Analytical investigation of salivary calculi, by mid-infrared spectroscopy

Sialolithiasis is common in salivary glands, especially in the submandibular and parotid ducts. X-Ray diffractometry was the principal technique used for their analysis, sometimes associated with scanning electron microscopy. Hydroxyapatite was the most frequently described constituent, in association with whitlockite and other calcium phosphates as brushite or octocalcium phosphate. Proteic matter was detected, as mucoproteins, albumin, nucleoproteins or as degenerative bacterial matter. This study presents the identification of constituents by mid-infrared spectrometry of 74 sialoliths. Their successive layers are analyzed from their crust to the nucleus, using absorbance measurements. Spectra are compared with reference mixtures of two or more constituents. Approximately 99% of sialoliths are constituted of calcium phosphates, under carbonated forms. More than three-quarters contain proteins, in which mucins represent the majority and albumin is found in 10% of all the specimens. Only 7% calculi are an association of two constituents, 66% are made of three and 27% have four or more components. For the 74 studied sialoliths, no specimen contains hydroxyapatite; but they are composed of carbonate apatites with irregular microcrystallized forms, even if proteins are present. Some of them have a pure protein nucleus, surrounded by carbonate apatite layers; the other stones are made of internal layers of apatites and covered with a dense and varnished crust of proteins.     

Simultaneous determination of composition of human urinary calculi by use of artificial neural networks

A new chemometric method, which uses artificial neural networks (ANN), is presented for determination of the composition of urinary calculi. The selected constituents were whewellite, weddellite, and uric acid from which approximately 40% of the urinary calculi obtained from Macedonia patients are composed. The results for the synthetic mixtures were better then those obtained by partial least squares (PLS) regression or by the principal component regression (PCR), because neural networks have better prediction capacity. The generalization abilities of the optimized neural networks were checked using the standard addition method on carefully selected real natural samples.     

用傅立叶红外光谱测定泌尿系结石中的草酸钙

对泌尿系结石的组成进行结构分析,为分析结石病的产生原因及预防复发提供参考,在临床诊断和治疗上具有重要意义.傅立叶红外光谱是研究泌尿系结石构造的一种较理想的分析手段.用红外光谱仪定量分析泌尿结石中草酸钙采用的内标法、吸光度比值法和一阶导数法进行了比较,建立的分析方法可以简便对草酸钙混合结石中一水草酸钙(COM)和二水草酸钙(COD)进行定量测定,分析结果可初步满足临床诊断的要求.     

Ultrasonic nebulization extraction-heating gas flow transfer-headspace single drop microextraction of essential oil from pericarp of Zanthoxylum bungeanum Maxim

The ultrasonic nebulization extraction-heating gas flow transfer coupled with headspace single drop microextraction (UNE-HGFT-HS-SDME) was developed for the extraction of essential oil from Zanthoxylum bungeanum Maxim. The gas chromatography-mass spectrometry was applied to the determination of the constituents in the essential oil. The contents of the constituents from essential oil obtained by the proposed method were found to be more similar to those obtained by hydro-distillation (HD) than those obtained by ultrasonic nebulization extraction coupled with headspace single drop microextraction (UNE-HS-SDME). The heating gas flow was firstly used in the analysis of the essential oil to transfer the analytes from the headspace to the solvent microdrop. The relative standard deviations for determining the five major constituents were in the range from 1.5 to 6.7%. The proposed method is a fast, sensitive, low cost and small sample consumption method for the determination of the volatile and semivolatile constituents in the plant materials.     

Determination of the composition of human urinary calculi composed of whewellite, weddellite and carbonate apatite using artificial neural networks

More than half of the analyzed calculi from patients from Macedonia are composed of whewellite, weddellite and carbonate apatite (as single components or in binary or ternary mixtures). In order to develop a simple and satisfactorily reliable method for quantitative analysis of urinary calculi, the possibility was explored to employ artificial neural networks (ANNs) as a tool for such a purpose. By changing the number of input and hidden neurons, a search was made for the three-layered feed-forward ANN which would give the best performance. The root-mean-square errors (RMSE) for the test samples are: 0.035 for whewellite, 0.064 for weddellite and 0.078 for carbonate apatite. The accuracy of the method was checked using standard-addition method on real samples. The discrepancies between calculated and predicted mass fraction of constituents were in the range acceptable for use of the proposed method in clinical laboratories.     

荧光分光光度法直接测定炎痢停片中的甲氧苄氨嘧啶和氟哌酸

应用荧光分光光度法通过更换溶剂,不经分离同时测定了炎痢停片中甲氧苄氨嘧啶（ＴＭＰ）和氟哌酸的含量。水相中直接测定了氟哌酸含量,乙醇相中测定ＴＭＰ时,引入光度分析中的Ｈ－点标准加入法求得ＴＭＰ的含量。该方法平均回收率：ＴＭＰ为１００．４４％（ＲＳＤ＝１．７６％,ｎ＝５）。氟哌酸为９９．９６％（ＲＳＤ＝１．６２％,ｎ＝５）。此法可作为样品的检测方法,也可应用于生产质量控制和药物代谢研究。     

On the structural diversity of sialoliths.

Sialoliths from parotid and submaxillar glands have been characterized. Fractured and polished surfaces revealed an intrinsic structural diversity across the calculi sections. In general, the calculi presented highly mineralized amorphous-looking cores surrounded by concentric alternating mineralized and organic layers. The thickness of these layers decreased from the outer regions toward the center of the sialolith, illustrating a sequence of growth stages. Nevertheless, a significant variability could be detected among the specimens. In some cases, the calculi displayed multiple cores and lacked concentric laminated structures. In other instances, the specimens exhibited extensive regions of globular structures. In these cases, the globule diameter decreased across the radius toward the center of the sialoliths, and the globular structures tended to reorganize, forming bright and dark laminated layers surrounding the core. The participation of globular structures in the layer formation process points to morphogenetic mechanisms not previously described.     

GC／MS法测定木香挥发油化学成分

用气相色谱-质谱法对木香挥发油进行化学成分的分析。采用水蒸气馏法从木香中提取挥发油。试用不同类型的毛细管柱进行分析,找出最佳分析条件,用归一化法测定其含量,并用气相色谱-质谱法对化学成分进行鉴定。共鉴定了38个成分,占挥发油总成分的86%以上。方法稳定可靠。重现性好,适用于中药挥发油的化学成分分析。     

Trace element study of kidney stones from subjects belonging to stone belt region of India

Kidney stones obtained from six patients belonging to the stone belt region of India (Punjab) were analyzed for inorganic constituents using instrumental neutron activation analysis (INAA) and energy dispersive X-ray fluorescence (EDXRF) techniques. For INAA, samples were irradiated along with IAEA RM Soil 7 as reference standard in CIRUS reactor, BARC, Mumbai. Gamma activity of irradiated samples was measured using a 45% relative efficiency HPGe detector coupled to 8?k channel analyzer. EDXRF method was used for determination of concentration of Ca. The concentrations of ten elements namely Ca, Na, K, Mn, Co, Cr, Zn, Br, Sm and Cl, are reported and discussed.     

Evaluation of the renal calculi compositions

Thermal analysis and infrared (IR) spectroscopy are modern physical–chemical methods suitable for the investigation of the kidney stones composition. The applications of these methods in our work were anticipated by performing the standard thermal analysis and standard infrared spectra on pure compounds: oxalates, phosphate, carbonate, and uric acid. This work reveals a logical algorithm for correlating the experimental data regarding urolithiasis types and compositions by thermal analysis methods, Fourier Transform Infrared (FT-IR) spectroscopy, and second derivative FT-IR spectra. The limits and performance of each analysis method have been highlighted, and by correlating the results of both methods we have obtained comprehensive information for the identification and/or determination of the main components as well as of organic and/or inert impurities in calculi composition. The data regarding the urinary calculi composition are important for the clinical guideline in the prophilaxy and methaphylaxy of urolithiasis.     

Polarographic determination of hydroxylamines: application to analysis of photographic processing solutions

Procedures for the determination of hydroxylamine and N,N-diethylhydroxyIamine (DEHA), based on anodic polarographic waves, are described. The importance of using a strongly alkaline supporting electrolyte and of complete removal of dissolved oxygen is illustrated. With rapid alternating current (a.c.) polarography, 3 × 10^-6 M hydroxylamine and 4 × 10^-5 M DEHA can be detected. Detection limits with the differential pulse technique are approximately tenfold lower. In a practical application, rapid a.c. polarography is shown to be suitable for the direct determination of hydroxylamine and DEHA in photographic processing solutions. The only pretreatment of samples is dilution with a strongly alkaline supporting electrolyte. Possible interferences from other constituents of the processing solutions are avoided by using the standard addition method.     

Trial Proteomic Qualitative and Quantitative Analysis of the Protein Matrix of Submandibular Sialoliths

Our studies aimed to explore the protein components of the matrix of human submandibular gland sialoliths. A qualitative analysis was carried out based on the filter aided sample preparation (FASP) methodology. In the protein extraction process, we evaluated the applicability of the standard demineralization step and the use of a lysis buffer containing sodium dodecyl sulfate (SDS) and dithiothreitol (DTT). The analysis of fragmentation spectra based on the human database allowed for the identification of 254 human proteins present in the deposits. In addition, the use of multi-round search in the PEAKS Studio program against the bacterial base allowed for the identification of 393 proteins of bacterial origin present in the extract obtained from sialolith, which so far has not been carried out for this biological material. Furthermore, we successfully applied the SWATH methodology, allowing for a relative quantitative analysis of human proteins present in deposits. The obtained results correlate with the classification of sialoliths proposed by Tretiakow. The performed functional analysis allowed for the first time the selection of proteins, the levels of which differ between the tested samples, which may suggest the role of these proteins in the calcification process in different types of sialoliths. These are preliminary studies, and drawing specific conclusions requires research on a larger group, but it provides us the basis for the continuation of the work that has already begun.     

Determination of Almitrine in Human Plasma by a Gas Chronatographic Method

Abstract

A sensitive and specific method for the quantitative determination of the new drug, almitrine bismesylate, was developed based on the procedure reported by Baune et al. The method depends on initial extraction of drug from plasma with cyclohexane followed by separation from acidic constituents by a base wash. Final separation and quantitation of almitrine was achieved by GLC with NPD detection using the dtmethyl derivative (S-2082) as internal standard. The method was shown to be specific for almitrfne and for internal standard by a parallel run using GC/MS and anaiysis of the M.S. species. The procedure was linear from 25 to 175 ng/ml. Assays were routinely performed with 1 ml of plasma though the analysis could be performed with 0.5 ml. The method is rugged, having been run by at least three analysts over a span of more than 5000 samples, and efficient, allowing 60 samples per day throughput on a routine basis.     

Determination of deoxyspergualin by gas chromatography-selected-ion monitoring

A gas chromatographic-selected-ion monitoring method was developed for the sensitive and specific determination of deoxyspergualin in dog plasma using a capillary column and a C8-amide homologue of deoxyspergualin as an internal standard. Extraction and purification of deoxyspergualin from its constituents, 7-guanidinoheptanamide and glyoxylylspermidine, and its possible metabolites was achieved by using CM-Sephadex C-25 column chromatography with stepwise elution with sodium chloride solution. Treatment of deoxyspergualin with acetylacetone resulted in the formation of the volatile pyrimidine derivative of 7-guanidinoheptanamide accompanying hydrolytic cleavage in the alpha-hydroxyglycine moiety of deoxyspergualin. Deoxyspergualin could be determined in a concentration of 10 ng/ml in plasma. The method was applied to the determination of deoxyspergualin in plasma during and after intravenous infusion into dogs.     

Determination of the nicotine metabolite trans-3'-hydroxycotinine in urine of smokers using gas chromatography with nitrogen-selective detection or selected ion monitoring.

A gas chromatographic method for the determination of the nicotine metabolite trans-3'-hydroxycotinine is described. The method involves conversion of the metabolite to the tert.-butyldimethylsilyl derivative, chromatography on a fused-silica capillary column, and determination using nitrogen-phosphorus detection or electron ionization mass spectrometry with selected ion monitoring. A structural analogue, trans-3-hydroxy-1-methyl-5-(2-pyridyl)pyrrolidin-2-one (trans-3'-hydroxy-ortho-cotinine), was used as an internal standard. Using selected ion monitoring, good precision and accuracy were obtained for determination of trans-3'-hydroxycotinine in urine over the concentration range 10-10,000 ng/ml. There was a good correlation between concentrations determined by selected ion monitoring and by nitrogen-phosphorus detection in urine of smokers, although low concentrations determined using nitrogen-phosphorus detection tended to be somewhat higher, suggesting some interference from urinary constituents. Concentrations and 24-h excretion of trans-3'-hydroxycotinine in the urine of 22 cigarette smokers are reported and compared to concentrations and excretion of nicotine, cotinine, nicotine 1'-N-oxide, nornicotine, and cotinine N-oxide.     

Quantitative determination of bitter principles in specimens of Ganoderma lucidum using high-performance liquid chromatography and its application to the evaluation of ganoderma products

For quantitative determination of 19 triterpene constituents, including six ganoderma alcohols (1-6) and 13 ganoderma acids (7-19), in the products of Ganoderma lucidum, an analytical system was developed using high-performance liquid chromatography with an ODS column. The mobile phase was a linear gradient of 1% AcOH/H(2)O-CH(3)CN and 2% AcOH/H(2)O-CH(3)CN, and the elution profile was monitored at 243 and 250 nm for ganoderma alcohols and acids, respectively. The relative standard deviations of this method were less than 2.35% and 2.18% (n=5) for intraday and interday assays, and the recoveries were 90.9-100.8% and 93.4-103.9% for constituents of alcohol and acid groups, respectively. This system was applied to a quantitative determination of the constituents in 10 different products of G. lucidum: six usual umbrella forms of the fruiting bodies, three antlered forms of the fruiting bodies and spores, and eight specimens from the same G. lucidum strain, which was parasitized on logs from different plants or different fungus beds. The analytical results indicated that the quantity and composition of these triterpenes differed appreciably among various specimens, but the relative ratio of the alcohols and acids was not significantly different when the same strain of G. lucidum was used.     

Ultrafast gas chromatography method with direct injection for the quantitative determination of benzene,toluene, ethylbenzene,and xylenes in commercial gasoline

Benzene, toluene, ethylbenzene, and xylenes are some of the most hazardous constituents found in commercial gasoline samples; therefore, these components must be monitored to avoid toxicological problems. We propose a new routine method of ultrafast gas chromatography coupled to flame ionization detection for the direct determination of benzene, toluene, ethylbenzene, and xylenes in commercial gasoline. This method is based on external standard calibration to quantify each compound, including the validation step of the study of linearity, detection and quantification limits, precision, and accuracy. The time of analysis was less than 3.2 min, with quantitative statements regarding the separation and quantification of all compounds in commercial gasoline samples. Ultrafast gas chromatography is a promising alternative method to official analytical techniques. Government laboratories could consider using this method for quality control.     

Determination of 6-N-trimethyllysine in urine by high-performance liquid chromatography

A method for determination of 6-N-trimethyllysine in urine is described. Trimethyllysine and the chemically analogous 6-N-triethyllysine internal standard were isolated from aqueous samples by microcolumn ion-exclusion chromatography. The specimens were derivatized by reaction with 1-fluoro-2,4-dinitrobenzene and reaction byproducts extracted by organic solvents. The trimethyllysine and internal standard derivatives were separated easily from other sample constituents by reversed-phase paired-ion high-performance liquid chromatography with spectrophotometric detection at 405 nm. Standard curves were linear over a sample concentration range of 10-150 nmol/ml; the detection limit corresponded with 0.1 nmol trimethyllysine injected into the chromatograph. The procedure was used for determination of trimethyllysine in human urine.     

Separation of very hydrophobic analytes by micellar electrokinetic chromatography. I. Optimization of the composition of the sample solution for the determination of the aromatic ingredients of sassafras and other essential oils of forensic interest

A micellar electrokinetic chromatographic method using UV and (UV)LIF detection in-line was developed for the determination of aromatic constituents, mainly allylbenzenes in essential oils. The method optimization included the optimization of the composition of the separation electrolyte using ACN and urea to reduce retention factors and CaCl(2) to widen the migration time window. In addition, it was necessary to optimize the composition of the sample solution which included the addition of a neutral surfactant at high concentration. With the optimized method, the determination of minor constituents in essential oils was possible despite of the presence of a structurally related compound being in a molar ratio excess of 1000:1. The use of UV and LIF-detection in-line enabled the direct comparison of the two detection traces using an electrophoretic mobility x-axis instead of the normal time-based scale. This simplifies the assignment of signals and enhances repeatability. The method developed was successfully applied to the determination of minor and major constituents in herbal essential oils, some of them being forensically relevant as sources of precursors for synthetic drugs.