Obesity is a complex multifactorial disease involving genetic and environmental factors and influencing several different metabolic pathways. In this regard, metabonomics, that is the study of ...complex metabolite profiles in biological samples, may provide a systems approach to understand the global metabolic regulation of the organism in relation to this peculiar pathology. In this pilot study, we have applied a nuclear magnetic resonance (NMR)-based metabolomic approach on urinary samples of morbidly obese subjects. Urine samples of 15 morbidly obese insulin-resistant (body mass index>40; homeostasis assessment model of insulin resistance>3) male patients and 10 age-matched controls were collected, frozen and analyzed by high-resolution 1H-NMR spectroscopy combined with partial least squares-discriminant analysis. Furthermore, two obese patients who underwent bariatric surgery (biliopancreatic diversion and gastric bypass, respectively) were monitored during the first 3 months after surgery and their urinary metabolic profiles were characterized. NMR-based metabolomic analysis allowed us to identify an obesity-associated metabolic phenotype (metabotype) that differs from that of lean controls. Gut flora-derived metabolites such as hippuric acid, trigonelline, 2-hydroxyisobutyrate and xanthine contributed most to the classification model and were responsible for the discrimination. These preliminary results confirmed that in humans the gut microflora metabolism is strongly linked to the obesity phenotype. Moreover, the typical obese metabotype is lost after weight loss induced by bariatric surgery.
It is well established that different factors affect milk composition in cows and that milk composition, in turn, affect both technological and nutritional qualities. In this respect the ...comprehension of the metabolic variability of milk composition in relation to the lactation time as well as to the genetic background may be of paramount importance for the agri-food industries. In the present study we investigated the variations of the metabolic profiles during lactation in milks obtained from Friesian and autochthonous races from Northern Italy by
1
H NMR metabolomics. Furthermore, the external factors influencing the milk composition were minimized: the cows were breeded in the same farm, were fed with the same diet and were paired for the lactation interval and lactation stage. Our results showed a difference in milk composition between races and in relation to late lactation. The PLS-DA analysis permitted to distinguish the Friesian and autochthonous cow milks at the investigated different lactation times. Interestingly, the metabolites significantly involved into the discrimination between races appeared to be also technological property parameters, highlighting the importance of maintaining the biodiversity of cow breeds. Therefore, NMR-based metabolomics of milk could represent an informative tool to identify metabolites involved in milk quality both from a nutritional and industrial perspective.
The metabolic profiling of kiwifruit (
Actinidia deliciosa,
Hayward cultivar) aqueous extracts and the water status of entire kiwifruits were monitored over the season (June–December) using nuclear ...magnetic resonance (NMR) methodologies. The metabolic profiling of aqueous kiwifruit extracts was investigated by means of high field NMR spectroscopy. A large number of water-soluble metabolites were assigned by means of 1D and 2D NMR experiments. The change in the metabolic profiles monitored over the season allowed the kiwifruit development to be investigated. Specific temporal trends of aminoacids, sugars, organic acids and other metabolites were observed.
The water status of kiwifruits was monitored directly on the intact fruit measuring the
T
2 spin–spin relaxation time by means of a portable unilateral NMR instrument, fully non-invasive. Again, clear trends of the relaxation time were observed during the monitoring period.
The results show that the monitoring of the metabolic profiling and the monitoring of the water status are two complementary means suitable to have a complete view of the investigated fruit.
In the Crabtree-negative Kluyveromyces lactis yeast the rag8 mutant is one of nineteen complementation groups constituting the fermentative-deficient model equivalent to the Saccharomyces cerevisiae ...respiratory petite mutants. These mutants display pleiotropic defects in membrane fatty acids and/or cell walls, osmo-sensitivity and the inability to grow under strictly anaerobic conditions (Rag− phenotype). RAG8 is an essential gene coding for the casein kinase I, an evolutionary conserved activity involved in a wide range of cellular processes coordinating morphogenesis and glycolytic flux with glucose/oxygen sensing.
A metabolomic approach was performed by NMR spectroscopy to investigate how the broad physiological roles of Rag8, taken as a model for all rag mutants, coordinate cellular responses.
Statistical analysis of metabolomic data showed a significant increase in the level of metabolites in reactions directly involved in the reoxidation of the NAD(P)H in rag8 mutant samples with respect to the wild type ones. We also observed an increased de novo synthesis of nicotinamide adenine dinucleotide. On the contrary, the production of metabolites in pathways leading to the reduction of the cofactors was reduced.
The changes in metabolite levels in rag8 showed a metabolic adaptation that is determined by the intracellular NAD(P)+/NAD(P)H redox balance state.
The inadequate glycolytic flux of the mutant leads to a reduced/asymmetric distribution of acetyl-CoA to the different cellular compartments with loss of the fatty acid dynamic respiratory/fermentative adaptive balance response.
•Metabolic changes taking place in casein kinase (CK) defective cells are depicted.•CK I depleted cells display mono/poly-unsaturated altered fatty acid content.•CK I depleted cells exert lipid homeostasis as a redox balancing adjusting mechanism.•Unequal distribution of acetyl-CoA to cellular compartments is proposed in CK I depleted cells.
The construction of efficient cell factories for the production of metabolites requires the rational improvement/engineering of the metabolism of microorganisms. The subject of this paper is directed ...towards the quantitative understanding of the respiratory/fermentative Kluyveromyces lactis yeast metabolism and its rag8 casein kinase mutant, taken as a model for all rag gene mutations.
13C NMR spectroscopy and 1,2-13C2glucose were used as metabolic stable-isotope tracer to define the metabolic profiling of a K. lactis yeast and its derivative mutants.
Rag8 showed a decrease of all 13C glutamate fractional enrichments, except for 4-13Cglutamate that was higher than wild type ones. A decrease of TCA cycle flux in rag8 mutants and a contribution of a 4-13Cketoglutarate pool not originating from mitochondria were suggested.
13C lysine enrichments confirmed the presence of two compartmentalized α-ketoglutarate (α-KG) pools participating to glutamate and lysine synthesis.
Moreover, an increased transaldolase, as compared to transketolase activity, was observed in the rag8 mutant by 13C NMR isotopomer analysis of alanine.
13C NMR-based isotopomer analysis showed the existence of different α-KG metabolic pools for glutamate and lysine biosynthesis. In the rag8 mutant, 13C labeled pentose phosphate intermediates participated in the synthesis of this compartmentalized α-KG pool.
A compartmentalization of the α-KG pools involved in lysine biosynthesis has been revealed for the first time in K. lactis. Given its great impact in metabolic engineering field, its existence should be validated/compared with other yeasts and/or fungal species.
•Metabolic pathways are characterized by 13C NMR-based isotopomer analysis.•α-Ketoglutarate (α-KG) pool involved in lysine synthesis is compartmentalized.•α-KG pool for lysine synthesis is mitochondrial-independent.•α-KG pool for lysine synthesis is PPP intermediates derived in rag8 cells.
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