochemistry of male, female and hermaphroditic cannabis plants is drastically different which allows for their confirmatory identification using a hand-held Raman spectrometer. Furthermore, the coupling of machine learning approaches enables the identification of hermaphrodites with 98.7% accuracy, whereas both male and female plants can be identified with 100% accuracy. Considering the label-free, non-invasive and non-destructive nature of RS, the developed optical sensing approach can transform cannabis farming in the U.S. and overseas.

Infrared and Raman chemical imaging and spectroscopy at the nanoscale.

The advent of nanotechnology, and the need to understand the chemical composition at the nanoscale, has stimulated the convergence of IR and Raman spectroscopy with scanning probe methods, resulting in new nanospectroscopy paradigms. Here we review two such methods, namely photothermal induced resonance (PTIR), also known as AFM-IR and tip-enhanced Raman spectroscopy (TERS). AFM-IR and TERS fundamentals will be reviewed in detail together with their recent crucial advances. The most recent applications, now spanning across materials science, nanotechnology, biology, medicine, geology, optics, catalysis, art conservation and other fields are also discussed. Even though AFM-IR and TERS have developed independently and have initially targeted different applications, rapid innovation in the last 5 years has pushed the performance of these, in principle spectroscopically complimentary, techniques well beyond initial expectations, thus opening new opportunities for their convergence. Therefore, subtle differences and complementarity will be highlighted together with emerging trends and opportunities.

Non-Invasive Identification of Nutrient Components in Grain.

Digital farming is a modern agricultural concept that aims to maximize the crop yield while simultaneously minimizing the environmental impact of farming. Successful implementation of digital farming requires development of sensors to detect and identify diseases and abiotic stresses in plants, as well as to probe the nutrient content of seeds and identify plant varieties. Experimental evidence of the suitability of Raman spectroscopy (RS) for confirmatory diagnostics of plant diseases was previously provided by our team and other research groups. In this study, we investigate the potential use of RS as a label-free, non-invasive and non-destructive analytical technique for the fast and accurate identification of nutrient components in the grains from 15 different rice genotypes. We demonstrate that spectroscopic analysis of intact rice seeds provides the accurate rice variety identification in ~86% of samples. These results suggest that RS can be used for fully automated, fast and accurate identification of seeds nutrient components.

Structural Characterization of Individual α-Synuclein Oligomers Formed at Different Stages of Protein Aggregation by Atomic Force Microscopy-Infrared Spectroscopy.

Aberrant α-synuclein aggregation is strongly associated with the onset and development of Parkinson's disease (PD). Therefore, characterizing the structure of toxic intermediate oligomers plays an essential role in better understanding their neurotoxicity. Using atomic force microscopy-infrared spectroscopy (AFM-IR), we were able to reveal the structure of α-synuclein oligomers present at different stages of protein aggregation and establish a relationship between morphology and structure on the single oligomer level. We were also able to probe the secondary structure evolution of individual oligomers. Moreover, the IR spectra of individual oligomers suggest structural rearrangement that is necessary for oligomers with an antiparallel ß-sheet to propagate into fibrils that have a parallel-ß-sheet secondary structure. Detailed investigation of structural organization of α-synuclein oligomers reported in this study is critically important to understand the toxicity of these protein species. We also anticipate that this work will help developing approaches for oligomer detection and consequently presymptomatic diagnostic of PD.

Raman-Based Differentiation of Hemp, Cannabidiol-Rich Hemp, and Cannabis.

Hemp (Cannabis sativa) has been used to treat pain as far back as 2900 B.C. Its pharmacological effects originate from a large variety of cannabinols. Although more than 100 different cannabinoids have been isolated from Cannabis plants, clear physiological effects of only a few of them have been determined, including delta-9 tetrahydrocannabinol (THC), cannabidiol (CBD), and cannabigerol (CBG). While THC is an illicit drug, CBD and CBG are legal substances that have a variety of unique pharmacological properties such as the reduction of chronic pain, inflammation, anxiety, and depression. Over the past decade, substantial efforts have been made to develop Cannabis varieties that would produce large amounts of CBD and CBG. Ideally, such plant varieties should produce very little (below 0.3%) if any THC to make their cultivation legal. The amount of cannabinoids in the plant material can be determined using high performance liquid chromatography (HPLC). This analysis, however, is nonportable, destructive, and time and labor consuming. Our group recently proposed to use Raman spectroscopy (RS) for confirmatory, noninvasive, and nondestructive differentiation between hemp and cannabis. The question to ask is whether RS can be used to detect CBD and CBG in hemp, as well as enable confirmatory differentiation between hemp, cannabis, and CBD-rich hemp. In this manuscript, we show that RS can be used to differentiate between cannabis, CBD-rich plants, and regular hemp. We also report spectroscopic signatures of CBG, cannabigerolic acid (CBGA), THC, delta-9-tetrahydrocannabinolic acid (THCA), CBD, and cannabidiolic acid (CBDA) that can be used for Raman-based quantitative diagnostics of these cannabinoids in plant material.

Nanoscale Structural Characterization of Individual Viral Particles Using Atomic Force Microscopy Infrared Spectroscopy (AFM-IR) and Tip-Enhanced Raman Spectroscopy (TERS).

Viruses are infections species that infect a large spectrum of living systems. Although displaying a wide variety of shapes and sizes, they are all composed of nucleic acid encapsulated into a protein capsid. After virions enter the host cell, they replicate to produce multiple copies of themselves. They then lyse the host, releasing virions to infect new cells. The high proliferation rate of viruses is the underlying cause of their fast transmission among living species. Although many viruses are harmless, some of them are responsible for severe diseases such as AIDS, viral hepatitis, and flu. Traditionally, electron microscopy is used to identify and characterize viruses. This approach is time- and labor-consuming, which is problematic upon pandemic proliferation of previously unknown viruses, such as H1N1 and COVID-19. Herein, we demonstrate a novel diagnosis approach for label-free identification and structural characterization of individual viruses that is based on a combination of nanoscale Raman and infrared spectroscopy. Using atomic force microscopy-infrared (AFM-IR) spectroscopy, we were able to probe structural organization of the virions of Herpes Simplex Type 1 viruses and bacteriophage MS2. We also showed that tip-enhanced Raman spectroscopy (TERS) could be used to reveal protein secondary structure and amino acid composition of the virus surface. Our results show that AFM-IR and TERS provide different but complementary information about the structure of complex biological specimens. This structural information can be used for fast and reliable identification of viruses. This nanoscale bimodal imaging approach can be also used to investigate the origin of viral polymorphism and study mechanisms of virion assembly.