Transacylation and hydrolysis of the acyl glucuronides of ibuprofen and its α-methyl-substituted analogues investigated by 1H NMR spectroscopy and computational chemistry: Implications for drug design.

Drugs and drug metabolites containing a carboxylic-acid moiety can undergo in vivo conjugation to form 1-ß-O-acyl-glucuronides (1-ß-O-AGs). In addition to hydrolysis, these conjugates can undergo spontaneous acyl migration, and anomerisation reactions, resulting in a range of positional isomers. Facile transacylation has been suggested as a mechanism contributing to the toxicity of acyl glucuronides, with the kinetics of these processes thought to be a factor. Previous 1H NMR spectroscopic and HPLC-MS studies have been conducted to measure the degradation rates of the 1-ß-O-AGs of three nonsteroidal anti-inflammatory drugs (ibufenac, R-ibuprofen, S-ibuprofen) and a dimethyl-analogue (termed here as "bibuprofen"). These studies have also determined the relative contributions of hydrolysis and acyl migration in both buffered aqueous solution, and human plasma. Here, a detailed kinetic analysis is reported, providing the individual rate constants for the acyl migration and hydrolysis reactions observed in buffer for each of the 4 AGs, together with the overall degradation rate constants of the parent 1-ß-O-AGs. Computational modelling of the reactants and transition states of the transacylation reaction using density functional theory indicated differences in the activation energies that reflected the influence of both substitution and stereochemistry on the rate of transacylation/hydrolysis.

Lifestyle and socioeconomic determinants of diabetes: Evidence from country-level data.

OBJECTIVE: The objectives of the study is to investigate the global socioeconomic risk factors associated with diabetes prevalence using evidence from available country-level data. DESIGN: A cross-sectional study based on (2010 & 2019) countrywide Health Nutrition and Population Statistics data. POPULATION: People ages 20-79 who have diabetes. SETTING: One hundred and thirty-two countries or territories in the world. PRIMARY OUTCOME MEASURE: Diabetes prevalence rates were determined from (2010 & 2019) countrywide Health Nutrition and Population Statistics (Health Stats, World Bank Group). RESULTS: In 2010, a 1% increase in per capita income and total tobacco consumption is associated with a 0.92% (95% CI 0.64% to 1.19%) and 0.02% (95% CI 0.006% to 0.047%) increase in diabetes prevalence respectively; and a 1% increase in alcohol consumption is associated with a -0.85% (95% CI -1.17% to -0.53%) decrease in diabetes prevalence. Statistically significant socioeconomic and lifestyle indices positively associated with diabetes prevalence included gross national income; overweight prevalence (BMI>25 kg/m2); and tobacco consumption. Statistically significant inverse associations with global diabetes prevalence included total population size; unemployment and alcohol consumption. The 2019 data was removed due to sparsity of data. CONCLUSION: Statistically significant global lifestyle and socioeconomic determinants of diabetes prevalence include alcohol consumption; tobacco consumption; overweight prevalence; per capita income; total population and unemployment rates. Determinants of diabetes include modifiable risk factors which are consistent at both the micro and macro level and include tobacco consumption and overweight prevalence. Factors which are non-modifiable and warrant further investigation include total population and unemployment rates, which were inversely associated with diabetes prevalence and are a product of other underlying factors. Other determinants such as alcohol consumption was also inversely associated with diabetes prevalence, but has been observed to have both negative and positive associations with diabetes at the micro-level. These associations were dependent upon the amount of alcohol consumed. Global cut-off point of alcohol consumption is critical to establish global policies to reduce diabetes prevalence. Overall, the use of cross-sectional based study for country level aggregate data is a critical tool that should be considered when making global joint strategies or policies against diabetes in both data analysis and decision making.

Kinetic modelling of acyl glucuronide and glucoside reactivity and development of structure-property relationships.

Acyl glucuronide metabolites have been implicated in the toxicity of several carboxylic acid-containing drugs, and the rate of their degradation via intramolecular transacylation and hydrolysis has been associated with the degree of protein adduct formation. Although not yet proven, the formation of protein adducts in vivo - and subsequent downstream effects - has been proposed as a mechanism of toxicity for carboxylic acid-containing xenobiotics capable of forming acyl glucuronides. A structurally-related series of metabolites, the acyl glucosides, have also been shown to undergo similar degradation reactions and consequently the potential to display a similar mode of toxicity. Here we report detailed kinetic models of each transacylation and hydrolysis reaction for a series of phenylacetic acid acyl glucuronides and their analogous acyl glucosides. Differences in reactivity were observed for the individual transacylation steps between the compound series; our findings suggest that the charged carboxylate ion and neutral hydroxyl group in the glucuronide and glucoside conjugates, respectively, are responsible for these differences. The transacylation reaction was modelled using density functional theory and the calculated activation energy for this reaction showed a close correlation with the degradation rate of the 1-ß anomer. Comparison of optimised geometries between the two series of conjugates revealed differences in hydrogen bonding which may further explain the differences in reactivity observed. Together, these models may find application in drug discovery for prediction of acyl glucuronide and glucoside metabolite behaviour.

Metabolic phenotype modulation by caloric restriction in a lifelong dog study.

Modeling aging and age-related pathologies presents a substantial analytical challenge given the complexity of gene-environment influences and interactions operating on an individual. A top-down systems approach is used to model the effects of lifelong caloric restriction, which is known to extend life span in several animal models. The metabolic phenotypes of caloric-restricted (CR; n = 24) and pair-housed control-fed (CF; n = 24) Labrador Retriever dogs were investigated by use of orthogonal projection to latent structures discriminant analysis (OPLS-DA) to model both generic and age-specific responses to caloric restriction from the ¹H NMR blood serum profiles of young and older dogs. Three aging metabolic phenotypes were resolved: (i) an aging metabolic phenotype independent of diet, characterized by high levels of glutamine, creatinine, methylamine, dimethylamine, trimethylamine N-oxide, and glycerophosphocholine and decreasing levels of glycine, aspartate, creatine and citrate indicative of metabolic changes associated largely with muscle mass; (ii) an aging metabolic phenotype specific to CR dogs that consisted of relatively lower levels of glucose, acetate, choline, and tyrosine and relatively higher serum levels of phosphocholine with increased age in the CR population; (iii) an aging metabolic phenotype specific to CF dogs including lower levels of liproprotein fatty acyl groups and allantoin and relatively higher levels of formate with increased age in the CF population. There was no diet metabotype that consistently differentiated the CF and CR dogs irrespective of age. Glucose consistently discriminated between feeding regimes in dogs (≥312 weeks), being relatively lower in the CR group. However, it was observed that creatine and amino acids (valine, leucine, isoleucine, lysine, and phenylalanine) were lower in the CR dogs (<312 weeks), suggestive of differences in energy source utilization. ¹H NMR spectroscopic analysis of longitudinal serum profiles enabled an unbiased evaluation of the metabolic markers modulated by a lifetime of caloric restriction and showed differences in the metabolic phenotype of aging due to caloric restriction, which contributes to longevity studies in caloric-restricted animals. Furthermore, OPLS-DA provided a framework such that significant metabolites relating to life extension could be differentiated and integrated with aging processes.

Colonization-induced host-gut microbial metabolic interaction.

UNLABELLED: The gut microbiota enhances the host's metabolic capacity for processing nutrients and drugs and modulate the activities of multiple pathways in a variety of organ systems. We have probed the systemic metabolic adaptation to gut colonization for 20 days following exposure of axenic mice (n = 35) to a typical environmental microbial background using high-resolution (1)H nuclear magnetic resonance (NMR) spectroscopy to analyze urine, plasma, liver, kidney, and colon (5 time points) metabolic profiles. Acquisition of the gut microbiota was associated with rapid increase in body weight (4%) over the first 5 days of colonization with parallel changes in multiple pathways in all compartments analyzed. The colonization process stimulated glycogenesis in the liver prior to triggering increases in hepatic triglyceride synthesis. These changes were associated with modifications of hepatic Cyp8b1 expression and the subsequent alteration of bile acid metabolites, including taurocholate and tauromuricholate, which are essential regulators of lipid absorption. Expression and activity of major drug-metabolizing enzymes (Cyp3a11 and Cyp2c29) were also significantly stimulated. Remarkably, statistical modeling of the interactions between hepatic metabolic profiles and microbial composition analyzed by 16S rRNA gene pyrosequencing revealed strong associations of the Coriobacteriaceae family with both the hepatic triglyceride, glucose, and glycogen levels and the metabolism of xenobiotics. These data demonstrate the importance of microbial activity in metabolic phenotype development, indicating that microbiota manipulation is a useful tool for beneficially modulating xenobiotic metabolism and pharmacokinetics in personalized health care. IMPORTANCE: Gut bacteria have been associated with various essential biological functions in humans such as energy harvest and regulation of blood pressure. Furthermore, gut microbial colonization occurs after birth in parallel with other critical processes such as immune and cognitive development. Thus, it is essential to understand the bidirectional interaction between the host metabolism and its symbionts. Here, we describe the first evidence of an in vivo association between a family of bacteria and hepatic lipid metabolism. These results provide new insights into the fundamental mechanisms that regulate host-gut microbiota interactions and are thus of wide interest to microbiological, nutrition, metabolic, systems biology, and pharmaceutical research communities. This work will also contribute to developing novel strategies in the alteration of host-gut microbiota relationships which can in turn beneficially modulate the host metabolism.

Synthesis of a series of phenylacetic acid 1-ß-O-acyl glucosides and comparison of their acyl migration and hydrolysis kinetics with the corresponding acyl glucuronides.

We report the synthesis of the 1-ß-O-acyl glucoside conjugates of phenylacetic acid (PAA), R- and S-α-methyl-PAA and α,α'-dimethyl-PAA, and measurement of their transacylation and hydrolysis reactivity by NMR methods. These are analogues of acyl glucuronides, the transacylation kinetics of which could be important in adverse drug effects. One aim of this work was to investigate whether, as previously postulated, the free carboxylate group of the acyl glucuronides plays a part in the mechanism of the internal acyl migration. In addition, such acyl glucosides are known to be endogenous biochemicals in their own right and investigation of their acyl migration propensities is novel. Our previously described selective acylation procedure has proved highly successful for 1-ß-O-acyl glucuronide synthesis and when subsequently applied to 6-O-trityl glucose, it gave good yields and excellent anomeric selectivity. Mild acidolysis of the O-trityl intermediates gave the desired acyl glucosides in excellent yield with essentially complete ß-selectivity. Measurement of the acyl glucoside transacylation kinetics by (1)H NMR spectroscopy, based simply on the disappearance of the 1-ß-isomer in aqueous buffer at pH 7.4, showed marked differences depending on the degree of methyl substitution. Further kinetic modelling of the isomerisation and hydrolysis reactions of the acyl glucosides showed considerable differences in kinetics for the various isomeric reactions. Reactions involving the -CH(2)OH group, presumably via a 6-membered ring ortho-ester intermediate, are facile and the α-glucoside anomers are significantly more reactive than their ß-counterparts. By comparison with degradation rates for the corresponding acyl glucuronides, it can be inferred that substitution of the carboxylate by -CH(2)OH in the acyl glucosides has a significant effect on acyl migration for those compounds, especially for rapidly transacylating molecules, and that thus the charged glucuronide carboxylate is a factor in the kinetics.

Self-modeling curve resolution recovery of temporal metabolite signal modulation in NMR spectroscopic data sets: application to a life-long caloric restriction study in dogs.

A novel model-free statistical approach (self modeling curve resolution, SMCR) has been applied to recover biochemical information from complex overlapping signals in (1)H NMR spectra of blood serum in a long-term study of caloric restriction (CR) in the dog (n = 24 control fed (CF) and n = 24 CR animals). A new statistical spectroscopic construct, the spectrotype, is proposed which is a spectroscopic subset description or component of a metabolic phenotype. Characterization of the (1)H NMR profiles according to their evolutionary contribution of each spectrotype gives clues to the kinetics of the macro-biochemical response profiles and the identity of the underlying biochemical constituents, governing the evolutionary global response to an intervention. This information can be used to monitor and predict the end point of the biological process and to identify the mechanisms responsible for those changes. Here a SMCR strategy together with a pattern recognition method, principal component analysis (PCA) was used to resolve sets of spectrotypes, without a priori information. From the (1)H NMR evolutionary response profiles, two spectrotypes were identified and resolved; spectrotype 1 dominated by lipids featuring contributions from phosphatidylcholine, lipoprotein lipid fatty acyl groups from triglycerides, phospholipids, and cholesteryl esters plus total cholesterol (i.e., both esterified and unesterified); spectrotype 2 comprising glucose signals and a poorly resolved envelope of albumin and N-acetylated glycoprotein resonances. The relative contributions of these spectrotypes in each sample were calculated. For both caloric restricted (CR) and control fed (CF) dogs between ages 1 and 9 years, the contribution of spectrotype 2 > spectrotype 1, whereas for dogs aged between 9 and 12 years spectrotype 1 > spectrotype 2. Therefore, SMCR analysis pinpointed ages where nutrition and aging metabolic changes became significant within serum samples as well as providing the individual longitudinal contribution profiles associated with each spectrotype, which could potentially be used as part of a strategy to monitor and predict longevity and morbidity in populations. Hence SMCR is a useful addition to the chemometric "toolbox" for metabolic analysis and should have diverse applications within other biomedical conditions characterized by subtle time-dependent changes.