N-(1-naphthyl) Ethylenediamine Dihydrochloride Research Articles

Conformational stability, quantum computational (DFT), vibrational, electronic and non-covalent interactions (QTAIM, RDG and IGM) of antibacterial compound N-(1-naphthyl)ethylenediamine dihydrochloride

Naphthalene derivatives are heterocyclic compounds with significant applications in the pharmaceutical and biomedical industries. Several spectroscopy methods, including FT-IR, FT-Raman, and UV–Vis, were used in this study to examine the molecular structure of N-(1-Naphthyl) ethylenediamine dihydrochloride (NEDD). By employing DFT through the WB97XD with the level of 6–311++G(d, p) as the basis set, the optimized geometrical characteristics and full vibrational allocations of frequencies by the potential energy distribution were generated and matched to the experimental data. The delocalization of charge caused by intermolecular connections becomes apparent in the natural bond orbital (NBO) study. Additionally, frontier molecular orbital bandgap energy and electron excitation analyses were acquired to provide a deeper understanding of the structural characteristics of NEDD. Frontier molecular orbital energies revealed an energy band gap of 5.505 eV, demonstrating the existence of charge transfer within the molecule. Electronic transitions seen in the experimentally recorded UV–visible spectra were assigned using the TD-DFT method whereas transition density matrix (TDM) plots showed the ability to transmit charges. Considering wave function-derived parameters like the Laplacian of electron density (LED), local information entropy (LIE), the average localized ionization energy (ALIE) as well as non-covalent interactions, provides essential insights into the reactivity and stability traits of molecular systems. Local reactivity descriptions aided in understanding the interactions of physiologically relevant molecular systems, thereby facilitating the discovery of potential pharmaceutical compounds. Drug-likeness and ADMET assessment were achieved to assess biological properties of NEDD. Antibacterial assay confirmed its effectiveness against bacterial strains, while molecular docking studies. The stability of the best docked protein has been investigated using 70 ns molecular dynamics simulation followed by effective free energy calculation using MMPBSA.

MALDI Mass Spectrometry Imaging sample preparation with wet-interface matrix deposition for lipid analysis.

Sample preparation is one of the most crucial steps for MALDI mass spectrometry imaging (MALDI-MSI). The scientists beginning their adventure with this technique may be overwhelmed by the variety of matrices, solvents, concentrations, the ways of their applications, and the lack of widely available knowledge about the influence of these parameters on the results. Here we would like to present our experiences with detailed aspects of matrices deposition, hopefully helpful for the scientific community. In our article, we have tested several MALDI matrices applied by the SunCollect® system: wet-interface matrix deposition in the context of lipids analysis. Among them: 2,5-dihydroxybenzoic acid (DHB), norharmane, N-(1-naphthyl) ethylenediamine dihydrochloride (NEDC), 9-aminoacridine (9AA). We have optimized the number of matrix layers and nozzle settings in terms of spectra intensity and the overall quality of obtained ion maps. Our research presents the influence of the number of matrix layers and nozzle settings on the results and allows for choosing the optimal parameters for the analyses. In positive ionization mode, DHB matrix could be chosen as the first selection. In the negative ionization mode, DAN matrix produces higher peak intensity in a lower mass range and seems to provide more information than 9AA. We recommended NEDC for particular tasks such as glucose analysis. Comparably to remaining matrices, norharmane significantly changes received ion maps. Dealing with such a great amount of data allows us to notice an interesting conclusion the obtained ion picture for a particular ion could differ dramatically with changing the matrix, the solvent composition, or even the number of matrix layers. This must be considered when interpreting the result and forces us to compare the results obtained with different matrices with extreme caution.

Enhancing metabolite coverage in MALDI-MSI using laser post-ionisation (MALDI-2).

Matrix-assisted laser desorption/ionisation mass spectrometry imaging (MALDI-MSI) of metabolites can reveal how metabolism is altered throughout heterogeneous tissues. Here negative ion mode MALDI-MSI has been coupled with laser post-ionisation (MALDI-2) and applied to the MSI of low molecular weight (LMW) metabolites (<m/z 600) to investigate the benefits MALDI-2 offers for spatial metabolomics in terms of metabolite coverage and sensitivity. When applied to mouse kidney tissue MALDI-2 provided almost double the number of on-tissue specific mass features compared to conventional MALDI. MALDI-2 also resulted in not only the increased detection sensitivity for multiple metabolite species but also permitted the imaging of LMW metabolites (e.g. uridine) that were not detected using conventional MALDI-MSI. When compared against ∼140 publically available kidney datasets submitted through the METASPACE analysis platform using the same N-(1-naphthyl) ethylenediamine dihydrochloride (NEDC) matrix, MALDI-2 provided 34 unique metabolite m/z features that were not consistently annotated previously. To further evaluate the usefulness of this MALDI-2 approach to metabolite imaging, MALDI-2 was applied to the imaging of mouse liver tissue containing a metastasised breast cancer at a pixel size of 20 μm. Using a co-localisation analysis, MALDI-2 detected six tumour-specific metabolites that were not detected using conventional MALDI, as well as providing an up to 20-fold increase in signal intensities for many others (e.g., glutamate). This work provides one of the first reports of MALDI-2 applied to metabolite imaging and demonstrates the dramatic improvements in sensitivity and metabolite coverage it provides.

Facile Detection of Naphthalene with a 5,10,15,20-tetrakis(4-methoxyphenyl)-21H,23H-Porphine Nickel (II)/N-(1-Naphthyl) Ethylenediamine Dihydrochloride Renewable Graphene Oxide Paste Electrode

We report the fabrication of a new paste electrode based on 5,10,15,20-tetrakis(4- methoxyphenyl)−21H,23H-porphine nickel (II) (NiTPP) and N-(1-Naphthyl)ethylenediamine dihydrochloride (N-NEDDH) for the electrochemical detection of naphthalene. The electrochemical behaviour of the modified N-NEDDH/NiTPP/GO/CPE was studied by cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS) and differential pulse voltammetry (DPV). Under the optimal conditions, the modified sensor presented a wide linear range from 1 × 10−8 to 1 × 10−5 mol l−1 in the detection of naphthalene, with a 3 × 10−9 mol l−1 detection limit. Other polycyclic aromatic hydrocarbons (PAHs), like phenanthrene, anthracene, fluorene and pyrene showed little interference on the detection. It also demonstrated a good applicability on naphthalene determination from four types of water samples, with good recovery rates.

Mapping glucose metabolites in the normal bovine lens: Evaluation and optimisation of a matrix-assisted laser desorption/ionisation imaging mass spectrometry method.

The spatial resolution of microdissection-based analytical methods to detect ocular lens glucose uptake, transport and metabolism are poor, whereas the multiplexing capability of fluorescence microscopy-based approaches to simultaneously detect multiple glucose metabolites is limited in comparison with mass spectrometry-based methods. To better understand lens glucose transport and metabolism, a more highly spatially resolved technique that maintains the fragile ocular lens tissue is required. In this study, a sample preparation method for matrix-assisted laser desorption/ionisation imaging mass spectrometry (MALDI IMS) analysis of ocular lens glucose uptake and metabolism has been evaluated and optimised. Matrix choice, tissue preparation and normalisation strategy were determined using negative ion mode MALDI-Fourier transform-ion cyclotron resonance MS of bovine lens tissue and validation performed using gas chromatography-MS. An internal standard was applied concurrently with N-(1-naphthyl)ethylenediamine dihydrochloride (NEDC) matrix to limit cracking of the fresh frozen lens tissue sections. MALDI IMS data were collected at a variety of spatial resolutions to detect both endogenous lens metabolites and stable isotopically labelled glucose introduced by ex vivo lens culture. Using this approach, initial steps in important metabolic processes that are linked to diabetic cataract formation were spatially mapped in the bovine lens. In the future, this method can be applied to study the dynamics of glucose uptake, transport and metabolic flux to aid in the study of diabetic lens cataract pathophysiology.

Direct identification of forensic body fluids by MALDI-MS.

The rapid identification of human body fluids is meaningful for forensic casework. However, current methods suffer from several limitations such as poor sensitivity, time consumption and big sample consumption. Herein, we developed a mass spectrometry method to distinguish human body fluids (blood, semen, urine, sweat, and saliva) based on small molecular regions with no pretreatment, microliter sample consumption and high throughput. A highly sensitive and high salt-tolerance matrix N-(1-naphthyl)ethylenediamine dihydrochloride (NEDC) was used to efficiently detect metabolites in complex humoral environment. Some characteristic small metabolic molecules such as heme, hemin, creatinine, phosphate acid, uric acid, citric acid and lactic acid were identified and served as potential biomarkers to differentiate different body fluid types. Further principal component analysis (PCA) was performed to cluster the body fluid samples and three principal components allowed 75% clustering of all body fluid types. Blind testing revealed that nine out of ten unknown body fluid samples could be correctly classified into their corresponding group. This novel method can efficiently differentiate five body fluids with minimal interferences due to the storage time (less than 12 months) and carrier materials (cotton, fabric and tissue). The whole process from sampling to recording of mass spectra of body fluids can be finished in less than 10 minutes. We believe that this developed strategy has significant implications for rapid and effective human body fluid screening in forensic casework.

Differentiation and Relative Quantitation of Disaccharide Isomers by MALDI-TOF/TOF Mass Spectrometry.

Saccharide isomer differentiation has been a challenge in glycomics, as the lack of technology to decipher fully the diverse structures of compositions, linkages, and anomeric configurations. Several mass spectrometry-based methods have been applied to the discrimination of disaccharide isomers, but limited quantitative analyses have been reported. In the present study, MALDI-LIFT-TOF/TOF has been investigated to differentiate and relatively quantify underivatized glucose-containing disaccharide isomers that differ in composition, connectivity or configuration. N-(1-naphthyl)ethylenediamine dihydrochloride (NEDC) was used as a highly sensitive matrix without matrix interferences in low mass range, thus yielding intense chloride-attached disaccharide ions [M + Cl]-, which could be fragmented to give diagnostic characteristic fragment patterns for distinguishing these isomers. Three different types of disaccharide isomers were successfully relatively quantified in a binary mixture using the specific product ion pairs. Finally, this method was utilized to identify and relatively quantify two disaccharide isomers in Medicago leaf (maltose and sucrose) without numerous preparation steps. In general, this method is a fast, effective, and robust method for rapid differentiation and quantitation of disaccharide isomers in complex medium.

Enhanced coverage of lipid analysis and imaging by matrix-assisted laser desorption/ionization mass spectrometry via a strategy with an optimized mixture of matrices

In matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS) analysis and imaging of lipids, comprehensive ionization of lipids simultaneously by a universal matrix is a very challenging problem. Ion suppression of readily ionizable lipids to others is common. To overcome this obstacle and enhance the coverage of MALDI MS analysis and imaging of lipids, we developed a novel strategy employing a mixture of matrices, each of which is capable of selective ionization of different lipid classes. Given that MALDI MS with either 9-aminoacridine (9-AA) or N-(1-naphthyl) ethylenediamine dihydrochloride (NEDC) yields weak in-source decay which is critical for analysis of complex biological samples and possesses orthogonal selectivity for ionization of lipid classes, we tested the mixtures of NEDC and 9-AA with different ratios for analysis of standard lipids and mouse brain lipid extracts. We determined 1.35 of NEDC/9-AA as an optimized molar ratio. It was demonstrated that an enhanced coverage with the optimized mixture was obtained, which enabled us to analyze and map all the major classes of phospholipids and sulfatide from either lipid extracts or tissue slides, respectively. We believe that this powerful novel strategy can enhance lipidomics analysis and MALDI MS imaging of lipids in a high-throughput and semi-quantitative fashion.

Electric Field-Assisted Matrix Coating Method Enhances the Detection of Small Molecule Metabolites for Mass Spectrometry Imaging.

Small molecule metabolites (SMMs, typically <500 Da) are important cellular constituents closely associated with tumor development and progression. However, in situ label-free detection of tissue SMMs has been limited due to interference from matrix and/or low sensitivity. Herein, we develop an electric field-assisted scanning-spraying (EFASS) matrix coating system to deposit matrix on tissue with crystal sizes of <10 μm, followed by matrix-assisted laser desorption/ionization-mass spectrometry imaging (MALDI-MSI) in negative ion mode. A comparison with other matrix deposition methods (i.e., airbrush and sublimation) using common matrixes (i.e., N-(1-naphthyl) ethylenediamine dihydrochloride (NEDC), 9-aminoacridine (9-AA), 2,5-dihydroxybenoic acid (DHB)) indicated that the EFASS system could effectively enhance detection sensitivity and the number of tissue SMMs detected. MSI of five gastric cancer tissues coated with NEDC by the EFASS system demonstrated that significantly increased levels of fatty acids (i.e., palmitic acid and oleic acid) and nucleosides monophosphate (i.e., uridine monophosphate, adenosine monophosphate, and guanosine monophosphate) and significantly decreased levels of nucleosides (i.e., inosine, guanosine, and uridine) and N-acetylneuraminic acid were observed in cancerous areas.

MALDI-TOF MS imaging of metabolites with a N-(1-naphthyl) ethylenediamine dihydrochloride matrix and its application to colorectal cancer liver metastasis.

Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI) is a label-free technique for identifying multiplex metabolites and determining both their distribution and relative abundance in situ. Our previous study showed that N-(1-naphthyl) ethylenediamine dihydrochloride (NEDC) could act as a matrix for laser desorption/ionization time-of-flight mass spectrometry (LDI-TOF MS) detection of oligosaccharides in solution. In the present study, NEDC-assisted LDI-TOF MSI yielded many more endogenous compound peaks between m/z 60 and m/z 1600 than 9-aminoacridine (9-AA). Our results show that NEDC-assisted LDI-TOF MSI is especially well-suited for examining distributions of glycerophospholipids (GPs) in addition to low molecular weight metabolites below m/z 400. Particularly, NEDC matrix allowed the LDI-TOF MSI of glucose in animal tissue. Furthermore, NEDC-assisted LDI-TOF MSI was applied to a mouse model of colorectal cancer liver metastasis. We revealed the distinct spatio-molecular signatures of many detected compounds in tumor or tumor-bearing liver, and we found that taurine, glucose, and some GPs decreased in tumor-bearing liver as the tumor developed in liver. Importantly, we also found a glucose gradient in metastatic tumor foci for the first time, which further confirms the energy competition between tumors and liver remnant due to the Warburg effect. Our results suggest that NEDC-assisted LDI MSI provides an in situ label-free analysis of multiple glycerophospholipids and low molecular weight metabolites (including glucose) with abundant peaks and high spatial resolution. This will allow future application to in situ definition of biomarkers, signaling pathways, and disease mechanisms.

Quantitative Analysis of Oligosaccharides Derived from Sulfated Glycosaminoglycans by Nanodiamond-Based Affinity Purification and Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry

Degraded fragments of sulfated glycosaminoglycans (GAGs) are key reporters for profiling the burden of mucopolysaccharidosis (MPS) disease at baseline and during therapy. Here, we present a high-throughput assay, which combines microwave-assisted degradation, solid-phase affinity purification, and matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS), for quantitative analysis of sulfated oligosaccharides in biological samples. First, sulfated oligosaccharides such as chondroitin-4-sulfate (CS) were efficiently isolated from highly diluted solutions or spiked artificial cerebrospinal fluid (aCSF) using polyarginine-coated nanodiamonds (PA-coated NDs) as affinity sorbents. Next, they were degraded to disaccharides through microwave-assisted methanolysis or enzymatic digestion for subsequent MALDI-TOF MS analysis. The reaction times for GAG depolymerization were significantly reduced from a few hours to less than 7 min under the microwave irradiation. Deuterium-labeled internal standards were then mixed with the CS-derived disaccharides for quantitative analysis by MALDI-TOF MS using the N-(1-naphthyl) ethylenediamine dihydrochloride (NEDC) matrix. The new assay is facile, specific (with distinct chlorine-isotope trait markers), sensitive (with a detection limit of ~70 pg), and potentially useful for clinical diagnosis of MPS.

High-Salt-Tolerance Matrix for Facile Detection of Glucose in Rat Brain Microdialysates by MALDI Mass Spectrometry

Due to its strong ultraviolet absorption, high salt tolerance, and little interference in the low molecular weight region, N-(1-naphthyl) ethylenediamine dihydrochloride (NEDC) has been applied as a matrix to measure the level of glucose in rat brain microdialysates by matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS) in combination with in vivo microdialysis. By monitoring the ion signals of (glucose + Cl)(-) in the mass spectra, we achieved a low detection limit of ~10 μM for glucose in 126 mM NaCl, which is a typical component in artificial cerebrospinal fluid, without prior sample purification. It is concluded that NEDC-assisted laser desorption/ionization (LDI) MS is a fast and general method for sensitive detection of small molecules (such as glucose and amino acids) in high ionic strength solutions.

Modification of AOAC Method 973.31 for Determination of Nitrite in Cured Meats

A modification of AOAC Method 973.31 is proposed to improve the extraction efficiency of nitrite from cured meat samples and its subsequent quantification based on the diazotization-coupling reaction of sulfanilamide with N-(1-naphthyl)ethylenediamine dihydrochloride (NED). The various experimental parameters were thoroughly investigated. A 5 g meat sample was mixed with 400 mL water; the pH of the mixture was adjusted to 5.5 +/- 0.3 and allowed to stand for 2 h on a water bath at 80 degrees C, with occasional shaking for the complete extraction of nitrite. After quantitative filtration, an aliquot was mixed with chloroacetic-chloroacetate buffer, pH 1.80 +/- 0.05, sulfanilamide, and NED, and the absorbance of the resulting azodye was recorded at 540 nm against water as a reference. Following the recommended procedure, a linear calibration graph was obtained for up to 0.8 microg/mL NO2(-), with a correlation coefficient of 0.9996 and a detection limit (based on the 3 Sb-criterion) of 5.6 ng/mL NO2(-). The proposed method was conveniently applied to various cured meat samples and was validated by comparison with the original AOAC method and by recovery experiments that gave quantitative results (94-98%) with convenient reproducibility. Statistical analysis of the analytical data could not detect any systematic error and revealed the high accuracy and precision of the proposed method.

Development of a Detection Tablet for a Portable NO2 Monitoring System

We have already developed a HCHO monitoring system which is called FP-30. In this experiment, we have developed a NO(2) detection tablet which can be used by the monitoring system. The detection tablet for the NO(2) was constructed with the sensing paper: porous cellulose paper that contains silica gel as an adsorbent, N-1-naphthylethylenediamine dihydrochloride (NED), and glycerin. The NO(2) in sample gas was blown over and adsorbed on the surface of the sensing paper. Then the NO(2) reacted with NED, producing a yellow compound. The coloring reaction took place on the surface of the sensing paper. The degree of color change of paper from white to yellow was monitored as a function of the intensity of the reflected light (lambda = 475 nm) of an LED. The detection limit was 0.01 ppm when the sampling time was 30 min, and the flow rate of sample gas was 250 ml/min. This sensing paper process was not interfered with by acetaldehyde, acetone, alcohols, hydrocarbons, carbon monoxide or carbon dioxide. The NO(2) concentrations in the rooms of a house or school were monitored using this monitoring system and the standard chemiluminescence method. The concentrations of NO(2) monitored by both methods were within 18% of the average. This highly sensitive, selective, and handy NO(2) gas monitoring system will be widely applicable and convenient for users who are not specialists in this field.

Studies on the self-assembly behavior of the amphiphilic block copolymer of PSt-b-PAA in apolar solvents with polar fluorescent probe

The block copolymer of polystyrene-b-poly(butyl acrylate) (PSt-b-PBA) with a well-defined structure was synthesized by atom transfer radical polymerization (ATRP); its structure was characterized, and the living polymerization was also validated by gel permeation chromatography, Fourier transform infrared, and 1H NMR measurements. Then, the amphiphilic block copolymer of polystyrene-b-poly(acrylic acid) (PSt-b-PAA) has been prepared by hydrolysis of PSt-b-PBA, and copolymers of PSt-b-PAA with longer PSt blocks and shorter PAA blocks were obtained by controlling the conditions of ATRP polymerization. The reversed micelle solution of PSt-b-PAA in toluene was prepared by using the single-solvent dissolving method, and the reverse micellization behavior of PSt-b-PAA in toluene was mainly investigated in this paper. The fluorescent probe technique was used by using polar fluorescence compound N-(1-Naphthyl)ethylenediamine dihydrochloride (NEAH) as a polar fluorescent probe to study the reverse micellization behavior of PSt-b-PAA. It was found that the reverse micellization behaviors of PSt-b-PAA in toluene can be clearly revealed by using NEAH as a polar fluorescence probe, and the critical micelle concentrations (cmcs) can be well displayed. The experimental results showed that the self-assembling behavior of PSt-b-PAA in toluene depends apparently on the microstructure of the macromolecules and is also influenced by the temperature. For the copolymers of PSt-b-PAA with the same length of hydrophobic PSt blocks, the copolymer with a longer hydrophilic block PAA has lower cmc, and at higher temperature, the copolymer has lower cmc.

Spectrophotometric determination of azathioprine in pharmaceutical formulations

Four simple and sensitive visible spectrophotometric methods (A–D) have been described for the assay of azathioprine (ATP) either in pure form or in pharmaceutical formulations. Methods A and B are based on the oxidation of ATP with excess N-bromosuccinimide (NBS) or chloramine-T (CAT) and determining the consumed NBS or CAT with a decrease in colour intensity of celestine blue (CB) (method A) or gallocyanine (GC) (method B), respectively. Methods C and D are based on the diazotisation of reduced azathioprine (RATP) with excess nitrous acid and estimating either the consumed nitrous acid (HNO 2) with cresyl fast violet acetate (CFVA) (method C) or by coupling reaction of the diazonium salt formed with N-1-naphthyl ethylene diamine dihydrochloride (NED) (method D). All of the variables have been optimized and the reactions presented. The concentration measurements are reproducible within a relative standard deviation of 1.0%. Recoveries are 99.2–100.3%.

A submersible flow injection-based sensor for the determination of total oxidised nitrogen in coastal waters

The design and construction of a remotely deployed submersible sensor for the determination of total oxidised nitrogen (nitrate plus nitrite) in seawater is described. It was based on the flow injection principle with solid state spectrophotometric detection of the diazotisation product from the reaction of nitrite with N-(1-naphthyl)ethylenediamine dihydrochloride (N1NED) and sulphanilamide. Nitrate was pre-reduced in-line with a copperised cadmium powder column. The limit of detection (3 s) was 0.1 μM nitrate (1.4 μg l −1 N) and the linear range was 0.1–55 μM nitrate with a 20 mm path length flow cell and a 260 μl sample loop. The linear range was modified for different environmental conditions by changing the sample loop size and - or the flow cell path length. Results are presented for preliminary laboratory studies and field trials in the Tamar Estuary and for deployments in the North Sea. The sensor was successfully deployed on three separate occasions for complete tidal cycles (13 h) at a depth of 4 m from the bow of RRS Challenger. Field data are compared with results from a shipboard air segmented analyser and correlated with turbidity and salinity data.

4-Chloro-6-methoxyindole is the precursor of a potent mutagen (4-chloro-6-methoxy-2-hydroxy-1-nitroso-indolin-3-one oxime) that forms during nitrosation of the fava bean (Vicia faba).

Fava beans (Vicia faba) upon treatment with nitrite under simulated gastric conditions, form a direct-acting bacterial mutagen, comparable in specific activity to the most potent known mutagens for several strains of Salmonella typhimurium. The precursor of the mutagen was isolated and identified as 4-chloro-6-methoxyindole by u.v., i.r., m.s. and n.m.r. The precursor was dechlorinated with NaBH4 and PdCl2 as the catalyst and the product obtained from this reaction was identified as 6-methoxyindole. Since synthetic 4-chloro-6-methoxyindole was not available, structure activity studies were conducted on substituted indoles. Nitrosation of 4-chloroindole closely follows the results for nitrosation of 4-chloro-6-methoxyindole. The major product of nitrosation of 4-chloroindole is 4-chloro-2-hydroxy-N1-nitroso-indolin-3-one oxime. Thus, it appears that the major nitrosation product of 4-chloroindole and of 4-chloro-6-methoxyindole is a stable alpha-hydroxy N-nitroso compound. This is the first reported case of stable alpha-hydroxy N-nitroso compounds. In the presence of N-(1-naphthyl)ethylenediamine dihydrochloride (NEDD), the alpha-hydroxy N-nitroso compound rearranges to an aromatic diazonium ion which couples with the diamine to form an azo dye. Studies on nitrosation kinetics indicate that the nitrosation of indoles are relatively fast reactions. Both the structural and rate studies give strong support to the hypothesis that intragastric nitrosation of fava beans yield the putative gastric carcinogen in the high-risk area in Colombia.