Thermal Degradation of Flavor Enhancers, Inosine 5'-monophosphate, and Guanosine 5'-monophosphate in Aqueous Solution

The mechanism of thermal degradation of inosine 5′-monophosphate (IMP) and guanosine (GMP) was investigated kinetically in aqueous solution as a function of pH and temperature. The degradation of IMP and GMP followed first order kinetics. The rate constants were considerably affected by pH and temperature. The half-life times at 100°C were: IMP, 8.7 hr (pH 4.0), 13.1 hr (pH 7.0), 46.2 hr (pH 9.0); GMP, 6.4 hr (pH 4.0), 8.2 hr (pH 7.0), 38.5 hr (pH 9.0). These times were shortened to about one-third by raising the temperature 10°C. The predominant degradation products were nucleosides and phosphoric acid, indicating that the main reaction of the thermal degradation was the hydrolysis of the phosphoric ester bond in the nucleotides.     

Decomposition of the Flavor Enhancers, Monosodium Glutamate, lnosine-5'-Monophosphate and Guanosine-5'-Monophosphate during Canning

Monosodium glutamate (MSG) was found to be very stable to canning conditions. Both flavor nucleotides inosine-5'-monophosphate (IMP) and guanosine-5'-monophosphate (GMP) were lost through hydrolysis, up to 50% or more, depending on the canning conditions. IMP was more stable than GMP with increased hydrolysis for both compounds occurring at lower pH and longer canning time. The loss of IMP and GMP was shown to go via phosphate hydrolysis to the corresponding nucleosides, inosine and guanosine, followed by base hydrolysis of the nucleosides to hypoxanthine and guanine.     

Effect of Adenosine-Nucleotides and Their Derivatives on the Denaturation of Myofibrillar Proteins in vitro during Frozen Storage at −20°C

To investigate the effect of adenosine-nucleotides and their derivatives on the denaturation of myofibrillar proteins, 235 nmoles/mL adenosine-5′-diphosphate (ADP), adenosine-5′-monophosphate (AMP), inosine-5′-monophosphate (IMP), inosine (HxR), or hypoxantine (Hx), was added to 3 mg/mL actomyosin (AM) solution suspended in 0.10M KC1 solution and stored at ?20°C for 12 wk. The AM was extracted from milktish dorsal muscle. Protein denaturation was evaluated by measuring solubility, Ca-ATPase and Mg(EGTA)-ATPase activity of AM, by analyzing changes in electrophoretic profiles and transmission electron microscopy. Inosine and hypoxanthine accelerated protein denaturation compared to control samples. Infrared spectrum analyses indicated that negatively charged groups of these nucleotides interacted with amino or imino groups on AM after addition. ADP, AMP, and IMP had a protective effect on denaturation of AM during frozen storage at ?20°C.     

Concentration of Umami Compounds in Pork Meat and Cooking Juice with Different Cooking Times and Temperatures

This study examined the concentrations of umami compounds in pork loins cooked at 3 different temperatures and 3 different lengths of cooking times. The pork loins were cooked with the sous vide technique. The free amino acids (FAAs), glutamic acid and aspartic acid; the 5′‐nucleotides, inosine‐5′‐monophosphate (IMP) and adenosine‐5′‐monophosphate (AMP); and corresponding nucleoside inosine of the cooked meat and its released juice were determined by high‐performance liquid chromatography. Under the experimental conditions used, the cooking temperature played a more important role than the cooking time in the concentration of the analyzed compounds. The amino acid concentrations in the meat did not remain constant under these experimental conditions. The most notable effect observed was that of the cooking temperature and the higher amino acid concentrations in the released juice of meat cooked at 80 °C compared with 60 and 70 °C. This is most likely due to the heat induced hydrolysis of proteins and peptides releasing water soluble FAAs from the meat into the cooking juice. In this experiment, the cooking time and temperature had no influence on the IMP concentrations observed. However, the AMP concentrations increased with the increasing temperature and time. This suggests that the choice of time and temperature in sous vide cooking affects the nucleotide concentration of pork meat. The Sous vide technique proved to be a good technique to preserve the cooking juice and the results presented here show that cooking juice is rich in umami compounds, which can be used to provide a savory or brothy taste.     

Sensory Changes in Umami Taste of Inosine 5'-Monophosphate Solution after Heating

Discrimination in umami taste of inosine 5′-monophosphate (IMP) solution caused by thermal degradation was investigated by sensory evaluation. The difference threshold of umami taste of 0.005% IMP solution in the presence of 0.05% monosodium glutamate (MSG) was 0.002%. The difference threshold of a 0.005% IMP solution decreased by about one half when heated at 95°C for 15 h. Inosine, one of the main products of the thermal degradation of IMP, had a bitter taste. The detection threshold of inosine varied widely among panelists. Heating a 0.005% IMP solution at 95°C for 15 h formed inosine at about one tenth of its lowest detection threshold.     

NUCLEOTIDES IN BEER

Nucleotidic material was isolated by anion exchange chromatography of beer on a Dowex 1 × 8 (formate) column. Ultraviolet spectroscopy and retention volume data suggested the presence of cytidine monophosphate (CMP) [max. 5 mg/litre], in one of the fractions. No other mononucleotides were detected. Chromatography of charcoal extracts of beer on DEAE Sephadex A25 failed to yield a fraction containing mononucleotides. Triangular taste tests indicated that addition of adenosine-5′-monophosphate (AMP) and guanosine-5′-monophosphate (GMP) to beer (30 mg/litre and 25 mg/litre respectively) did not cause significant flavour changes.     

Hypoxanthine Ratio Determination in Fish Extract Using Capillary Electrophoresis and Immobilized Enzymes

Fish freshness was assessed using capillary electrophoresis and an immobilized enzyme procedure to monitor degradation of inosine-5′-monophosphate (IMP), inosine (HxR) and hypoxanthine (Hx). The enzymatic method used an amperometric probe at + 0.7 V (platinum vs silver/silver chloride) with immobilized xanthine oxidase, catalase, nucleoside phosphorylase, and nucleotidase for converting Hx, HxR or IMP to uric acid. Capillary electrophoresis resolved IMP, inosine and Hx by migration rates resulting from an applied electric field (416 V/cm, 50 μA). Components were detected at 250 nm. The H ratio of Hx/[IMP + HxR + Hx] and simplified K value of [HxR + Hx]/ [IMP + HxR + Hx] were determined in cod, salmon and trout stored on ice (0-4°C) and at 20°C. The two procedures agreed and for all species H ratio and K values increased with storage time.     

Flavor Compounds of Beef Broth as Affected by Cooking Temperature

Of various cooking temperatures studied, broth from beef obtained on heating at 85°C for 60 min had the most intense brothy flavor. The components of the fractions with brothy flavor were isolated by gel filtration chromatography and characterized. A significant correlation between cooking temperature of the broth and the concentration of certain compounds such as free amino acids (FAA) (r = 0.88, p<0.005), camosine (r = 0.83, p <0.005) and inosine 5′-monophosphate (IMP) (r = 0.94, p<0.005) was detected. Because these results correspond to those of sensory evaluation, it was concluded that a wide mixture of FAA, peptides of low molecular weight (< 300 daltons) and IMP played an important role in the flavor intensity of beef broth.     

Effect of gamma-irradiation on flavour 5′-nucleotides,tyrosine, and phenylalanine in mushrooms (Agaricus bisporus)

The impact of gamma-irradiation on 5′-nucleotides and on the free amino acids tyrosine and phenylalanine in fresh mushrooms was studied. After irradiation the samples were freeze-dried to avoid enzyme induced chemical changes. Three 5′-nucleotides could be detected using HPLC–UV and LC–ESI-MS: adenosine 5′-monophosphate (AMP), guanosine 5′-monophosphate (GMP) and guanosine 5′-diphosphate (GDP). Irradiation significantly reduced (p = 0.05) the GDP concentration (22%). AMP showed a marked reduction (46%) only at 5 kGy. GMP, tyrosine, and phenylalanine were not affected by gamma-irradiation.     

Nonvolatile taste compounds in cooked Chinese Nanjing duck meat following postproduction heat treatment

Abstract: Taste‐active compounds, including free amino acids, succinic acid and 5′‐nucleotides, and chemical components including moisture, pH, protein, crude fat, and sodium chloride were evaluated in cooked and packaged Chinese Nanjing ducks following heat treatments including control, 99 °C for 40 min, 108 °C for 20 min, 92 °C microwave followed by water heating, 95 °C for 30 min, 121 °C for 25 min. Heat treatment decreased (P < 0.05) the content of Alamine and moisture and reduced the pH value in muscle, but increased (P < 0.05) the protein and 5′‐nucleotides content. The 99 °C group had a significantly lower crude fat content compared with other groups (P < 0.05). The succinic acid content in the control group was significantly higher than in the 121 °C group (P < 0.05). Groups treated at higher temperatures (108 °C, 121 °C, and microwave) had similar equivalent umami concentrations and 5′‐nucleotides, free amino acids content, as well as the derived bitter and sweet taste components compared with the groups treated at lower temperatures. It can be speculated that these differences account for the enhanced flavor of the meat in the 99 °C, 108 °C, 121 °C, and microwave groups compared with the untreated control. Therefore, heat treatment at lower temperature after packaging may prolong product shelf life without any detrimental effects on taste. The results of this study indicate that it is important to use lower temperatures in this type of food processing. However, it may be possible to modify the primary processing steps to improve the content of umami‐like taste compounds such as 5′‐nucleotides. Practical Application: Heat treatment of packaged products is an effective method for eradication of microbes, therefore increasing the shelf‐life. However, such treatment can result in major changes in the sensory perception of meat products, particularly the formation of off‐flavors. This study investigated changes in taste‐active compounds in duck meat following a number of types of heat treatment.     

Analysis of ATP and Its Breakdown Products in Beef by Reversed-Phase HPLC

Analytical conditions for the quantitative determination of adenosine 5′-triphosphate(ATP), adenosine 5′-diphosphate(ADP), inosine 5′-monophosphate(IMP), adenosine 5′-monophosphate(AMP), hypoxanthine(Hyp), xanthine(Xan), inosine(Ino) and adenosine(Ado) in meat extracts by reversed-phase high-performance liquid chromatography (HPLC) were examined. A commercial ODS column with a 5-μm particle diameter was used, and expeimental parameters affecting the separation were discussed. Peaks in chromatograms of meat extracts were identified by retention time, co-injection with standards, absorbance ratios and the enzymatic peak shift method. The procedure proposed was adaptable as an indication of freshness of meat.     

Reversed-phase high-performance liquid chromatographic determination of inosine 5′-monophosphate dephosphorylation in turbot muscle during storage in different chilling and freezing conditions

Reversed-phase high-performance liquid chromatography was used to study post-mortem variation in dephosphorylation of inosine 5′-monophosphate and accumulation of its breakdown products in turbot (Scophthalmus maximus) muscle during storage at low temperatures. The relative concentrations of these compounds can be used as an index of freshness. Nucleotide decomposition and nucleoside and base accumulation are influenced by the following storage conditions: chilling at 4°Cfor 1 to 15 days; freezing for 15 days of pre- or post-rigor muscle at various temperatures between -10 and -196°C; and quick or slow thawing.     

Simultaneous determination of flavor enhancers in stock cube samples by using spectrophotometric data and multivariate calibration

Spectrophotometric data followed by a suitable treatment of chemometric analysis were used for the simultaneous determination of monosodium glutamate (MSG), guanosine 5′-monophosphate (GMP) and inosine 5′-monophosphate (IMP) in stock cube samples, without any previous extraction step. By this way, the overlapping of the absorption spectra was resolved using a PLS-1 model. The concentration for experimental calibration matrix were varied between 5.03–34.2 μg mL⁻¹ for IMP and GMP, and 448–1399 μg mL⁻¹ for MSG. The relative errors of prediction (REPCV %) were 1.8, 2.8 and 3.1 for IMP, GMP and MSG, respectively.     

Acclimation Temperature affects Activities of 5'-Nucleotidase and Acid Phosphatase and Lipid and Fatty Acid Composition in Carp Muscle Microsomes

The Km values of 5′-nucleotidase against IMP markedly increased below 6°C in the case of 32°C-acclimated carp muscle microsomes. This caused a break point at 6°C in the Arrhenius plot of 5′-nucleotidase activity against IMP at a low concentration. The discontinuity of temperature dependency was not found when a surfactant had been introduced. Muscle microsomes of carp acclimated at 32°C contained much more cholesterol and saturated fatty acid than those of 10°C-acclimated carp. Therefore, the difference of lipid composition seemed to be closely related to the change in kinetics of microsomal 5′-nucleotidase.     

A Simple Method for Following Postmortem ATP Depletion in Lamb Muscle

Relationship between adenosine 5′-triphosphate (ATP) concentration and R-value, defined as the absorption ratio of the degradation derivatives inosine 5′-monophosphate (IMP), inosine and hypoxanthine (E₂₅₀) to adenosine nucleotides (E₂₆₀), was studied in postmortem lamb Longissimus dorsi, until onset of rigor mortis, at different temperatures of muscle storage and lamb carcass weight. Results showed a high overall correlation coefficient (r =?0.960), nondependent of storage temperature or carcass weight. Thus, lamb postmortem ATP concentrations can be indirectly calculated through a simple, rapid, economic and reliable method such as R-value measurements.     

Effect of cooking conditions on the flavour compounds and composition of shrimp (Parapenaeus longirostris) broth

 To achieve a shrimp broth with the best flavour, the whole shrimp must be cooked at 85°C for 30 min in a 0.5% NaCl solution in proportions 1 : 2 (w : v). Quantitatively, the main components of the broth were nitrogen substances, the most abundant of these being peptides of molecular weight less than 600 Da. Levels of ATP metabolites were also determined [the more abundant compounds were inosine, guanosine 5′-monophosphate (GMP) and inosine 5′-monophosphate (IMP)], as were free sugars (glucose, fructose and ribose) and fat content. The free amino acid composition was also determined. A significant correlation (P <0.0001) between cooking temperature and different nitrogen fractions was observed. Received: 22 September 1997 / Revised version: 13 November 1997     

Enzymatic Production of Ribonucleotides from Autolysates of Kluyveromyces marxianus Grown on Whey

After 20h fermentation of medium containing 5% (w/v) dehydrated whey, at 30°C, pH 4.5, yeast cells were harvested, diluted in 0.1M KH₂PO₄, and autolyzed at different pHs (6.5–7.5) and temperatures (45–55°C). Phosphodiesterase (0.2–1.0% w/v, 65°C, pH 6.5, 6h) and adenyl deaminase (0.5-1.0% w/v, 60°C, pH 5.5, 4h) were added to the autolysates. After heat treatment (100°C, 15 min), samples were analyzed by RP-HPLC and LC/MS. Production of 5′-ribonucleotides was maximized at 50°C, pH 6.5. Yields of 5′-AMP (800 μg/g of biomass) and 5′-GMP (2000 μg/g) increased considerably after addition of 1.0% phosphodiesterase. 5′-IMP increased only after addition of 1.0% adenyl deaminase.     

Effect of Phytate and Other Myo-inositol Phosphate Esters on α-Amylase Digestion of Starch

The effects of phytate and other myo-inositol phosphate esters (containing one or more phosphate groups) on α-amylase digestion of soluble potato starch were evaluated by an in vitro procedure. Human salivary or Bacillus subtilisα-amylase was treated with either 2 mM or 5 mM phytate, myo-inositol-2-monophosphate (l–2-MP), or phytate hydrolyzed to various degrees, and then incubated at 37°C with the starch at pH 4.15 or 6.90. Starch digestion varied with degree of phosphorylation of inositol, inositol phosphate ester concentration, pH and enzyme source. At pH 4.15, phytate (2 mM) and I-2-MP (2 mM) reduced starch digestion by salivary α-amylase to 8.5 and 78.3%, respectively, of the control.     

Determination of Inosine-5-Monophosphate in Fish Tissue with an Enzyme Sensor

An enzyme sensor for IMP consisted of immobilized enzymes and an oxygen electrode. A nucleotidase (E.C. 3.1.3.5.), nucleoside phosphorylase (E.C. 2.4.2.1.) and xanthine oxidase (E.C. 1.2.3.2.) were immobilized on a membrane prepared from cellulose triacetate, 1,8-diamino-4-aminomethyloctane and glutaraldehyde. Optimum conditions for IMP determination were pH 7.8, 30°C, a flow rate of 1 ml min⁻¹ and sample volume of 20 μl. A calibration curve for IMP was linear up to 15 mM. This sensor could be applied to the determination of IMP in meats of sea bass, mackerel, yellowfish and saurel. Good correlative results were observed between the values obtained by the sensor and the conventional method. The sensor was stable for more than 15 days at 5°C and 100 assays.     

Characterization of Quality Deterioration in Yellowfin Tuna

The spoilage pattern of thawed yellowfin tuna carcasses was examined. After five days storage at 10° and 18°C, bacterial counts were low (< 10³) although visible softening of the flesh was apparent. Direct measurement of ethanol as an indicator of pre-process spoilage also failed to reflect differences among carcasses which were aged for up to five days. Analysis of nucleotides by high performance liquid chro-matography suggested that the ratio of inosine monophosphate (IMP) to other nucleotides may be a valuable indication of spoilage time where the IMP ratio decreased linearly with time after thawing. Average recoveries of adenosine monophosphate, inosine monophosphate, inosine and hypoxanthine were 50%, 75%, 64% and 92%, respectively, from spiked tuna which had been cooked at 122°C for 67 min.