Measurement of very low-molecular weight metabolites by traveling wave ion mobility and its use in human urine samples

The collision cross-sections(CCS)measurement using ion mobility spectrometry(IMS)in combination with mass spectrometry(MS)offers a great opportunity to increase confidence in metabolite identification.However,owing to the lack of sensitivity and resolution,IMS has an analytical challenge in studying the CCS values of very low-molecular-weight metabolites(VLMs250 Da).Here,we describe an analytical method using ultrahigh-performance liquid chromatography(UPLC)coupled to a traveling wave ion mobility-quadrupole-time-of-flight mass spectrometer optimized for the measurement of VLMs in human urine samples.The experimental CCS values,along with mass spectral properties,were reported for the 174 metabolites.The experimental data included the mass-to-charge ratio(m/z),retention time(RT),tandem MS(MS/MS)spectra,and CCS values.Among the studied metabolites,263 traveling wave ion mobility spectrometry(TWIMS)-derived CCS values(TWCCSN2)were reported for the first time,and more than 70%of these were CCS values of VLMs.The TWCCSN2 values were highly repeatable,with inter-day variations of<1%relative standard deviation(RSD).The developed method revealed excellent TWCCSN2 accuracy with a CCS difference(DCCS)within±2%of the reported drift tube IMS(DTIMS)and TWIMS CCS values.The complexity of the urine matrix did not affect the precision of the method,as evidenced by DCCS within±1.92%.According to the Metabolomics Standards Initiative,55 urinary metabolites were identified with a confidence level of 1.Among these 55 metabolites,53(96%)were VLMs.The larger number of confirmed compounds found in this study was a result of the addition of TWCCSN2 values,which clearly increased metabolite identification confidence.

Urea hydrolysis and recovery of nitrogen and phosphorous as MAP from stale human urine

Laboratory-scale tests for magnesium ammonium phosphate（MAP）precipitation following urea hydrolysis of human urine were conducted using orthogonal experiment design.The effects of initial pH,temperature and the volumetric ratios of stale urine to fresh urine,on urea hydrolysis in urine were studied to determine the final hydrolysis time to recover most nitrogen from separated human urine by MAP.With a volumetric ratio of stale to fresh urine>10% and at temperature≥20℃,urea hydrolysis could be completed in two days.Alkaline pH inhibited urea hydrolysis progress.The final pH values were all around 9.0 following urine hydrolysis,while the suspension pH might act as an indicator to detect the start and extent of urea hydrolysis.Over 95% of ammonium nitrogen and over 85% of phosphorus from hydrolyzed urine as MAP precipitate were obtained using MgCl;·6H;O and Na;HPO;·12H;O as precipitation agents at pH 8.5,molar ratio of Mg;:NH;-N:PO;-P at（1.2-1.3）:1:1,mixing speed of 120 r/min,and precipitation time and reaction time of 3 h and 15 min,respectively.The precipitate has a structure resembling pure MAP crystal.

Enantioselective assay of S(+)- and R(-)-propafenone in human urine by using RP-HPLC with pre-column chiral derivatization

The enantioselective assay for S(+)- and R(-)-propafenone (PPF) in human urine that developed in this work involves extraction of propafenone from human urine and using S(+)-propafenone as internal standard, chiral derivatization with 2,3,4,6-tetra-O-b-D-glucopranosyl isothiocyanate, and quantitation by an RP-HPLC system with UV detection (l=220 nm). A baseline separation of propafenone enantiomers was achieved on a 5-mm reverse phase ODS column, with a mixture of methanol:water:glacial acetic acid (25:12:0.02,v/v) as mobile phase. There was good linear relationship from 24.9 ng/ml to 1875.0 ng/ml for both of enantiomers. The regression equations of the standard curves based on CS-PPF (or CR-PPF ) versus ratio of AS-PPF/AS (or AR-PPF/AS ) were y=0.0032x-0.081, (r=0.999) for S-PPF and y=0.0033x+0.0039, (r=0.998) for R-PPF, respectively. The method抯 limit of detection was 12.5 ng/ml for both enantiomers, and the method抯 limit of quantitation was 28.20.52 ng/ml for S-PPF, 30.40.53 ng/ml for R-PPF (RSD<8%, n=5). The analytical method yielded average recovery of 98.9% and 100.4% for S-PPF and R-PPF, respectively. The relative standard deviation was no more than 6.11% and 6.22% for S-PPF and R-PPF, respectively. The method enabled study of metabolism of S(+)- and R(-)-propafenone in human urine. The results from 7 volunteers administered 150 mg racemic propafenone indicated that propafenone enantiomers undergo stereoselective metabolism and that in the human body, S(+)-propafenone is metabolized more extensively than R(-)- propafenone.

Science Letters:Culture of Spirulina platensis in human urine for biomass production and O_2 evolution

Attempts were made to culture Spirulina platensis in human urine directly to achieve biomass production and O2 evolution, for potential application to nutrient regeneration and air revitalization in life support system. The culture results showed that Spirulinaplatensis grows successfully in diluted human urine, and yields maximal biomass at urine dilution ratios of 140-240. Accumulation of lipid and decreasing of protein occurred due to N deficiency. O2 release rate of Spirulina platensis in diluted human urine was higher than that in Zarrouk medium.

Effects of N source and nitrification pretreatment on growth of Arthrospira platensis in human urine

Culture ofArthrospiraplatensis （Spirulinaplatens） in human urine was investigated to get valuable biomass. NO3-N was the proper N source, in comparison with other N source, including urea, NH4-N and NO2-N. As a result, aerobic nitrification of human urine was performed, with above 93.6% nitrification percentage finally achieved with total-N （TN） load of 46.52 mg/（L.d）, in which Arthrospira platensis was successfully grown. The main compositions of the obtained biomass are close to those in Zarrouk medium. Thus, it is possible to culture Arthrospiraplatensis in nitrified human urine for food production within bioregenerative life support systems （BLSSs）.

Determination of Sparfloxacin in Human Urine by Reversed-Phase High Performance Liquid Chromatography With Nitrous Acid and Hydroiodic Pre-Column Derivatization

Sparfloxacin can be oxidized by nitrous acid, then react with hydroiodic acid to form a fluorescent derivative. Based on this, a reversed-phase high performance liquid chromatographic pre-column derivatization new method is described for the determination of sparfloxacin in human urine. The linear range is 0.05 mg/L to 4.0 mg/L, the recoveries are 91.5%similar to 95.7% and the RSD is 1.2%similar to4.2%. The results showed that this method is suitable for the determination of sparfloxacin in human urine.

GAS CHROMATOGRAPHIC-MASS SPECTROMETRIC ANALYSIS OF ANABOLIC STEROIDS IN HUMAN URINE

A new GC/MS method for detection and identification of 19 anabolic steroids in human urine is presented.The procedure involves adsorption and isolation on a macroporous XAD-2 resin,enzymatic hydrolysis,alkaline extraction,derivatization,GC separation and MS detec- tion.Gas chromatographic-mass spectrometric data illustrate artifacts arising from enzymatic hydrolysis of steroid glucuronides and the structural characterization of their metabolites. Using this method,metabolic studies of these steroids in human urine were made after their ingestion by normal and healthy male volunteers.This method was proven to be suitable for large-scale routine analysis of anabolic steroids and was used successfully in passing the doping control test held by the Medical Commission of the International Olympic Committee.

GC/MS CONFIRMATORY METHOD OF BENZTHIAZIDE IN HUMAN URINE

A GC-MS method for the confirmation of benzthiazide is reported for the first time. This method is based on, the main decomposed product of the methylated derivative and provides a reliable detection of this drug. The detection limit of the method is 1.0 ng with selected ion monitoring.

Nutrient Recycling Using Human Urine： Potential for Low Input Farming

Recycling human urine for farming was assessed in a peri-urban Kyanja parish, Kampala district, and in a rural Migyera parish, Nakasongola district, to demonstrate its potential and develop local use guidelines. Test crops were maize, Nakati （Solanum aethiopicum）, kale, spinach, cabbage, tomatoes, egg plants. Urine-water mixtures （0, 10%, 20%, 30% urine） were applied weekly or bi-weekly. At Kyanja, 30% urine weekly gave the highest maize yields. Within 2 months, 10% urine weekly increased Nakati yield from 5,444 to 24,667 kg ha^-1. 20% Urine weekly increased kale yield （7,556 to 16,111 kg ha^-1） and spinach （4,222 to 19,022 kg ha^-1）. At Migyera, 10% urine weekly increased cabbage yield （4,975 to 16,113 kg ha^-1） but 30% urine weekly decreased cabbage head-weight by 36%. Weekly applied urine produced heavier cabbage heads than bi-weekly （548 g vs. 427 g, P 〈 0.05）. LeafN was higher for weekly than bi-weekly applied urine （3.3% vs. 3.0%）, implying more protein in the former than the latter. From this study, the following guidelines are proposed： Kyanja area, maize： apply 30% urine weekly for 8-weeks; Nakati： apply 10% urine weekly for 8-weeks; Kale and spinach： apply 20% urine weekly; For Migyera area, cabbage and spinach： apply 10% urine weekly. Apply urine 15 cm around each plant starting 2-weeks after transplanting. So kale and spinach prolong urine application for continued harvesting.

Factors hindering the degradation of pharmaceuticals from human urine in an iron-activated persulfate system

This study investigated the degradation of clofibric acid(CFA),bezafibrate(BZF),and sulfamethoxazole(SMX)in synthetic human urine using a novel mesoporous iron powderactivated persulfate system(mFe-PS system),and identified the factors limiting their degradation in synthetic human urine.A kinetic model was established to expose the radical production in various reaction conditions,and experiments were conducted to verify the modeling results.In the phosphate-containing mFe-PS system,the 120 min removal efficiency of CFA decreased from 95.1%to 76.6%as the phosphate concentration increased from 0.32 to 6.45 mmol/L,but recovered to 90.5%when phosphate concentration increased to 16.10 mmol/L.Meanwhile,the increased concentration of phosphate from 0.32 to 16.10mmol/L reduced the BZF degradation efficacy from 91.5%to 79.0%,whereas SMX removal improved from 37.3%to 62.9%.The m Fe-PS system containing(bi)carbonate,from 4.20 to166.70 mmol/L,reduced CFA and BZF removal efficiencies from 100%to 76.8%and 80.4%,respectively,and SMX from 83.5%to 56.7%within a 120-min reaction time.In addition,alkaline conditions(pH≥8.0)inhibited CFA and BZF degradations,while nonacidic pH(pH≥7.0)remarkably inhibited SMX degradation.Results of the kinetic model indicated the formation of phosphate(H_(2)PO_(4)^(·)/HPO_(4)^(·-))and/or carbonate radicals(CO_(3)^(·-))could limit pharmaceutical removal.The transformation products(TPs)of the pharmaceuticals revealed more incompletely oxidized TPs occurred in the phosphate-and(bi)carbonate-containing m Fe-PS systems,and indicated that H_(2)PO_(4)^(·)/HPO_(4)^(·-)mainly degraded pharmaceuticals via a benzene ring-opening reaction while CO_(3)^(·-)preferentially oxidized pharmaceuticals via a hydroxylation reaction.

Generation of tooth-like structures from integration-free human urine induced pluripotent stem cells

Background:Tooth is vital not only for a good smile,but also good health.Yet,we lose tooth regularly due to accidents or diseases.An ideal solution to this problem is to regenerate tooth with patients’own cells.Here we describe the generation of tooth-like structures from integration-free human urine induced pluripotent stem cells(ifhU-iPSCs).Results:We first differentiated ifhU-iPSCs to epithelial sheets,which were then recombined with E14.5 mouse dental mesenchymes.Tooth-like structures were recovered from these recombinants in 3 weeks with success rate up to 30%for 8 different iPSC lines,comparable to H1 hESC.We further detected that ifhU-iPSC derived epithelial sheets differentiated into enamel-secreting ameloblasts in the tooth-like structures,possessing physical properties such as elastic modulus and hardness found in the regular human tooth.Conclusion:Our results demonstrate that ifhU-iPSCs can be used to regenerate patient specific dental tissues or even tooth for further drug screening or regenerative therapies.

Determination of Uric Acid in Human Urine by Ion-exclusion Chromatography with UV Detection Using Pure Water as Mobile Phase

A simple, rapid and accurate ion-exclusion chromatographic method coupled with a UV detector for the determination of uric acid in human urine samples has been developed. The separation was carried out on an ion-exclusion column using only pure water as mobile phase. The detection wavelength was 254 nm and urine sample was injected directly without any pretreatment. Furthermore, the retention behavior of uric acid on the ion-exclusion column was researched when pure water and 1 mmol·L-1 HCI were used as mobile phase, respectively. The stability of uric acid was also further investigated within 28 days, In this method, the linear range of the calibration curve for uric acid was 0.25--100 mg·L-1, and the detection limit calculated at S/N=3 was 0.02mg·L-1 The proposed ion-exclusion chromatographic method has been used for the determination of uric acid in human urine.

Application of machine learning algorithms to screen potential biomarkers under cadmium exposure based on human urine metabolic profiles

Exposure to environmental cadmium increases the health risk of residents.Early urine metabolic detection using high-resolution mass spectrometry and machine learning algorithms would be advantageous to predict the adverse health effects.Here,we conducted machine learning approaches to screen potential biomarkers under cadmium exposure in 403 urine samples.In positive and negative ionization mode,4207 and 3558 features were extracted,respectively.We compared seven machine learning algorithms and found that the extreme gradient boosting(XGBoost)and random forest(RF)classifiers showed better accuracy and predictive performance than others.Following 5-fold cross-validation,the value of area under curve(AUC)was both 0.93 for positive and negative ionization modes in XGBoost classifier.In the RF classifier,AUC were 0.80 and 0.84 for positive and negative ionization modes,respectively.We then identified a biomarker panel based on XGBoost and RF classifiers.The incorporation of machine learning models into urine analysis using high-resolution mass spectrometry could allow a convenient assessment of cadmium exposure.

Quantification of selected monohydroxy metabolites of polycyclic aromatic hydrocarbons in human urine

An analytical method was developed to quantitatively determine selected monohydroxy metabolites of polycyclic aromatic hydrocarbons(PAHs) in human urine. The procedure included enzymatic hydrolysis to cleave the conjugated metabolites, solid-phase microextraction enrichment, and gas chromatography-mass spectrometry analysis. The method proved to be sensitive enough to detect the selected PAH metabolites in human urine.

Quantification of human urine and serum iodine by inductively coupled plasma mass spectrometry

Objective This paper aims at establishing an inductively coupled plasma mass specrometry(ICP-MS)method for quantification and evaluation of iodine in human urine and serum in routine clinical laboratory.Methods This study was methodology validation research on iodine evaluation using ICP-MS.Ammonia,isopropanol and ultrapure water were mixed at certain ratio

Dispersive Liquid-liquid Microextraction Combined with High-performance Liquid Chromatography for the Determination of Clozapine and Chlorpromazine in Urine

A simple method has been proposed for the determination of clozapine （CLZ） and chlorpromazine （CPZ） in human urine by dispersive liquid-liquid microextraction （DLLME） in combination with high-performance liquid chromatography-ultraviolet detector （HPLC-UV）. All important variables influencing the extraction efficiency, such as pH, types of the extraction solvent and the disperser solvent and their volume, ionic strength and centrifugation time were investigated and optimized. Under the optimal conditions, the limit of detection （LODs） and quantification （LOQs） of the method were 13 and 39 ng/mL for CLZ, and 2 and 6 ng/mL for CPZ, respectively. The relative standard deviations （RSDs） of the targets were less than 5.1% （C=0.100 μg/mL, n=9）. Good linear behaviors over the tested concentration ranges were obtained with the values of R20.999 for the targets. The absolute extraction efficiencies of CLZ and CPZ from the spiked blank urine samples were 98.3% and 97.8%, respectively. The applicability of the technique was validated by analyzing urine samples and the mean recoveries for spiked urine samples ranged from 93.3% to 105.0%. The method was successfully applied for the determination of CLZ and CPZ in real human urine.

A New Glucuronidated Metabolite of Andrographolide in Human

A new andrographolide metabolite 1 was isolated from human urine samples after oral administration. The structure was determined to be 3-carbonylandrographolide-19-O-β-D-glu- curonide on the basis of chemical evidences and spectral analysis, especially by 2D-NMR techni- ques.

Measurement of Orotic Acid in Urine by Supercritical Fluid Chromatography

This work presents a simple, rapid and reliable supercritical fluid chromatography (SFC) method for a sensitive measurement of orotic acid in human urine. The samples were diluted with deionized water and analyzed directly without any pretreatment.

Study of the Optimal Conditions for Anaerobic Digestion of Cow Dung in Households in Cote d’Ivoire

This paper investigates the optimal conditions for methanisation applied to cow dung. Four experimental 25 liters digesters were used in this work. The best biogas yield is obtained when the digester is installed in a metal box and exposed to sunlight. The temperature in this digester varied between 25˚C and 37˚C. The dry matter content of the collected cow dung was 15.5%. The digester was fed with 9 kg of cow dung mixed with 8.5 litres of water, one litre of cassava effluent and 200 ml of human urine. After a retention period of 22 days, the biogas obtained was 67% methane and 21% carbon dioxide. The use of human urine and cassava effluent improved the quality of the biogas.

人尿中美托拉宗的液相色谱-质谱研究

A sensitive and reliable procedure for analysis of metolazone in human urine by HPLC-MS was describes.The extraction recovery of liquid-liquid extraction (LLE) at various pH were studied.The detection limit of the compound was below 0.2ng at absolute amount.It is suitable for metolazone screening and confirmation in doping control.