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Identifier 000445882
Title Μεταβολομική ανάλυση τυροκομικών προϊόντων με τη φασματοσκοπία NMR
Alternative Title Metabolomic analysis of dairy products by NMR spectroscopy
Author Ράλλη, Ευαγγελία
Thesis advisor Σπύρος, Απόστολος
Reviewer Περγαντής, Σπυρίδων
Χανιωτάκης, Νικόλαος
Δημάδης, Κωνσταντίνος
Κατερινόπουλος, Χαράλαμπος
Παυλίδης, Ιωάννης
Stapf, Siegfried
Abstract This dissertation deals with the study of PDO Cretan graviera cheese and several white PDO Cretan cheeses produced solely on the island, using high field and low field (LF) NMR spectroscopy and metabolomics for the development of multivariate statistical analysis models based on the NMR experimental data. The research results are presented in three main parts; the first deals with the study of the maturation process of Cretan graviera cheese, the second part examines the effect of milk composition and state (liquid or powder) on the chemical composition of graviera, while the third part deals with the analysis of several sour white PDO cheeses produced in Crete. Three (3) Cretan graviera samples produced in different dairy producer sites on the island of Crete were studied in the first part of the thesis, were followed during maturation, and sampled regularly for analysis. Polar and apolar extracts of the graviera cheese samples were studied by high resolution NMR spectroscopy, and the integration of the NMR spectra was used for the quantitative determination of the chemical composition (metabolite) profile of the graviera samples as a function of the maturation time. The analytical results were used to calculate the rate of proteolysis (aminoacid production, µmol/d), lipolysis (1,2- Diglycerides/Triglycerides ratio) and tyramine (µmol/d) production for each of the three different graviera samples studied under maturation. Furthermore, multivariate statistical analysis PLS models based on the NMR data were developed in order to elucidate important metabolites correlated with maturation, and to identify possible differences in the maturation procedure in different dairy production facilities. These PLS models were applied to the compositional analysis of a large set of commercial graviera samples in an effort to assess the potential of NMR metabolomics in the determination of the maturation time of unknown samples. One of the graviera samples, obtained from Kargakis dairy farm, was used to monitor the maturation procedure by performing low field NMR experiments in the same samples and time frame as the high resolution NMR analysis. The parameters measured in these experiments were the T1 and T2 relaxation times and diffusion coefficients D of the water and lipid protons as a function of maturation time, and the proton relaxation rate as a function of magnetic field strength obtained from Fast Field Cycling (FFC) NMR experiments. The proton T1 and T2 relaxation times of water and lipid fat in the graviera were found to be similar in value to other yellow hard and semi-hard cheeses reported in the literature, and did not change much with maturation time. The diffusion coefficient of fat, Df, displayed a small xx increase as a function of maturation time, while the diffusion coefficient of water Dw decreased with maturation time. A comparison of the PLS maturation models developed showed that the multivariate model incorporating both low and high field NMR data demonstrated better validation and statistical significance characteristics compared to models using either just the low or only the high field NMR data. The second part of the thesis involved the laboratory production of five different “model” sheep cheeses based on the cheese-making protocol used for Cretan graviera, in order to study the effect of milk quality (sheep milk – commercial sheep milk powder) on the composition and quality attributes of the graviera samples. The milk used was either 100% sheep milk obtained from a dairy farm, or a mixture of such milk with increasing amounts of commercial sheep milk powder (25-100%). After the production of the model graviera cheeses they were left to mature for 90 days, as per PDO directions, during which they were sampled for extraction and NMR spectral analysis. This series of experiments showed that all cheeses prepared with some amount of milk powder contained 1-monoglycerides (1-MG) the quantity of which was increasing with % milk powder used and maturation time and were low in amino acids and lactate. NMR analysis of the commercial milk powder identified 1-MG as preexisting components of the lipid matrix which were transferred to the graviera cheese during the cheese-making procedure. Retroanalysis of NMR spectra of a large number of commercial graviera samples revealed the presence of 1-MG in a small number of samples, indicating either the (illegal in case of PDO products) use of milk powder, or the suboptimal practices in milk treatment before cheese-making. The third part of the thesis deals with the qualitative and quantitative high resolution NMR analysis of different types of “sour” Cretan PDO white cheeses, namely Xynomizithra Kritis, Xygalo Sitias, Piktogalo Chanion and xynogalo (non PDO). The water extract of these cheeses was found to contain sugars, organic acids, amino acids, alcohols and nitrogen compounds, all of which were successfully quantified using NMR spectroscopy, and the same was done for the apolar lipid extract of the cheeses. It was found that the polar metabolite profile of the cheeses showed the greater variability, while the lipid fraction only showed minor differences. The obtained 1H NMR metabolite profiles of the white Cretan cheeses were used to construct multivariate statistical analysis models that classified the type and production method of the different white “sour” cheeses. It was also shown that PDO Xinomizithra Kritis could not be differentiated according to geographical origin within the general area of production on the island of Crete.
Language Greek
Subject Amino acids
Cretan PDO cheeses
Low field NMR
Relaxation time
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Issue date 2022-04-12
Collection   School/Department--School of Sciences and Engineering--Department of Chemistry--Doctoral theses
  Type of Work--Doctoral theses
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