High-performance liquid chromatographic measurement of guanidino compounds of clinical importance in human urine and serum by pre-column fluorescence derivatization using benzoin

High-performance liquid chromatographic microanalyses for guanidino compounds in human physiological fluids have been accomplished by means of a pre-column fluorescence derivatization method using benzoin. The guanidino compounds in urine or deproteinized serum after ultrafiltration are converted to the fluorescent derivatives with benzoin in an alkaline medium, and the derivatives are separated simultaneously within 25 min on a reversed-phase column (mu Bondapak Phenyl) with a linear gradient elution of methanol in aqueous mobile phase (pH 8.5). The method permits the quantitative determination of guanidinosuccinic acid, methylguanidine, taurocyamine and guanidinobutyric acid at concentrations of as low as 8-78 pmol/ml in human urine and serum.     

Bestimmung von Guanidin, Methylguanidin und asymmetrischem Dimethylguanidin in menschlichem Serum

Zusammenfassung Durch Umsatz von Guanidin, Methylguanidin oder Dimethylguanidin mit Acetylaceton lassen sich zur Gas-Chromatographie geeignete Pyrimidinderivate gewinnen. Ihr Verhalten bei Dünnschicht-Chromatographie und Flüssig-flüssig-Verteilungen, ihre NMR, IR- und UV-Spektren und ihre Flüchtigkeit werden beschrieben. Die Bedingungen zur Synthese werden untersucht und der Syntheseverlauf, ausgehend von Nanogramm-, Mikrogramm oder Milligrammengen der Guanidine, wird diskutiert. Die besten Bedingungen zur Gas-Chromatographie der Guanidine werden angegeben und die Methoden zur Analyse von Serumproben verwandt. Der Gehalt im pool-Serum Gesunder beträgt etwa 0,2 ppm Guanidin, 0,1 ppm Methylguanidin. Dimethylguanidin konnte nicht gefunden werden. Seine Konzentration muß unter 0,1 ppm liegen.

---

Determination of guanidine, methylguanidine and asymmetric dimethylguanidine in human serumPart II. Gas-chromatographic determination

Derivatives of pyrimidine are prepared by reacting guanidine, methylguanidine or dimethylguanidine with acetylacetone. The derivatives are used for a gas-chromatographic determination. Their NMR-, IR- and UV-spectra, their behaviour in distribution processes and thin-layer chromatographic separations and their volatility is described.Conditions for synthesis, starting with nanogram, microgram or milligram amounts of guanidines are elaborated and compared. The procedures proposed are applied to the determination of the concentration of guanidines in serum. Serum of healthy persons contains about 0.2 ppm of guanidine, 0.1 ppm of methylguanidine. Dimethylguanidine could not be detected. Its concentration must be smaller than 0.1 ppm.

---

---

Teil I: diese Z. 245, 311 (1969).

---

Herrn Dr. A. F. Moroni (Institut für physikalische Chemie, Universität Mainz) danken wir für die bereitwillige Unterstützung bei der Auswertung der kinetischen Ergebnisse und Herrn Dr. F. Caesar für die Diskussion der Kernresonanzspektren.     

Gas chromatographic determination of guanidino compounds in uremic patients using glyoxal as derivatizing reagent

The guanidino compounds guanidine, methylguanidine, guanidinoacetic acid, guanidinopropionic acid, guanidinobutyric acid and guanidinosuccinic acid were eluted and separated after pre-column derivatization with glyoxal from an HP-5 column (30 m × 0.32 mm i.d.) with film thickness 0.25 μm at an initial column temperature of 100 °C for 2 min, with ramping of 20°C/min up to 250 °C and a nitrogen flow rate of 3 mL/min. Detection was by flame ionization detection. Linear calibrations were observed within 0.1-20.0 μmol/L, with limit of detection within 0.024-0.034 μmol/L for each compound. The separation was repeatable with relative standard deviation (RSD) (n = 6) within 1.2-1.8 and 1.1-1.6% in terms of retention time and peak height/peak area, respectively. The method was applied for the determination of the guanidino compounds from serum of uremic patients (n = 7) and healthy volunteers (n = 8), and amounts were observed within 1.33-11.71 and 0.07-0.39 μmol/L with RSD 1.1-3.5 and 1.1-3.0%, respectively. The results were further supported by the standard addition method.     

Synthesis of pyrimidine derivatives by reaction of pyrylium salts with guanidine and compounds of the guanidine series

2,4,6-Triphenylpyrylium perchlorate reacts with methylguanidine to give 1-methyl-2-amino-4,6-diphenylpyrimidinium perchlorate, which undergoes the Dimroth rearrangement to give 2-methylamino-4,6-diphenylpyrimidine. 2,4,6-Triarylpyrylium perchlorates react with guanidine to give N-pyrimidinylpyridinium salts. Pyrylium salts react with aminoguanidine, sulfanilylguanidine, and symmetrical diphenylguanidine at one amino group to give the corresponding pyridinium salts. Translated from Khimiya Geterotsiklicheskikh Soedinenii, No. 6, pp. 748–752, June, 1980.     

Bestimmung von Guanidin, Methylguanidin und asymmetrischem Dimethylguanidin in menschlichem Serum

Zusammenfassung Die Abschätzung der biochemischen Bedeutung von Guanidin, Methylguanidin und Dimethylguanidin macht die Ausarbeitung zuverlässigerer Analysenmethoden nötig. Zwei unabhängige Verfahren, die auf verschiedenen Prinzipien beruhen, werden vorgeschlagen. Im Teil I wird die Enteiweißung mit Trichloressigsäure oder Alkohol-Aceton und die nachfolgende Extraktion der Guanidin-Tetraphenylborate aus Lösungen von pH 8 beschrieben. Nach dieser Voranreicherung wird die Dünnschicht-Chromatographie zur Trennung der einzelnen Guanidine voneinander verwandt. Diese werden von der Celluloseschicht eluiert und durch Fluorimetrie bestimmt, wobei Ninhydrin in alkalischer Lösung als Reagens benutzt wird. Im Teil II wird die gas-chromatographische Bestimmung der Guanidine beschrieben.

---

Determination of guanidine, methylguanidine and asymmetric dimethylguanidine in human serumPart I. Determination by thin-layer chromatography and subsequent fluorimetry

An evaluation of the biochemical significance of guanidine, methylguanidine and dimethylguanidine requires the elaboration of more reliable methods. Two independent procedures based on different principles are suggested. Part I describes the deproteinization, using trichloracetic acid or alcohol-acetone, and the extraction of the tetraphenyloborates of guanidines at pH 8. After such a preconcentration thin-layer chromatography is used for the separation of the individual guanidines. These are eluted from the cellulose carrier and determined by means of a fluorescent reaction with ninhydrin in alcaline solution as reagent.Part II will describe a gas-chromatographic determination of the guanidines.

     

High-performance liquid chromatographic analysis of guanidino compounds using ninhydrin reagent. II. Guanidino compounds in blood of patients on haemodialysis therapy

An automated high-performance liquid chromatographic method using alkali-ninhydrin reagent for the post-column derivatization of guanidines has been developed. This procedure was applied to measurements made before and after haemodialysis. Among the guanidino compounds found in human blood, methylguanidine showed the lowest removal rate. The removal rate of guanidinosuccinic acid correlated with the plasma alpha 1-globulin fraction. The removal rate of each guanidino compound decreased with the period of dialysis.     

Synthesis of Amphiphilic Guanylated Polymers as Potential Gene Delivery Carriers

Abstract

We have developed novel polymeric systems that aggregate in aqueous media to form nanospheric particles. Our polymeric system constitutes guanidine functional groups and poly(ethylene glycol) (PEG) units. The guanidine group is an important structural component in many biologically active compounds. Because of their strongly basic character, guanidines are fully protonated under physiological conditions. The positive charge thus imposed on the molecule forms the basis for specific interactions between the ligand and receptor or the enzyme and substrate. The PEG units may form a characteristic micelle structure with a hydrophilic shell layer surrounding the core of the polyion complex (PIC) formed between DNA and the cation segments (PIC micelles).     

A rapid,direct microquantitative method for determination of cyclohexanone in physiological solutions and biological fluids and of the retention behavior of some volatile compounds by gas chromatography

A gas Chromatographic method for a rapid, direct microquantitative determination of cyclohexanone in physiological solutions and biological fluids is described. The retention behavior of some volatile organic compounds is also presented. The effect of column temperature on the retention behavior of cyclohexanone is briefly discussed. Other Chromatographic factors which might affect the analytical results are mentioned.     

Amino acid analysis of physiological fluids by high-performance liquid chromatography with phenylisothiocyanate derivatization and comparison with ion-exchange chromatography

The suitability of pre-column derivatization with phenylisothiocyanate followed by high-performance liquid chromatography was investigated as a means of analyzing free amino acids in plasma and other physiological fluids. A comparison was made between this method and a conventional ion-exchange method. The correlation coefficient for all the amino acids tested was greater than 0.9, except for proline and tryptophan. Various forms of sample preparation were tried for plasma and amniotic fluid; it was finally decided that protein precipitation with acetonitrile was most suitable. Ultrafiltration was used for cerebrospinal fluid preparation while urine was treated the same as a standard mixture. The retention times relative to the internal standard (nor-leucine) are given for over 90 compounds. Some of these were chromatographed underivatized because they are known to be present in some physiological fluids and absorb at 254 nm because of their aromaticity. The imprecision for this method compared favourably with the standard ion-exchange method although each had specific amino acids for which the imprecision was poor. The technique is suitable for the same routine clinical analysis purposes as high-resolution ion-exchange chromatography. It also offers the advantages of speed of analysis, sensitivity and equipment versatility over the conventional ion-exchange methods.     

The chemistry and biology of organic guanidine derivatives

Organic guanidine compounds are reviewed, with emphasis on natural products isolation, identification, synthesis and biological activities. The literature survey includes purely synthetic guanidine derivatives, guanidine alkaloids and non-ribosomal peptides from bacteria and cyanobacteria, as well as related compounds isolated from marine and terrestrial invertebrates and higher plants.     

Determination of Guanidino Compounds in Serum by Micellar Electrokinetic Capillary Chromatography Using Benzoin as the Derivatizing Reagent

An analytical procedure has been evolved for the determination of seven guanidino compounds; guanidine (G), methylguanidine (MG), guanidinoacetic acid (GAA), guanidinopropionic acid (GPA), guanidinobutyric acid (GBA), arginine (Arg), and guanidinosuccinic acid (GSA) by micellar electrokinetic capillary chromatography (MEKC) within 6 min using benzoin as derivatizing reagent. Sodium dodecyl sulfate (SDS) was used as the micellar medium in a sodium tetraborate (0.1 M) buffer at pH 8.5. Uncoated fused silica capillary was used with an effective length of 39 cm and 75 μm id. Applied voltage was 25 kV and photo diode array detection was set at 228 nm. Linear calibrations obtained from 0.057 to 14.11 μmol/L and limits of detection (LOD) were within 0.019–0.03 μmol/L. The derivatization and separation was repeatable with relative standard deviation (RSD) within 1.9–3.8%. Serum of healthy volunteers and uremic patients was analyzed and amounts found in uremic patients were G 1.98–3.03, MG 1.21–1.79, GAA 3.67–6.09, GPA 1.17–1.37, GBA 1.29–1.46, Arg 9.49–19.17, and GSA 6.83–10.91 μmol/L with RSD (n = 4) within 1.3–4.5%. The amount of guanidino compounds was higher in uremic patients than in healthy volunteers.     

Optimization of derivatization parameters with 9-fluorenylmethyl chloroformate for amphetamine and catecholamine separation by supercritical fluid chromatography

Summary The separation of amino compounds by supercritical fluid chromatography (SFC) is a difficult problem to solve, owing to the apolar nature of CO₂. The derivatization of amino functions with the 9-fluorenylmethyl chloroformate (FMOC-Cl) allows to obtain apolar UV-absorbing compounds easily eluted with a supercritical mobile phase. Optimization of derivatization parameters allows us to analyze quantitatively amphetamines and catecholamines. These compounds can be separated in less than 5 min with a small addition of methanol as polar modifier. The total procedure takes no more than 15 min and can be automatized to gain time. As presented in this study, this method can be employed to physiological fluids as urine.     

Gas Chromatographic Determination of Guanidino Compounds Using Hexafluoroacetylacetone and Ethyl Chloroformate as Derivatizing Reagents

Guanidino compounds guanidine, methylguanidine, guanidinoacetic acid, guanidinobutyric acid, guanidinopropionic acid, and guanidinosuccinic acid after derivatization with hexafluoroacetylacetone and ethyl chloroformate at pH 9 in aqueous phase, eluted, and separated from gas chromatographic column HP-5 (30 m × 0.32 mm id) with film thickness of 0.25 μm at an initial column temperature 90 °C for 2 min, followed by heating rate of 10 °C min^?1 up to 220 °C with nitrogen flow rate of 1 mL min^?1. The detection was by flame ionization detector. The linear calibration ranges of each of guanidino compounds were obtained within 1–10 μg mL^?1, and the limit of detection was within 0.014–0.19 μg mL^?1. The derivatization and gas chromatography elution and separation were repeatable in terms of retention time and peak height/peak area with relative standard deviation (RSD) (n = 4) within 1.7–2.9 % and 1.4–2.8 %, respectively. The method was applied for the determination of guanidino compounds from deproteinized serum of uremic patients and healthy volunteers, and was found in the range below the limit of quantitation (BLOQ) to 1.25 μg mL^?1 with RSD within 1.4–3.6 %, and BLOQ to 0.4 μg mL^?1 with RSD 1.3–3.4 %, respectively. A number of pharmaceutical additives did not effect the determination with RSD within ±3.1 %.     

Development of a Promising Method for Producing Oligomeric Mixture of Branched Alkylene Guanidines to Improve Substance Quality and Evaluate Their Antiviral Activity against SARS-CoV-2

This paper reports the synthesis of branched alkylene guanidines using microfluidic technologies. We describe the preparation of guanidine derivatives at lower temperatures, and with significantly less time than that required in the previously applicable method. Furthermore, the use of microfluidics allows the attainment of high-purity products with a low residual monomer content, which can expand the range of applications of this class of compounds. For all the samples obtained, the molecular-weight characteristics are calculated, based on which the optimal condensation conditions are established. Additionally, in this work, the antiviral activity of the alkylene guanidine salt against the SARS-CoV-2 virus is confirmed.     

Three New Compounds Based on 4,4'-Azo-1,2,4-triazol-5-one: Synthesis,Crystal Structure and Thermal Properties

A series of guanidine salts of 4,4'-azo-1,2,4-triazol-5-one with guanidine(1), aminoguanidine(2), diaminoguanidine(3) and triaminoguanidine(4) was prepared. Compounds 2-4 were characterized by infrared(IR) spectroscopy, elemental analysis and single-crystal X-ray diffraction. Thermal decomposition processes of compounds 1-4 were investigated by differential scanning calorimetry(DSC), and all the compounds showed good thermal stability up to 190℃. Moreover, these four guanidine salts are more unstable with the increasing number of amino groups. Thermal stability parameters(T_e,0 and T_b) and thermodynamic functions(ΔS^≠, ΔH^≠ and ΔG^≠) for compounds 1-4 were calculated. The constant-volume combustion heats(Δ_cU) for compounds 2-4 were determined and tended to increase with the increase of the number of amino groups. The calculated standard molar enthalpies of formation(Δ_fH⁰m) of compounds 2-4 are -541.04, -178.67 and -83.08 kJ/mol, respectively. The impact sensitivities results indicate that these four energetic salts are less sensitive than 1,3,5-trinitrotriazacyccohexane(RDX) and 1,3,5,7-tetranitrotetraqza-cyclo-octane(HMX).     

TFA-sensitive arylsulfonylthiourea-assisted synthesis of N,N'-substituted guanidines

An efficient synthesis of N,N'-substituted guanidine derivatives was developed via an aromatic sulfonyl-activated thiourea intermediate. The use of certain aromatic sulfonamides, such as PbfNH(2), as the key reagent to incorporate a TFA-labile guanidine protection group greatly facilitates solid-phase synthesis of N,N'-substituted guanidine compounds.     

Cyclic Guanidine Compounds from Toxic Newts Support the Hypothesis that Tetrodotoxin is Derived from a Monoterpene

The biosynthesis of tetrodotoxin (TTX), a potent neurotoxin consisting of a 2,4‐dioxaadamantane skeleton and a guanidine moiety, is an unsolved problem in natural product chemistry. Recently, the first C5–C10 directly bonded TTX analogue, 4,9‐anhydro‐10‐hemiketal‐5‐deoxyTTX, was obtained from toxic newts and its carbon skeleton suggested a possible monoterpene origin. On the basis of this hypothesis, screening of predicted biosynthetic intermediates of TTX was performed using two MS‐guided methods. Herein, five novel cyclic guanidine compounds from toxic newts are reported which commonly contain a cis‐fused bicyclic structure including a six‐membered cyclic guanidine. These structures could be biosynthetically derived from geranyl guanidine through oxidation, cyclization, and/or isomerization steps. LC–MS analysis confirmed the widespread distribution of the five novel compounds in toxic newt species. These results support the hypothesis that TTX is derived from a monoterpene.     

Thermal degradation of wood treated with guanidine compounds in air: Flammability study

Wood has been treated with guanidine phosphate, guanidine nitrate, guanidine carbonate and guanidine chloride to impart flame retardancy. The samples were subjected to differential thermal analysis (DTA) and thermogravimetry (TG) from ambient temperature to 800°C in air to study their thermal behaviors. From the resulting data, kinetic parameters for different stages of thermal degradation were obtained following the method of Broido. For the decomposition of wood and flame retardant wood, the activation energy was found to decrease from 116 to 54 kJ mol^–1; the char yield was found to increase from 5.6 to 34.9%, LOI from 18 to 41.5, which indicated that the flame retardancy of treated wood was improved. Effects of the different compounds on the degradation and flammability of wood have also been proposed.This revised version was published online in November 2005 with corrections to the Cover Date.     

Synthesis and Crystal Structure of a Novel 4-Guanidinosalicylic Acid

The guanidine functionality is found widely in natural products, pharmaceutically active compounds and in molecules used for molecular recognition.[1]A number of compounds containing guanidine groups possess the potent antitumour activity and antifungal activity.[2]     

Pyrimidines

A method for the preparation of 2-amino(acetamido)pyrimidines by condensation of α,β-unsaturated carbonyl compounds with guanidine (acetylguanidine) was developed.