Metabolite characterisation in peritoneal dialysis effluent using high-resolution (1) H and (1) H-(13) C NMR spectroscopy.

Metabolite analysis of peritoneal dialysis (PD) effluent may provide information regarding onset and progression of complications associated with prolonged PD therapy. In this context, the nuclear magnetic resonance detectable small metabolites of PD effluent samples were characterised using high-resolution (1) H and (1) H-(13) C NMR spectroscopy. The various spectra were recorded (at 800 MHz proton frequency) on PD effluent samples obtained after 4-h (intraperitoneal) dwell time from patients with end-stage renal failure and continuing normally on PD therapy. In spite of devastating spectral feature of PD effluent due to the presence of intense resonances from glucose and lactate, we were able to identify 53 small endogenous metabolites (including many complex coupled spin systems) and more than 90% of the total CH cross peaks of (1) H-(13) C heteronuclear single-quantum correlation spectrum specific to various metabolites of PD effluent. We foresee that the characteristic fingerprints of various metabolites of control PD effluent samples will be used to identify and distinguish metabolic differences from PD-related complications.

"Perfect echo" INEPT: more efficient heteronuclear polarization transfer by refocusing homonuclear J-coupling interaction.

A "perfect echo" based INEPT experiment that demonstrates more efficient heteronuclear polarization transfer over conventional INEPT has been developed. This scheme refocuses the effect of homonuclear (1)H-(1)H J-evolution and simultaneously allows heteronuclear (13)C-(1)H J-evolution to continue during INEPT. This improves one bond heteronuclear polarization transfer efficiency at longer INEPT transfer delays and also enhances the sensitivity of long range INEPT. The refocusing of homonuclear (1)H-(1)H J-coupling could be achieved by doubling the INEPT transfer period leading to a doubling of T2 losses. Therefore, the sensitivity gain is observed when loss of magnetization due to homonuclear (1)H-(1)H J-modulation is more severe than that of T2 decay. However, in general, INEPT transfer period is rather short compared to the longer T2 observed in small molecules. The long range PE-INEPT transfer to carbonyl carbon in beta-butyrolactone, showed much faster build up of C-13 signal than conventional long range INEPT as the long range heteronuclear J-coupling is comparable in magnitude to homonuclear (1)H-(1)H J-coupling in this case. For one bond heteronuclear polarization transfer at shorter delay, PE-INEPT and conventional INEPT displays equal transfer efficiency. Efficient polarization transfer is observed for small molecules dissolved in isotropic as well as weakly aligned media. Further, simulation results obtained using the full propagator and product operator analysis agree well with the experimental observations.

Effect of SOHAM meditation on human brain: a voxel-based morphometry study.

The anatomical correlates of long-term meditators involved in practice of "SOHAM" meditation have been studied using voxel-based morphometry (VBM). The VBM analysis indicates significantly higher gray matter density in brain stem, ventral pallidum, and supplementary motor area in the meditators as compared with age-matched nonmeditators. The observed changes in brain structure are compared with other forms of meditation.

"Perfect echo" HMQC: sensitivity and resolution enhancement by broadband homonuclear decoupling.

Homonuclear (1)H-(1)H J-modulation leads to J-multiplets in F1 dimension of 2D (1)H-(13)C HMQC spectra. This hampers unambiguous signal assignment for overcrowded (13)C spectra. Broadband homonuclear decoupling has been achieved in the indirect t1 evolution period by incorporating blocks of perfect echo. This method of perfect echo HMQC demonstrates better resolution and sensitivity than conventional HMQC spectra. The results on Cyclosporine demonstrate that the method is very efficient for refocusing geminal couplings in weakly coupled -(13)CH2 groups. Partial refocusing of vicinal couplings is also observed for -(13)CH and -(13)CH3 groups. Interpretation of the result based on product operator formalism is also given. Comparison of pe-HMQC, HMQC and HSQC reveals that the F1 linewidth of pe-HMQC is much narrower than HMQC and very close to that of HSQC for CH2 groups.

Distinctive neural signatures for negative sentences in Hindi: an fMRI study.

We examined cortical activations using functional magnetic resonance imaging (fMRI) technique in skilled native Hindi readers while they performed a 'target-probe' semantic judgment task on affirmative and negative sentences. Hindi, an Indo-Aryan language widely spoken in India, follows subject-object-verb (SOV) order canonically but allows free word order. The common cortical regions involved in affirmative and negative sentence conditions included bilateral inferior frontal gyrus (IFG), left parietal cortex (BA 7/40), left fusiform (BA 37), bilateral supplementary motor area (SMA) (BA 6), bilateral middle temporal gyrus (BA 21), and bilateral occipital area (BA 17/18). While no distinct region was activated for affirmative sentences, we observed activations in the region of bilateral anterior temporal pole for negative sentence. The behavioral results showed no significant mean difference for reaction times (RT) and accuracy measures between affirmative and negative sentences. However, the imaging results suggest the recruitment of anterior temporal pole in processing of negative sentences. Region of interest (ROI) analysis for selected regions showed higher signal intensity for negative sentences possibly indicating the associated inherent difficulty level of processing, especially when integrating information related to negations.

Frequency and factors associated with increased small intestinal permeability in patients with portal hypertension.

AIM: Cirrhosis with portal hypertension (PHT) may be associated with increased small intestinal permeability (SIP), predisposing to malnutrition and bacterial translocation causing septicaemia, endotoxaemia and spontaneous bacterial peritonitis. However, data on SIP in extrahepatic portal venous obstruction (EHPVO), in which PHT occurs without hepatic dysfunction, are scanty. Such studies would help to know the effect of PHT on SIP independent of hepatic dysfunction; hence, we undertook this study. METHODS: A total of 96 patients with PHT (cirrhosis 71, EHPVO 25) underwent evaluation of SIP using urinary lactulose/mannitol excretion ratio over 6 hours after oral administration of 15 mL (10 g) lactulose and 5 g mannitol using 1H-NMR spectroscopy by a method described by us previously. RESULTS: Gender of patients with EHPVO and cirrhosis was comparable but patients with EHPVO were younger in age. The causes of cirrhosis were cryptogenic (n = 22), alcohol (n = 20), post-viral (n = 21) and others (n = 8). Twenty-seven (38%) patients with cirrhosis had ascites. Abnormal SIP was detected in 47 (49%) patients (40/71,56% with cirrhosis vs. 7/25, 28% with EHPVO, p = 0.01). Patients with cirrhosis had a higher urinary lactulose/mannitol excretion ratio than those with EHPVO (0.09, range 0-0.87 mmol vs. 0.05, 0-0.19 mmol; p = 0.008). Patients with abnormal SIP had a higher Child score, and more often had cirrhosis than EHPVO, ascites and deranged liver function. On multivariate analysis, presence of cirrhosis, ascites, high serum bilirubin level and prothrombin time were associated with abnormal SIP. CONCLUSIONS: Cirrhosis was associated with abnormal SIP, which was related to liver dysfunction. However, SIP was normal in patients with EHPVO.

Metabolic alterations of Withania somnifera (L.) dunal fruits at different developmental stages by NMR spectroscopy.

INTRODUCTION: Withania somnifera (Ashwagandha) is a high-value Ayurvedic medicinal plant and an important constituent of several dietary supplements. In order to substantiate the health claims, the herb has drawn considerable scientific attention. OBJECTIVE: The objective of the study was to investigate the alterations in primary and secondary metabolites of W. somnifera fruits during its maturity using NMR spectroscopy. METHODOLOGY: Fruits at different stages of development from one week after fertilisation until maturity, classified in seven developmental stages, were analysed by a combined use of one- and two-dimensional NMR experiments. RESULTS: Seventeen metabolites were characterised and quantified from non-polar and polar extracts of different fruit development stages of W. somnifera. The principal component analysis of polar metabolites at different stages could be grossly classified into three metabolic phases, viz. initial phase, developmental phase and maturation phase. CONCLUSION: Qualitative and quantitative analysis of metabolites in W. somnifera fruits indicated specific stages when fruits can be harvested for obtaining substantial bioactive ingredients for desirable pharmacological activity. This study potentially provides a complementary tool for quality control of herbal medicinal products when W. somnifera fruits are used.

Abnormal small intestinal permeability in patients with idiopathic malabsorption in tropics (tropical sprue) does not change even after successful treatment.

BACKGROUND: Though tropical sprue (TS) is common in tropics, studies on small intestinal permeability (SIP) in TS are scant. METHOD: SIP was evaluated using (1)H nuclear magnetic resonance (NMR) spectroscopy of urinary lactulose and mannitol in 24 patients with TS (22 before and 15 after treatment with tetracycline and folate) and in 31 healthy subjects (HS). Effect of treatment of TS on SIP and its relationship with outcome were studied. RESULT: Subjects were comparable in terms of age and gender. Before treatment, urinary lactulose (0.24 mmol, 0-1.09 mmol versus 0.09 mmol, 0-0.68 mmol, P=0.02) and lactulose-to-mannitol (L/M) ratio (0.11, 0-0.41 versus 0.042, 0-0.26, P=0.001) were higher in TS than in HS, though mannitol was comparable (2.7 mmol, 0.61-10.5 mmol versus 3.8 mmol, 1.3-16.4 mmol, P=0.08). Patients improved after treatment [stool frequency (9, 4-20/day versus 1, 1-2/day, P<0.0001), weight (44.4, 32-69 kg, versus 56, 39-84 kg, P<0.0001), fecal fat (10.1, 6-26 g/24 h versus 4.4, 3.0-6.7 g/24 h, P<0.0001), D-xylose (0.57, 0.28-1.2 g/5 g/5 h versus 1.1, 0.2-2.1 g/5 g/5 h, P<0.0001), and small intestinal bacterial overgrowth (SIBO) resolved in 10/24 (41.7%) versus 1/15 (6.6%), P=0.02]. Though urinary lactulose (0.17, 0-4.3 mmol versus 0.09, 0-0.68 mmol, P=0.11) and mannitol (2.17, 0.8-36.7 mmol versus 3.84, 1.3-16.4 mmol, P=0.06) were comparable, L/M ratio was higher in TS than in HS (0.09, 0-0.22 versus 0.042, 0-0.26, P=0.002). L/M ratio was more often abnormal (cutoff 0.078) in TS than in HS [14/22 (63.6%) versus 3/31 (9.7%); P=0.0001], which persisted even after treatment [9/15 (60%) as compared with HS; P=0.0006]. Persistently abnormal SIP was associated with less weight gain and frequent stools following treatment. CONCLUSION: SIP is often abnormal in TS and remains unchanged even after successful treatment that was associated with less weight gain and more frequent stool.

Comprehensive metabolic fingerprinting of Withania somnifera leaf and root extracts.

Profiling of metabolites is a rapidly expanding area of research for resolving metabolic pathways. Metabolic fingerprinting in medicinally important plants is critical to establishing the quality of herbal medicines. In the present study, metabolic profiling of crude extracts of leaf and root of Withania somnifera (Ashwagandha), an important medicinal plant of Indian system of medicine (ISM) was carried out using NMR and chromatographic (HPLC and GC-MS) techniques. A total of 62 major and minor primary and secondary metabolites from leaves and 48 from roots were unambiguously identified. Twenty-nine of these were common to the two tissues. These included fatty acids, organic acids, amino acids, sugars and sterol based compounds. Eleven bioactive sterol-lactone molecules were also identified. Twenty-seven of the identified metabolites were quantified. Highly significant qualitative and quantitative differences were noticed between the leaf and root tissues, particularly with respect to the secondary metabolites.

Metabolic and histopathological alterations of Jatropha mosaic begomovirus-infected Jatropha curcas L. by HR-MAS NMR spectroscopy and magnetic resonance imaging.

Alterations in the anatomical structures, sap translocation and metabolic profiles in Jatropha curcas L. (Euphorbiaceae), infected with Jatropha mosaic virus (JMV) have been investigated using MRI and HR-MAS NMR spectroscopy. The contrast of MRI images distinguishes abnormalities in anatomical structures of infected and healthy stem. The HR-MAS NMR spectroscopic analysis indicated that viral infection significantly affected the plant metabolism. Higher accumulation of TCA cycle intermediates, such as citrate and malate, in JMV-infected plants suggested a higher rate of respiration. The respiration rate was more than twofold as compared to healthy ones. The viral stress also significantly increases the concentrations of alanine, arginine, glutamine, valine, GABA and choline as compared to healthy ones. Microscopic examination revealed severe hyperplasia caused by JMV with a considerable reduction in the size of stem cells. Lower concentration of glucose and sucrose in viral-infected stem tissues indicates decreased translocation of photosynthates from leaves to stem due to hyperplasia caused by JMV. The MR images distinguished stele, cortical and pith regions of JMV-infected and healthy stems. Contrast of T(1)- and T(2)-weighted images showed significant differences in the spatial distribution of water, lipids and macromolecules in virus-infected and healthy stem tissues. The results demonstrated the value of MRI and HR-MAS NMR spectroscopy in studying viral infection and metabolic shift in plants. The present methodology may help in better understanding the metabolic alterations during biotic stress in other plant species of agricultural and commercial importance.

Metabolomic analysis of serum by (1) H NMR spectroscopy in amyotrophic lateral sclerosis.

BACKGROUND: Amyotrophic lateral sclerosis (ALS), an invariably fatal neurological disorder shows complicated pathogenesis that poses challenges with respect to diagnosis as well as monitoring of disease progression. METHODS: We investigated metabolite profiles in the serum of 30 patients with ALS, 10 patients of Hirayama disease, which served as a neurological disease control and 25 healthy controls by using (1) H NMR spectroscopy. RESULTS: Compared to healthy controls, the ALS patients had higher quantities of glutamate (P<0.001), beta-hydroxybutyrate (P<0.001), acetate (P<0.01), acetone (P<0.05), and formate (P<0.001), and lower concentrations of glutamine (P<0.02), histidine (P<0.001) and N-acetyl derivatives. On the other hand, Hirayama disease patients had significantly higher median concentrations of pyruvate (P<0.05), glutamate (P<0.001), formate (P<0.05) and lower median concentrations of N-acetyl derivatives. Furthermore, we also found that serum glutamate showed a positive correlation (P<0.001, r=0.6487) whereas, histidine showed a negative correlation (P<0.001, r=-0.5641) with the duration of the disease in ALS. CONCLUSIONS: Such (1) H NMR study of serum may reveal abnormal metabolite patterns, which could have the potential to serve as surrogate markers for monitoring ALS disease progression.

1H NMR spectroscopic study of blood serum for the assessment of liver function in liver transplant patients.

BACKGROUND AND AIMS: Plausible reasons for the failure of liver graft in liver transplantation are explored. 1H-NMR spectroscopy of serum is employed for assessment of liver graft function. Differences in concentrations of specific metabolites between patients with successful and unsuccessful liver grafts following transplantation were used as possible markers to assess the graft quality. METHODS: Blood samples from the patients undergoing liver transplantation were obtained preoperatively, immediately after transplant followed by every 24 hrs of post-transplantation until patients were discharged or expired. 1H-NMR spectroscopic studies of serum were performed at each time point and concentrations of various metabolites measured. Conventional biological tests were also performed at each time point. RESULTS: Elevation of concentrations of the nine metabolites (lactate, alanine, lysine, glutamine, methionine, asparagine, tyrosine, histidine and phenylalanine) in non-survivors using NMR was attributed to the graft dysfunction. The information on the graft dysfunction using conventional biological tests was obtained much later. However, elevation in aminotransferases and bilirubin levels was indicated after about one week and 3 days respectively in non-survivors. Hepatic failure causes alteration in the concentrations of amino acids due to impairment of amino acid metabolism and urea cycle. 1H-NMR spectroscopy provides the information of all the metabolites in a single step without involving any chemical pretreatment implying better accuracy since each step involved can introduce its own experimental error. CONCLUSION: Distinct metabolic profile in non-survivors compared to survivors following transplantation promises potential of 1H-NMR studies in the assessment of liver graft function.

(1)H and (31)P NMR studies indicate reduced bile constituents in patients with biliary obstruction and infection.

Patients with extrahepatic biliary obstruction present with impairment of the normal bile flow, with jaundice and cholangitis as common complications. (1)H and (31)P NMR quantitative analysis of bile specimens from patients with extrahepatic biliary obstruction (n = 80) (with/without jaundice and cholangitis separately and together) was carried out for the chief biliary constituents to determine the relationship between biliary constituents and jaundice (serum bilirubin concentration >or=1.0 mg/dL) and cholangitis (total leucocyte count >11,000 cells/mm(3) and/or fever >38.5 degrees C with/without bile culture positivity). Compared with controls (patients without jaundice and cholangitis), median indices of the chief biliary constituents (total bile acids, cholesterol, phosphatidylcholine and inorganic phosphate) were significantly suppressed in patients with cholangitis and/or jaundice. Quantities of total bile acids, cholesterol and phosphatidylcholine correlated negatively with the quantity of bilirubin and with cholangitis, i.e. total leucocyte count. Suppression of biliary constituents correlated significantly with the severity of jaundice and cholangitis. The decrease in biliary constituents in the presence of jaundice and cholangitis is possibly the result of downregulation of the function of transporters located at the canalicular side of hepatocytes, leading to their suppressed indices in bile. This information may have implications in the examination of bile for clinical studies.

Room-temperature and low-ordered, amphotropic-lyotropic ionic liquid crystal phases induced by alcohols in phosphonium halides.

Tri- n-decylmethylphosphonium chloride and bromide ( 1P10X) salts are not liquid crystalline. However, mesophases are induced by adding very small amounts of an alcohol or water. The temperature ranges of the induced smectic A 2 (SmA 2) liquid-crystalline phases can be very broad and the onset temperatures can be below room temperature depending upon the concentration of the alcohol or water and the structure of the alcohol. At least one molar equivalent of hydroxyl groups is necessary to convert the 1P10X completely into a liquid crystal. Strong association between the hydroxyl groups of an alcohol or water and the head groups of the 1P10X is indicated by spectroscopic, diffraction, and thermochemical data. Unlike many other smectic phases, those of the 1P10X/alcohol complexes are easily aligned in strong magnetic fields and the order parameters of selectively deuterated alcohols as measured by (2)H NMR spectroscopy, approximately 10 (-2), are much lower than the values found when the host is a commonly employed thermotropic liquid crystal. The dependence of the specific values of the order parameters on temperature, the nature of the halide anion, and the structure and concentration of the alcohol are reported. In sum, a detailed picture is presented to explain how and why an alcohol or water induces liquid crystallinity in the 1P10X salts. The data also provide a blueprint for designing media with even lower order parameters that can be hosts to determine the conformations and shapes of guest molecules.

Lipid profiling of developing Jatropha curcas L. seeds using (1)H NMR spectroscopy.

Seed development in Jatropha curcas L. was studied with respect to phenology, oil content, lipid profile and concentration of sterols. Seeds were collected at various stages of development starting from one week after fertilization and in an interval of five days thereafter till maturity. These were classified as stage I to stage VII. Moisture content of the seeds ranged from 8.8 to 90.3%; the lowest in mature seeds in stage VII and highest in stage I. The seed area increased as the seed grew from stage I to stage VI (0.2-10.2mm(2) per seed), however, the seed area shrunk at stage VII. Increase in seed area corresponded to increase in fresh weight of the seeds. (1)H NMR spectroscopy of hexane extracts made at different stages of seed development revealed the presence of free fatty acids (FFA), methyl esters of fatty acids (FAME) and triglycerol esters (TAG), along with small quantity of sterols. The young seeds synthesized predominantly polar lipids. Lipid synthesis was noticed nearly three weeks after fertilization. From the fourth week the seeds actively synthesized TAG. Stage III is a turning point in seed development since at this stage, the concentration of sterols decreased to negligible, there was very little FAME formation, accumulation of TAG increased substantially, and there was a sudden decrease in FFA concentration. The findings can be helpful in understanding the biosynthesis and in efforts to improve biosynthesis of TAG and reduce FFA content in the mature seeds.

Aluminum-mediated metabolic changes in rat serum and urine: a proton nuclear magnetic resonance study.

The toxic effects of Al(3+) have been studied in 90-days AlCl(3) orally treated male albino rats (n = 7) using (1)H NMR spectroscopy-based metabolic profile of rat serum and urine, serum enzyme tests, behavioral impairment, and histopathology of kidney and liver. Metabolic profile of 90-days Al(3+)-treated rat sera showed significantly elevated levels of alanine, glutamine, beta-hydroxy-butyrate, and acetoacetate and significantly decreased level of acetone when compared with that of control rats. However, metabolic profile of 90-days Al(3+)-treated rat urine showed significantly decreased levels of citrate, creatinine, allantoin, trans-aconitate, and succinate and significantly increased level of acetate when compared to control rats. The overall perturbations observed in the metabolic profile of serum and urine demonstrate the impairment in the tricarboxylic acid cycle, liver and kidney metabolism, which was further reinstated by clinical chemistry and histopathological observations. Moreover, "in vivo" behavioral impairment has also been observed as the indication of aluminum neurotoxicity.

Induced amphotropic and thermotropic ionic liquid crystallinity in phosphonium halides: "lubrication" by hydroxyl groups.

The influence of covalently attaching hydroxymethylene to the methyl groups of methyl-tri-n-alkylphosphonium halides (where the alkyl chains are decyl, tetradecyl, or octadecyl and the halide is chloride or bromide) or adding methanol as a solute to the salts on their solid, liquid-crystalline (smectic A2), and isotropic phases has been investigated using a variety of experimental techniques. These structural and compositional changes are found to induce liquid crystallinity in some cases and to enhance the temperature range and lower the onset temperature of the liquid-crystalline phases in some others. The results are interpreted in terms of the lengths of the three n-alkyl chains attached to the phosphorus cation, the nature of the halide anion, the influence of H-bonding interactions at the head group regions of the layered phases, and other solvent-solute interactions. The fact that at least 1 molar equiv of methanol must be added to effect complete (isothermal) conversion of a solid methyl-tri-n-alkylphosphonium salt to a liquid crystal demonstrates a direct and strong association between individual methanol molecules and the phosphonium salts. Possible applications of such systems are suggested.

(1)H NMR spectroscopy of ascitic fluid: discrimination between malignant and benign ascites and comparison of the results with conventional methods.

It is often difficult to distinguish benign ascites from malignant ascites by conventional examination of ascitic fluid. Therefore, (1)H NMR spectroscopy of ascitic fluid specimens was explored as a one-shot experiment to identify potentially interesting metabolic indices that might help to differentiate between the two. Seventy ascitic fluid specimens (15 cytologically positive for malignant cells, eight cytologically negative for malignant cells but remaining suspicious for malignant ascites, and 47 due to liver cirrhosis) were subjected to (1)H NMR spectroscopy for quantitative estimation of 14 metabolites. Mean concentrations of the metabolites were compared with the Mann-Whitney U test. Multivariate discriminant function analysis was performed to determine important descriptors in the discrimination process. The sensitivity and specificity of the proposed model were compared with conventional methods using ascitic fluid protein and serum ascitic albumin gradient. Then, probable predictions for the doubtful cases were made using the proposed model. Patients with malignant ascites had significantly higher mean concentrations (microM) of beta-hydroxybutyrate (594 vs 61), lactate (5384 vs 2104), acetone (136 vs 69), and acetoacetate (122 vs 48) than patients with cirrhotic ascites, and significantly lower concentrations of glutamine (359 vs 615), citrate (62 vs 118), glucose (4933 vs 8411), tyrosine (44 vs 124), and phenylalanine (51 vs 93) (P < 0.05 for all). In the discriminant function analysis model, the best discrimination (P < 0.001) was achieved when beta-hydroxybutyrate, lactate, citrate and tyrosine were considered together as markers. Sensitivity and specificity of the proposed model, ascitic fluid protein and serum ascitic albumin gradient were found to be 100% and 97.9%, 53.3% and 76.6%, and 60% and 87.2%, respectively. The proposed model put five of the eight doubtful cases in the malignant group. This is encouraging and may provide useful information for clinical purposes.

One-step analysis of major bile components in human bile using 1H NMR spectroscopy.

Human gallbladder bile dissolved in dimethylsulfoxide provides sharp and resolved signals for major bile components in 1H NMR spectra. Characteristic well-resolved marker signals that invariably appear in 1H NMR spectra of bile were identified for cholesterol (H18 methyl signal at 0.643 ppm), lipids (glycerol CH signal at 5.064 ppm), total bile acids (H18 signals in the range 0.520-0.626 ppm), total glycine conjugated bile acids (NH signal at 6.958 ppm), total taurine conjugated bile acids (NH signal at 7.646 ppm), and urea (NH2 signal near 5.48 ppm), which enabled their rapid and accurate analysis. Excellent linearity and precision of quantitative analysis was observed for all the identified bile components (R2 > 0.99 for all). The method was demonstrated on gallbladder bile from 19 patients with gallbladder diseases. Urea in bile was identified by NMR for the first time and its quantitative analysis, along with several other bile components, is presented. The majority of the bile components could be analyzed in a single step. Accurate and rapid quantification of several bile components noninvasively by using the method presented herein may have far-reaching implications in the study of bile acid metabolism and pathophysiology of various hepatobiliary and gastrointestinal diseases.

Single-step analysis of individual conjugated bile acids in human bile using 1H NMR spectroscopy.

1H and 13C NMR spectra of intact human bile were assigned using one-dimensional (1H and 13C) and two-dimensional (1H-1H and 1H-13C) experiments. Individual conjugated bile acids--glycocholic acid, glycodeoxycholic acid, glycochenodeoxycholic acid, taurocholic acid, taurodeoxycholic acid, and taurochenodeoxycholic acid--were identified. The bile acids were quantified accurately and individually in a single step by using distinct and characteristic amide signals. Making use of 13C NMR, the study also suggests a way to analyze unconjugated bile acids separately, if present. Chemical shift assignments and rapid single-step analysis of individual conjugated bile acids from intact bile presented herein may have immense utility in the study of bile acid metabolism and deeper understanding of hepatobiliary diseases.