Analysis of 10 nucleotides and related compounds in Litopenaeus vannamei during chilled storage by HPLC-DAD
Introduction
The Pacific white shrimp, Litopenaeus vannamei, is native to the Eastern Pacific coast through central Gulf of Mexico. It is one of the most commercially farmed species worldwide, accounting for over 70% of all farmed shrimp. Due to its high protein content, low fat content and delicate texture, L. vannamei is gaining popularity among consumers (Qian et al., 2013, Xie et al., 2012).
Nucleotides and related compounds (NRCs) are a group of compounds composed of a nitrogen-containing unit (purine, pyrimidine, nicotinamide or others) linked to a sugar and a phosphate group. Examples include adenosine triphosphate (ATP), adenosine diphosphate (ADP), adenosine monophosphate (AMP), inosinic acid (IMP), inosine (HxR), hypoxanthine (Hx), guanosine monophosphate (GMP), adenosine (Ado), adenine (Ad), xanthine (Xt), etc. NRCs are important substances in the muscles of aquatic animals such as shrimp. Close relationship has been found between the flavor and freshness of aquatic foods and NRCs. In terms of flavor, AMP can suppress bitterness and produce pleasant sweetness and saltiness, acting as a good flavor enhancer in aquatic foods. Hx can interact with certain amino acids and peptides resulting in bitterness (Tikk et al., 2006). IMP and GMP, the major flavor enhancer nucleotides in aquatic foodstuff, can be used with monosodium glutamate (MSG), producing stronger flavor enhancing effect than MSG alone. They have been widely used as new flavor enhancers in all types of food (Aristoy and Toldra, 2009, Matoba et al., 1988). As for freshness, it is commonly accepted that the ATP in fish muscle sequentially degrades after death to ADP, AMP, IMP, HxR, and Hx. Hx is subsequently oxidized to Xt (Venugopal, 2002). K value, a freshness indicator, is defined as the ratio of the sum of HxR and Hx to that of ATP, ADP, AMP, IMP, HxR, and Hx. K value has proven to be an important chemical indicator for fish freshness according to freshness studies of horse mackerel, turbot, and yellowfin tuna during chilled storage (Nejib et al., 2005, Santiago et al., 2005, Vanesa et al., 2005). K value has been widely used for the evaluation of fish quality (Kuda et al., 2007, Mendes et al., 2001). Recent studies showed that the degradation pathway of ATP in certain ocean invertebrates was different from that in fish. The ATP degradation pathway in the muscle tissues of clams and scallops was found to follow ATP→ADP→AMP→Ado→HxR→Hx (Arai and Saito, 1961, Hiltz and Dyer, 2011). There was no IMP intermediate in between AMP and HxR. In the muscle tissues of ark clam, mogai, and abalone, this process followed ATP→ADP→AMP→Ado→Ad (Arai and Saito, 1961, Iwamoto et al., 1991, Saito et al., 1958, Yoneda et al., 2002). On the other hand, two ATP degradation pathways were proposed in the muscle tissues of Dungeness crab and kuruma prawn, i.e. ATP→ADP→AMP→IMP→HxR→Hx and ATP→ADP→AMP→Ado→HxR→Hx (Groniger and Brandt, 1969, Shirai and Kikuchi, 2002). Therefore, it remains inconclusive whether K value can be used as a freshness indicator in invertebrates. Regardless, fast and accurate determination of NRCs in L. vannamei can provide preliminary identification of the ATP degradation pathway in postmortem muscles, which is of great significance for the flavor and freshness of aquatic foodstuff.
Current methods for the analysis of NRCs include enzymatic method (Angel & Fermin, 1981), ion exchange chromatography (Gao et al., 2006, Iwamoto et al., 1987), capillary electrophoresis (Kaname et al., 2005, Tsoda, 1983), reverse phase liquid chromatography (Bhatt et al., 2012, Graven et al., 2014, Isabel et al., 2001, Özogul et al., 2000, Wang et al., 2007; ), sensor method (Devi et al., 2013, Devi et al., 2011), etc. Reverse phase HPLC has the advantages of high speed and simplicity, which render it the principal method currently used in biological and food sciences (Jose, Barat, Eduardo, et al., 2008). However, for the analysis of ionic species, ion pairing is usually required. Ion pair reagents are pricy and may contaminate the column, shortening the lifetime of the column. Moreover, many methods have limitations on the type of analytes and some require long analysis time. Particularly, no fast and facile HPLC method is available for the analysis of 10 NRCs in L. vannamei. In this study, a high performance liquid chromatography coupled with diode array detection (HPLC-DAD) method for the analysis of 10 NRCs in L. vannamei was established. Validation of the method was performed and good results were obtained. The concentration change of the 10 target NRCs during chilled storage (4 °C) was analyzed.
Section snippets
Instruments and materials
Live shrimp (L. vannamei) with a body weight of 10–15 g were purchased from Luchaogang Market of Shanghai and put in an oxygenated sampling vessel, which was quickly taken back to the lab. Upon arrival, the shrimp were washed with ice water and blotted dry. They were stored at 4 ± 1 °C before further processing.
HPLC analysis was performed on a Waters Alliance 2695 HPLC system equipped with a Waters 2996 photodiode array (PDA) detector and Empower 2 software package. A Shimadzu Kubota 520
Optimization of chromatographic conditions
Separation of the 10 NRCs was optimized in terms of peak shape, degree of separation, and analysis efficiency. Optimal, column, pH and composition of the mobile phase were selected.
Conclusion
In this study, an HPLC-DAD method for the analysis of 10 NRCs in L. vannamei was developed and evaluated. The target 10 NRCs were ATP, ADP, AMP, IMP, HxR, Hx, GMP, Ado, Ad, and Xt. This new method is faster, more efficient, more sensitive, and has better separation power than current methods. It meets the requirements for precision, repeatability, linearity, limit of detection and spike recovery. This method is effective in determining NRCs and deciphering the ATP degradation pathway in L.
Acknowledgments
This work was supported in part by National “Twelfth Five-Year” Plan for Science & Technology Support (Grant No: 2012BAD29B06), Shanghai Science and Technology Key Project on Agriculture from Shanghai Municipal Agricultural Commission (Grant No: (2013)3rd-4).
References (41)
- et al.
A sensitive HPLC-based method to quantify adenine nucleotides in primary astrocyte cell cultures
Journal of Chromatography B
(2012) - et al.
A method for determination of xanthine in meat by amperometric biosensor based on silver nanoparticles/cysteine modified Au electrode
Process Biochemistry
(2013) - et al.
Determination of the big head carp myofibrillar (Aristichthys nobilis) adenosine triphosphatase activity by ion chromatography
Journal of Chromatography A
(2006) - et al.
Purine metabolite and energy charge analysis of trypanosoma brucei cells in different growth phases using an optimized ion-pair RP-HPLC/UV for the quantification of adenine and guanine pools
Experimental Parasitology
(2014) - et al.
Simple, rapid, and sensitive liquid chromatography-fluorescence method for the quantification of tranexamic acid in blood
Journal of Chromatography A
(2007) - et al.
Intracellular nucleotide and nucleotide sugar contents of cultured CHO cells determined by a fast, sensitive, and high-resolution ion-pair RPHPLC
Analytical Biochemistry
(2006) - et al.
Effects of freshness on ATP related compounds in retorted chub mackerel Scomber japonicas
Food Science and Technology
(2007) - et al.
Inhibitory effect of a quercetin-based soaking formulation and modified atmospheric packaging (MAP) on muscle degradation of Pacific white shrimp (Litopenaeus vannamei)
Food Science and Technology
(2015) Biosensors in fish production and quality control
Biosensors & Bioelectronics
(2002)- et al.
Identification of the distribution of adenosine phosphates, nucleosides and nucleobases in royal jelly
Food Science and Technology
(2015)
Dietary lysine requirement of juvenile Pacific white shrimp, Litopenaeus vannamei
Aquaculture
Acid-soluble nucleotides of cow's, goat's and sheep's milks, at different stages of lactation
Journal of Dairy Research
Changes in adenine nucleotides in the muscle of some marine invertebrates
Nature
Nucleotides and its derived compounds
Electrochemical detection of xanthine in fish meat by xanthine oxidase immobilized on carboxylated multiwalled carbon nanotubes/polyaniline composite film
Biochemical Engineering Journal
Degradation of adenine nucleotides in the muscle of dungeness crab (cancer magister) and king crab (Paralithodes camtschatica) during storage and cooking
Journal of Milk and Food Technology
Principal acid-soluble nucleotides in adductor muscle of the scallop Placopecten magellanicus and their degradation during postmortem storage in ice
Journal of the Fisheries Board of Canada
The determination and distribution of nucleotides in dairy products using HPLC and diode array detection
Food Chemistry
Effect of storage temperature on rigor-mortis and ATP degradation in plaice Paralichtithys olivaceus muscle
Journal of Food Science
Change in ATP and related breakdown compounds in the adductor muscle of itayagai scallop pecten ablicans during storage
Nippon Suisan Gakkaishi
Cited by (34)
Freshness evaluation of grouper fillets by inexpensive e-Nose and spectroscopy sensors
2024, Microchemical JournalEffect of different thermal processing methods on water-soluble taste substances of tilapia fillets
2022, Journal of Food Composition and AnalysisEffects of thermoultrasonic treatment on characteristics of micro-nano particles and flavor in Greenland halibut bone soup
2021, Ultrasonics SonochemistryCitation Excerpt :Thus, halibut bone soup was rich in volatile compounds such as ketones, aldehydes and alcohols. This is consistent with the results that volatile flavor substances such as aldehydes, ketones and alcohols are usually contained in fish products [42]. However, the response value of W5S in the TUT group was significantly increased, while the response value of W2S, W3S and W1S decreased.
Application of UHPLC-Q-TOF-MS/MS metabolomics approach to investigate the taste and nutrition changes in tilapia fillets treated with different thermal processing methods
2021, Food ChemistryCitation Excerpt :Compared to the fresh samples, the nucleotide contents in the tilapia fillet meat after thermal processing showed an overall upwards trend with certain differences between the different processing methods. Inosincacid (IMP) and AMP are the two main taste nucleotides in fish (Qiu, Chen, Xie, Qu, & Song, 2016), both of which can be decomposed by ATP. The degradation process of ATP is affected and controlled by ATPase, adenosine diphosphate (ADP) enzyme, AMP dehydrogenase, phospholipase and nucleoside enzyme.