Derivatization Method Research Articles

Determination of biogenic amines in chicken, beef, and mutton by dansyl chloride microwave derivatization in Malaysia

In this study, an improved dansyl-chloride derivatization technique using a microwave synthesizer was used for the qualitative and quantitative analyses of biogenic amine in the fresh meat samples. The derivatization technique was optimized in terms of temperature, reaction time, and spinning speed. The derivatization method together with a validated reversed-phase HPLC-DAD method was used for the determination of biogenic amines in chicken, beef, and mutton sold in the wet market. The results of the analyses showed that tryptamine, putrescine, and histamine were generally detected in all the three types of meat. Higher levels of histamine were found in chicken and beef. However, low levels of histamine were observed in mutton. Tyramine was either detected low or moderate in all the three types of meat. The biogenic amines of the fresh meat sold in the wet market is generally higher than the reported values. The mechanisms of biogenic amines-dansyl-chloride formation were investigated and proposed. PRACTICAL APPLICATION: The biogenic amine derivatization method was improved. The improved derivatization method can be potentially used for various food products beside meats for routine biogenic amine analyses due to its fast analysis time and simplicity. High levels of biogenic amines were generally found in the meat sold in the wet markets. However, proper handling of the raw meat can reduce the risk of infection.

Ultrasensitive quantification of estrogens in serum and plasma by liquid chromatography-tandem mass spectrometry.

Stable isotope dilution (SID) methodology coupled with liquid chromatography-tandem mass spectrometry (LC-MS) is rapidly becoming the gold standard for measuring estrogens in serum and plasma due to improved specificity, high accuracy, and the ability to conduct a more comprehensive analysis. A general consideration of the problems associated with measuring estrogens and two detailed derivatization methods are described in this chapter. These methods quantify estrogens and their metabolites in serum and plasma samples using this state-of-art technology, which is applicable to the routine clinical laboratory.

An effective ultra‐performance liquid chromatography and derivatization method for the quantification of potential genotoxic impurity Hydrazine in Gliclazide and its formulation – Robustness study by the design of experiments

AbstractThe current study presents an accurate, precise, and highly selective ultra‐performance liquid chromatography derivatization method for the quantification of Hydrazine in Gliclazide and its formulations. Gliclazide, derivatizing agent‐ 3,4,5‐trimethoxy benzaldehyde, and Hydrazine derivatives were separated on YMC Pack pro C18 column (50 × 3.0 mm id, 2 μm), column oven temperature maintained at 40°C, with flow rate 0.6 ml/min on ultra‐performance liquid chromatography system with gradient elution monitored at 333 nm. 0.1% trifluoroacetic acid and acetonitrile were used as mobile phase‐A and mobile phase‐B. Evaluated the robustness of the optimized method by employing a quality‐by‐design‐based design of experiments. The method was successfully validated as per the International Conference on Harmonization Q2 (R1) guideline and the United States Pharmacopeia < 1225 > general chapter. The United States Pharmacopeia resolution between the Gliclazide and Hydrazine derivative was 3.6. The limit of detection and limit of quantitation for Hydrazine derivative are 0.10 μg/ml and 0.30 μg/ml, respectively. The obtained regression coefficient (R2) was > 0.9998, and the %recoveries from the limit of quantitation to 150% level were 96.9%–100.8% from the linearity and accuracy studies, respectively. The method validation data and quality‐by‐design‐based robustness study indicate that the developed derivatization method is suitable for routine use in Quality Control laboratories.

A highly sensitive pre-column derivatization method for fluorescence detection of tobramycin using 2-naphthalenesulfonyl chloride as a derivatizing agent

A new, simple, rapid and highly sensitive high performance liquid chromatographic method was developed for the determination of tobramycin (TOB) in ophthalmic solutions. The developed method was based on derivatizing this non-chromophoric drug with 2-naphthalenesulfonyl chloride (NSCl), at 80 ºC for 40 min, and measuring the fluorescent product. Elution from the column was isocratic, using a mobile phase consisting of acetonitrile –1% phosphoric acid in water, (90:10, v/v), with a flow rate of 2 mL/min, at 40 ºC. Determination was developed using fluorescence detection at an excitation wavelength of 300 nm and an emission wavelength of 380 nm. The chromatographic separation was achieved on Knauer-Eurospher 100–5 C18 silica column (300 mm × 4 mm i.d, 5 μm). Different approaches were used to optimize the conditions of the method and to reach the maximum yield. The linear regression data for the calibration plot showed acceptable correlation coefficient (r2 = 0.9998) at the concentration ranges of 2.5–1000 ng/mL for TOB. The method was successfully validated according to the guidelines of the International Council on Harmonization, and was found suitable for pharmaceutical applications. The method was new, simple and precise for analysis of TOB and applicable for routine analysis in quality control laboratories.

Not all bad: Gyromitrin has a limited distribution in the false morels as determined by a new ultra high-performance liquid chromatography method

ABSTRACT Gyromitrin (acetaldehyde N-methyl-N-formylhydrazone) and its homologs are deadly mycotoxins produced most infamously by the lorchel (also known as false morel) Gyromitra esculenta, which is paradoxically consumed as a delicacy in some parts of the world. There is much speculation about the presence of gyromitrin in other species of the lorchel family (Discinaceae), but no studies have broadly assessed its distribution. Given the history of poisonings associated with the consumption of G. esculenta and G. ambigua, we hypothesized that gyromitrin evolved in the last common ancestor of these taxa and would be present in their descendants with adaptive loss of function in the nested truffle clade, Hydnotrya. To test this hypothesis, we developed a sensitive analytical derivatization method for the detection of gyromitrin using 2,4-dinitrobenzaldehyde as the derivatization reagent. In total, we analyzed 66 specimens for the presence of gyromitrin over 105 tests. Moreover, we sequenced the nuc rDNA internal transcribed spacer region ITS1-5.8S-ITS2 (ITS barcode) and nuc 28S rDNA to assist in species identification and to infer a supporting phylogenetic tree. We detected gyromitrin in all tested specimens from the G. esculenta group as well as G. leucoxantha. This distribution is consistent with a model of rapid evolution coupled with horizontal transfer, which is typical for secondary metabolites. We clarified that gyromitrin production in Discinaceae is both discontinuous and more limited than previously thought. Further research is required to elucidate the gyromitrin biosynthesis gene cluster and its evolutionary history in lorchels.

Твердофазные сорбенты на основе синтетических нановолокон и глауконита для извлечения магнезона I из водных сред

A comparative evaluation of the sorption capacity of synthetic and natural sorbents with respect to phenol and some of its derivatives has been carried out. Electrospun nanofibers based on polyamide-6 (PA-6) were obtained under optimal conditions from a spinning solution of a polymer dissolved in a mixture of formic and acetic acids. The preparation of the clay mineral by the sieve method has been carried out, while the enriched fraction of glauconite (200–500 microns) has been isolated. The morphology of unmodified nanofibers, as well as glauconite, has been studied by scanning electron microscopy. Preliminary experiments showed low efficiency of sorption of phenol and some of its nitro- and chlorine derivatives by the sorbents used in this work. To improve the sorption characteristics, the method of derivatization of initialanalytes by diazotization and azo coupling reactions has been used. The concentrations of magneson I (nitrophenylazo-derivative of resorcinol) have been determined spectrophotometrically using a calibration dependence in the range of magneson I molar concentrations from 3·10-6 to 2·10-5 mol/l. The sorption capacity of glauconite from the Saratov region (Beloozerskoye deposit) and synthetic nanofibers based on PA-6 with respect to magneson I has been studied. The effect of pH on the sorption efficiency of magnezone I by glauconite and nanofibers based on PA-6 has been evaluated, in addition, the extraction coefficients of magnezone I by these sorbents have been calculated, which were in the range of 88–100% at optimal pH (≈3–6). The experiments performed on a model compound, magnesone I, led to the conclusion about the potential possibility of using glauconite from the Beloozerskoye deposit in the Saratov region and nanofibers based on polyamide-6 for the extraction of phenols in the form of their azo derivatives from various waters.

The Use of Polydialkylsiloxanes/Triflic Acid as Derivatization Agents in the Analysis of Sulfur-Containing Aromatics by “Soft”-Ionization Mass Spectrometry

Polycyclic aromatic sulfur-containing compounds are widely distributed in oil, especially in its low-volatile and heavy fractions (resins, asphaltenes), and this dictates the need for their determination when reliable methods for sulfur removing, cleaning and processing oil are developed. In these cases, "soft" ionization mass spectrometry methods, based on electrospray ionization (ESI) and matrix-assisted laser desorption/ionization (MALDI), are particularly effective. However, aromatic sulfur-containing compounds have low polarity and cannot be readily ionized by these methods. To overcome the problem, their preliminary conversion into sulfonium salts by the action of alkyl iodides and a silver-containing agent is widely used. In the process of developing more economical derivatization methods, we found a rather unexpected possibility of implementing S-alkylation of organic sulfides with commercial polydialkylsiloxanes (alkyl = CH3 or C2H5) in the presence of triflic acid (CF3SO3H) as a superacid co-alkylating agent. For homologous dibenzothiophenes as a typical model representative of petroleum sulfur-containing aromatic compounds, ESI and MALDI mass spectra exhibited the signals of corresponding S-alkylsulfonium salts with a high signal-to-noise ratio. A rational mechanism for the described chemical transformation is proposed, including the indispensable activation by triflic acid and the cleavage of the Si-C bond. Specific collision-induced dissociation of corresponding S-alkylated sulfonium cations is considered. The applicability of the derivatization approach to the analysis of petroleum products by high-resolution mass spectrometry is demonstrated.

Bimetallic MOF-derived CeO2/Co3O4 microflowers with synergy of oxygen vacancy and p-n heterojunction for high-performance n-butanol sensors

In this work, CeO 2 /Co 3 O 4 microflowers have been prepared by a facile bimetallic metal-organic framework (MOF) derivatization method. By optimizing the calcination temperature, the CeO 2 /Co 3 O 4 -350 with the calcination temperature as 350 ℃ has been prepared with the optimal morphology and gas sensing performance. The microflowers composed of nanosheet subunits can fully expose the surface-active sites of CeO 2 /Co 3 O 4 -350, expanding its contact surface with the detected gas. Moreover, for the larger atomic radius and polyvalent state of Ce, more oxygen vacancies can be generated on the surface of CeO 2 /Co 3 O 4 -350 composites. And the p-n heterojunction formed at the junction of CeO 2 and Co 3 O 4 can expand the depletion layer at the interfaces, increasing the amount of adsorbed oxygen, which conduce to heighten the response of CeO 2 /Co 3 O 4 -350. Benefiting from the synergy of oxygen vacancy and p-n heterojunction, CeO 2 /Co 3 O 4 -350 exhibits excellent sensing properties toward n-butanol. At the operating temperature of 190°C, CeO 2 /Co 3 O 4 -350 can make a fast response/recovery (63 s/11 s) to 100 ppm n-butanol with a response value of 87.96, and the lowest limit (LOD) of detection is 2 ppm. Moreover, the theoretical detection limit of CeO 2 /Co 3 O 4 -350 to n-butanol has been evaluated to be 105 ppm, which means that it can respond to ppb-level n-butanol. After 15 days long-term test, the response still maintains about 94%, showing good stability of CeO 2 /Co 3 O 4 -350, which is of great significance to the practical application of gas sensors. Our work uses simple MOF-derived method to construct sensing materials with larger active surface and optimized internal electronic structure, providing a facile and practical strategy to design gas sensing materials. • Bimetallic MOF-derived CeO 2 /Co 3 O 4 microflowers with p-n heterojunction and rich oxygen vacancy were synthesized. • The CeO 2 /Co 3 O 4 sensor with high-performance n-butanol response can be obtained by optimizing calcination temperature. • The excellent sensing performance of CeO 2 /Co 3 O 4 is attributed to the synergy between oxygen vacancy and p-n heterojunction.

Whole blood and plasma taurine reference intervals in adult Cavalier King Charles Spaniels and correlations between taurine concentration, diet and mitral valve disease.

To determine breed-specific reference intervals for whole blood (WB) and plasma taurine concentrations in adult, overtly healthy Cavalier King Charles Spaniels (CKCSs) and determine whether taurine concentrations differ across preclinical myxomatous mitral valve disease (MMVD) stages or between CKCSs eating diets that meet World Small Animal Veterinary Association (WSAVA) nutritional guidelines versus other diets. 200 privately owned CKCSs. Clinically healthy adult CKCSs were recruited prospectively. Diet and supplement history was collected. Dogs were staged by echocardiography using MMVD consensus guidelines. Taurine concentrations were measured in deproteinized lithium heparin WB and plasma samples with the postcolumn ninhydrin derivatization method on a dedicated amino acid analyzer. There were 12 stage A (6%), 150 stage B1 (75%), and 38 stage B2 (19%) CKCSs. Seventy-eight dogs (39%) were reported by their owners to be eating diets meeting WSAVA nutritional guidelines; 116 (58%) were not. Taurine concentrations in plasma (P = .444) and WB (P = .073) were not significantly different across MMVD stages or between CKCSs eating diets meeting WSAVA nutritional guidelines versus other diets (P = .345 and P = .527, respectively). Reference intervals for WB taurine (152 to 373 µM) and plasma taurine (51 to 217 µM) concentrations in CKCSs were generated. In CKCSs, taurine concentrations do not differ significantly based on preclinical MMVD stage, nor do they differ significantly based on consumption of a diet that does or does not meet WSAVA nutritional guidelines.

Pyrylium-Based Derivatization for Rapid Labeling and Enhanced Detection of Cholesterol in Mass Spectrometry Imaging.

Cholesterol in the central nervous system has been increasingly found to be closely related to neurodegenerative diseases. Defects in cholesterol metabolism can cause structural and functional disorders of the central nervous system. The detection of abnormal cholesterol is of great significance for the cognition of physiological and pathological states of organisms, and the spatial distribution of cholesterol can also provide more clues for our understanding of the complex mechanism of disease. Here, we developed a novel pyrylium-based derivatization reagent combined with matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) to visualize cholesterol in biological tissues. A new class of charged hydroxyl derivatization reagents was designed and synthesized, and finally 1-(carboxymethyl)-2,4,6-trimethylpyridinium (CTMP) was screened for tissue derivatization of cholesterol. Different from the shortcomings of traditional hydroxyl labeling methods such as harsh reaction conditions and long reaction time, in our study, we combined the advantages of CTMP itself and the EDCl/HOBt reaction system to achieve instant labeling of cholesterol on tissues through two-step activation. In addition, we also reported changes in cholesterol content in different stages and different brain regions during disease development in SOD1 mutant mouse model. The cholesterol derivatization method we developed provides an efficient way to explore the distribution and spatial metabolic network of cholesterol in biological tissues.

Single pot in situ aqueous derivatization and subsequent determination of streptomycin and dihydrostreptomycin residues in honey by means of mass spectrometry

Streptomycin (STR) and dihydrostreptomycin (DSTR) are the typically encountered aminoglycoside (AMG) residues in honey. For AMG analysis, studies in literature involve impractical and expensive applications such as ion-pairing chromatography, immunoassays, pre and post column derivatizations, or SPE approaches. Pretreatments of these methods are toilsome and costly. Herein, one-pot, aqueous in-situ derivatization method was presented as a superior protocol. Time and cost-efficient UHPLC-MS/MS method has been developed, and practical sample preparation was introduced. Satisfactory results were reported in method verification studies. The mean recovery values were 102.6% for STR and 101.3% for DSTR. Average values between 1.5% and 9.9% RSDs were found at intra and inter-day precisions. CCα (5.7 and 5.8 μg/kg) and CCβ (6.2 and 6.4 μg/kg) values were calculated for STR and DSTR respectively. AMG residues were found in 29 out of 110 analyzed samples using validated method. Described novelty enabled comprehensive analysis in an inexpensive and straightforward manner.

Life Underground: Investigating Microbial Communities and Their Biomarkers in Mars‐Analog Lava Tubes at Craters of the Moon National Monument and Preserve

AbstractCraters of the Moon National Monument and Preserve (CotM) is a strong terrestrial analog for lava tube formations on Mars. The commonality of its basalt composition to Martian lava tubes makes it especially useful for probing how interactions between water, rock, and life have developed over time, and what traces of these microbial communities may be detectable by current flight‐capable instrumentation. Our investigations found that secondary mineral deposits within these caves contain a range of underlying compositions that support diverse and active microbial communities. Examining the taxonomy, activity, and metabolic potential of these communities revealed largely heterotrophic life strategies supported by contributions from chemolithoautotrophs that facilitate key elemental cycles. Finally, traces of these microbial communities were detectable by flight‐capable pyrolysis and wet chemistry gas chromatography‐mass spectrometry methods comparable to those employed by the Sample Analysis at Mars (SAM) instrument aboard the Curiosity rover and the Mars Organic Molecule Analyzer (MOMA) on the upcoming Rosalind Franklin rover. Using a suite of methods for chemical derivatization of organic compounds is beneficial for resolving the greatest variety of biosignatures. Tetramethylammonium hydroxide (TMAH), for example, allowed for optimal resolution of long chain fatty acids. Taken together, these results have implications for the direction of mass spectrometry as a tool for biosignature detection on Mars, as well as informing the selection of sampling sites that could potentially host biosignatures.

Fast removal of phenolic compounds from water using hierarchical porous carbon nanofibers membrane

Water pollution is becoming increasingly serious and seriously endangering human health, especially the direct emissions of phenolic compounds. An integrated sample pre-treatment and derivatization method based on a biopolymers/TEOS-based carbon nanofibers microextraction that allows rapid extraction (5 min), followed by separation and highly sensitive detection of phenolic compounds by gas chromatography‒mass spectrometry (GC-MS) analysis, is described. The developed methodology, coupled with GC-MS, allowed low detection limits (0.03–0.32 ng mL‒1), good linearities (0.5–200 ng mL‒1) and recoveries (73.58–85.76%) to be achieved in a few steps and short time. Based on the high adsorption properties of materials, the on-line removal device of phenolic compounds in water was designed. The elimination rate of phenolic compounds in water was higher than 73.58% at 1 mL min‒1 and three cycles. The elimination strategy for the phenolic compounds is very versatile and is easy to apply.

3-Acetylpyridine On-Tissue Paternò-Büchi Derivatization Enabling High Coverage Lipid C═C Location-Resolved MS Imaging in Biological Tissues.

Unsaturated lipids containing single or more carbon-carbon double bonds (C═C) within tissues are closely associated with various types of diseases. Mass spectrometry imaging (MSI) has been used to study the spatial distribution of lipid C═C location isomers in tissue sections. However, comprehensive characterization of lipid C═C location isomers using MSI remains challenging. Herein, we established an on-tissue charge-switching Paternò-Büchi (PB) derivatization method using 3-acetylpyridine (3-AP) as a reaction reagent, which can be used to detect and assign C═C location of glycerophospholipids (GPLs) as well as neutral lipids, such as fatty acids (FAs), under the same experimental workflow using matrix-assisted laser desorption/ionization (MALDI)-MSI. High coverage of mono- and poly-unsaturated C═C location isomers among various lipid classes including FA, phosphatidylcholine (PC), and sulfatide (SHexCer) in distinct regions of the mouse brain and kidney was visualized using MALDI-MS/MS imaging. This method has also been applied to map the spatial distribution of lipid C═C location isomers in the Alzheimer's disease (AD) mice model for the first time, which provides a new tool to study the relationships between the distribution of lipid structural diversity and neurodegenerative diseases.

Evaluation of the Perceptual Interaction among Ester Odorants and Nonvolatile Organic Acids in Baijiu by GC-MS, GC-O, Odor Threshold, and Sensory Analysis.

By applying the aroma extract dilution analysis, 13 esters were found to have high FD factors in the Laobaigan flavor type of Baijiu. These were then quantified using GC-MS. In addition, 35 nonvolatile organic acids were quantified by a derivatization method combined with GC-MS. The perceptual interactions of lactic acid and ethyl lactate and that of lactic acid and ethyl acetate were studied through the olfactory threshold. The S curve was used to evaluate the changes in the olfactory thresholds. After adding lactic acid (1142, 20 000, and 53 703 mg/L), the olfactory thresholds of ethyl lactate (1000, 724, and 295 mg/L) and ethyl acetate (398, 324, and 257 mg/L) decreased obviously, which revealed that lactic acid gave additive or synergistic odor effects for the two esters. Furthermore, it was discovered that as the concentrations of lactic acid increased, the extent of the interaction among these compounds was also greater in the mixture.

Discovery of Novel N-Pyridylpyrazole Thiazole Derivatives as Insecticide Leads

To develop effective insecticides against Lepidoptera pests, 25 novel N-pyridylpyrazole derivatives containing thiazole moiety were designed and synthesized based on the intermediate derivatization method (IDM). The insecticidal activities of these target compounds against Plutella xylostella (P. xylostella), Spodoptera exigua (S. exigua), and Spodoptera frugiperda (S. frugiperda) were evaluated. Bioassays indicated that compound 7g−7j exhibited good insecticidal activities. Compound 7g showed especially excellent insecticidal activities against P. xylostella, S. exigua, and S. frugiperda with LC50 values of 5.32 mg/L, 6.75 mg/L, and 7.64 mg/L, respectively, which were adequate for that of commercial insecticide indoxacarb. A preliminary structure-activity relationship analysis showed that the insecticidal activities of thiazole amides were better than that of thiazole esters, and the amides with electron-withdrawing groups on the benzene ring were better than the ones with electron-donating groups. This work provides important information for designing novel N-pyridylpyrazole thiazole candidate compounds and suggests that the 7g is a promising insecticide lead for further studies.

Enzymatic Nitrogen Insertion into Unactivated C-H Bonds.

Selective functionalization of aliphatic C-H bonds, ubiquitous in molecular structures, could allow ready access to diverse chemical products. While enzymatic oxygenation of C-H bonds is well established, the analogous enzymatic nitrogen functionalization is still unknown; nature is reliant on preoxidized compounds for nitrogen incorporation. Likewise, synthetic methods for selective nitrogen derivatization of unbiased C-H bonds remain elusive. In this work, new-to-nature heme-containing nitrene transferases were used as starting points for the directed evolution of enzymes to selectively aminate and amidate unactivated C(sp3)-H sites. The desymmetrization of methyl- and ethylcyclohexane with divergent site selectivity is offered as demonstration. The evolved enzymes in these lineages are highly promiscuous and show activity toward a wide array of substrates, providing a foundation for further evolution of nitrene transferase function. Computational studies and kinetic isotope effects (KIEs) are consistent with a stepwise radical pathway involving an irreversible, enantiodetermining hydrogen atom transfer (HAT), followed by a lower-barrier diastereoselectivity-determining radical rebound step. In-enzyme molecular dynamics (MD) simulations reveal a predominantly hydrophobic pocket with favorable dispersion interactions with the substrate. By offering a direct path from saturated precursors, these enzymes present a new biochemical logic for accessing nitrogen-containing compounds.

Enhanced levofloxacin degradation by hierarchical porous Co3O4 with rich oxygen vacancies activating peroxymonosulfate: Performance and mechanism

Hierarchical porous Co3O4 with rich oxygen vacancies were fabricated by simple biotemplates derivatization method, and largely promoted the efficiency of peroxymonosulfate (PMS) activation to enhance levofloxacin (LVF) degradation by enlarging the specific surface area and introducing rich oxygen vacancy (Ovac). Results shows that hierarchical porous Co3O4 exposed more surface active sites, Co2+ and Ovac as active sites for PMS activation compared with that of bare Co3O4 obtained without biotemplates, giving rise to largely enhancement of catalytic performance, completely degrading 100% of LVF within 12 min by the optimum Co3O4-II. Besides, experimental studies confirmed that SO4− and electron transfer mechanism mainly contributed to LVF degradation in Co3O4-II/PMS system. The possible LVF degradation pathway was established on the base of the detected intermediates. This study not only improves the fundamental understanding of active sites in morphologically tunable metal oxides via biotemplates derivatization, but also provides guidance for the research and engineering application of peroxymonosulfate based advanced oxidation process in the future.

Synthesis, Anticancer Activities, and Mechanism of N-heptyl-containing Biguanide Derivatives.

In recent years, the anticancer effects of biguanide drugs have received considerable attention. However, the effective concentration of biguanide drugs to kill cancer cells is relatively high. Thus, we focus on structural modification of biguanides to obtain better antitumor candidates. A previous study in our laboratory has found that a biguanide compound containing the n-heptyl group has potent anticancer activity. However, the effect of different substituents on the benzene ringside of the biguanides on the anti-proliferative activity is unknown. A series of n-heptyl-containing biguanide derivatives whose benzene rings were modified by halogen substitution based on the intermediate derivatization method were further synthesized to find new compounds with improved antiproliferative activities. Ten n-heptyl-containing biguanide derivatives were synthesized via established chemical procedures. The activities of these derivatives were explored by MTT assay, clonogenic assay, and scratch assay. The protein levels were detected via Western blotting to explore the underlying mechanisms. The optimal biguanide derivatives 10a-10c, 11d exhibited IC50 values of 2.21-9.59 μΜ for five human cancer cell lines, significantly better than the control drug proguanil. The results of clonogenic and scratch wound healing assays also confirmed the inhibitory effects of derivatives 10a- 10c, 11d on the proliferation and migration of human cancer cell lines. Western blot results demonstrated that one representative derivative, 10c upregulates the AMPK signal pathway and downregulates mTOR/4EBP1/p70S6K. All biguanide derivatives containing n-heptyl groups are more active than proguanil, indicating that the modification of n-heptyl-containing biguanide derivatives provides a novel approach for the development of novel high efficient antitumor drugs.

Precolumn derivatization LC/MS method for observation of efficient hydrogen sulfide supply to the kidney via d-cysteine degradation pathway

d-Cysteine (d-Cys) is metabolized to hydrogen sulfide (H2S) by d-amino acid oxidase (DAO)/3-mercaptopyruvate sulfurtransferase pathway. The pathway is required for H2S supplementation that ameliorates acute kidney injury after the oral administration of d-Cys in mice. However, whether the rate-limiting activity of DAO regulates the tissue-selectivity or the extent of d-Cys degradation and H2S supplementation remains unclear. Here, to analyze the levels of d-Cys and H2S, we use two derivatization methods, a new method with no detectable isomerization of Cys and an established method for H2S. The derivatives were determined by LC/MS using a C18 column. With the methods, we show that inhibition of DAO significantly suppresses the H2S supplementation and d-Cys degradation in the mouse kidney. Additionally, we found that d-Cys is more efficiently metabolized into H2S than l-Cys in the kidney. Our results reveal the utility of the method and support the advantage of d-Cys administration in improving the supply of H2S to the kidneys.