Water-Soluble Flavor and Odor Precursors of Meat. 4. Influence of Cooking on Nucleosides and Bases of Beef Steaks and Roasts and Their Relationship to Flavor, Aroma and Juiciness

SUMMARY: Cooking resulted in significant increases in adenylic acid, total purine nucleosides and bases of 80 beef roasts of eight different cuts. It decreased the contents of inosinic acid, guanylic acid and sum of individual nucleotides (adenylic, cytidylic, uridylic, inosinic and guanylic acids) in these samples. Significant differences were also found between the various constituents of raw and cooked samples of the beef cuts.     

酿酒酵母和大肠杆菌鸟嘌呤脱氨酶基因的克隆、原核表达及活性测定

与植物体内合成路径不同,微生物体内合成咖啡碱存在一条以黄嘌呤为底物,利用鸟嘌呤脱氨酶催化鸟嘌呤生成黄嘌呤有效合成咖啡碱的新途径。为克隆鸟嘌呤脱氨酶的基因,构建可高效合成黄嘌呤的原核表达载体并对外源蛋白活性进行检测,分别以酿酒酵母和大肠杆菌为研究材料,根据GenBank中酿酒酵母和大肠杆菌中鸟嘌呤脱氨酶基因gud1和egud序列设计引物,聚合酶链式反应特异扩增其基因片段,将目的基因连接至pMAL-c5X载体,转入大肠杆菌BL21（DE3）中诱导蛋白表达,并用高效液相色谱法鉴定其目的蛋白的催化活性。结果表明重组载体pMAL-gud1、pMAL-egud均可用来合成黄嘌呤,且GUD1比EGUD合成黄嘌呤的效率更高。研究结果将进一步丰富黑茶加工技术理论,同时为体外构建高效咖啡碱生物工程菌提供理论支持。     

Water-Soluble Flavor and Odor Precursors of Meat. 3. Changes in Nucleotides, Total Nusleosides and Bases of Beef, Pork and Lamb During Heating

SUMMARY: Studies were made of the influence of heating on nucleotides and total purine nucleosides and bases of beef, pork and lamb muscle. lnosinic acid was the predominant nucleotide in all three species and it was degraded by heating. Adenylic acid increased during cooking in meat from all three species. Cytidylic, uridylic and guanylic acids were present in relatively low concentrations in meat from all three species and changed little during cooking. A rapid method for estimating total nucleotides resulted in greater variation than a specific method for measuring individual nucleotides.     

DETERMINATION OF DNA AND OF OTHER NUCLEIC ACID FRACTIONS IN SACCHAROMYCES CEREVISIAE

After extraction overnight of Saccharomyces cerevisiae (baker's yeast) with dilute alkali, to remove nucleotide pools and RNA, acidification gave an insoluble residue from which DNA guanine and adenine were extracted, in the free form, by 1M perchloric acid at room temperature, and determined by cation-exchange chromatography. Percentage DNA in the yeast was computed as 0.062 × μmol/g of guanine + adenine. Preliminary extraction of nucleotides was unnecessary, and gave low results if acid reagents (perchloric or trichloroacetic acids) were used. Several reagents were tested for their ability to extract the nucleotide pool from intact yeast without significant effect on cell RNA content. The content of guanine + adenine in pool-free yeast was taken as giving the sum of RNA + DNA purines, RNA purines being found by difference.     

Purine and pyrimidine utilisation by chicks fed nitrogen-limiting diets.

The responses of chicks to purine and pyrimidine supplementation of diets severely limiting in the dispensable amino acids have been measured in three experiments. In experiment 1, combined additions of adenine and uracil to a diet with a nitrogen (N) content of 19.84 g kg^?1 DM enhanced growth and dry matter intakes to levels similar to those attained by chicks receiving an equinitrogenous supplement of glutamic acid. Chicks in the latter group, however, had the best efficiencies of carcass N deposition. In experiment 2, growth, dry matter intakes and efficiency of food conversion of chicks fed a diet with 23.36 g N kg^?1 DM were improved markedly by combined supplements of adenine, uracil, guanine and cytosine. Equinitrogenous inclusions of guanine; guanine with thymine; uracil with thymine; or uracil with adenine were ineffective. Efficiency of carcass N deposition declined appreciably on supplementing the basal diet with purines and pyrimidines in different combinations, appreciable reductions occurring with additions of thymine; uracil with thymine and uracil with adenine. The low N efficiencies observed in the last two groups were improved by combined additions of adenine, uracil, guanine and cytosine. The highest efficiencies of N utilisation occurred in chicks fed a glutamic acid supplement. In experiment 3, the simultaneous addition of uracil, adenine and guanine was as effective in stimulating the growth performance of chicks fed a diet with 18.89 g N kg^?1 DM as a more complex mixture including cytosine. The best growth and efficiency of food conversion values were recorded in the group receiving supplementary glutamic acid.     

QUANTITATIVE ANALYSIS OF PURINE NUCLEOSIDES AND FREE BASES IN WORT AND BEER

Low molecular mass compounds containing purine or indole groups exhibit adsorption-based retention characteristics when chromatographed on high performance size exclusion chromatography supports (Superose 12 and Superdex 75). Exploitation of this phenomenon enabled the development of a method for the quantitative analysis of purine nucleosides and free bases in wort and beer. Using this technique, a survey of 18 commercial beers showed the following range of variation of concentrations of these compounds: guanosine + deoxyguanosine 19–110 mg/L, adenosine + deoxyadenosine 3–43 mg/L, xanthine 1–41 mg/L, guanine < 1–11 mg/L and adenine < 1–7 mg/L. Analysis of wort and beer from a commercial brewery demonstrated a decline in both adenine and guanine and an increase in xanthine during the course of a typical fermentation.     

Determination of uric acid precursors in dried yeast and other forms of single-cell protein

Total purine content, expressed as μmol/g, or as percentage RNA of a conventionally defined composition, is suggested as a convenient index of the amount of uric acid precursors in SCP. A simple but reproducible analytical procedure was based on the extraction of RNA and nucleotides from dried microbial cells with 1 N-perchloric acid at room temperature, followed by hydrolysis to free bases by heating the cell-free extract for 1 h at 100°C. After precipitation as the silver complexes, washing, and regeneration with HCI, purines (mainly adenine + guanine) were determined by spectrophotometry at 252 nm. If absorbance at 264.5 nm was measured as well, adenine and guanine could be determined separately. Results with dried yeast and with fungal mycelium were confirmed by cation-exchange chromatography, when purines were eluted from the column by citrate buffers (pH 3.0–5.0).     

Growth of Lactobacillus bulgaricus in milk. 2. Characteristics of purine nucleotides, pyrimidine nucleotides, and nucleic acid synthesis

Lactobacillus bulgaricus incorporated exogenous guanine only into guanine nucleotides and not into adenine nucleotides. Lactobacillus bulgaricus lacks the ability to interconvert adenine nucleotides and guanine nucleotides. Biosynthesis of pyrimidine nucleotides from orotic acid was depressed by addition of guanosine monophosphate or adenosine monophosphate. Some step in pyrimidine biosynthesis from orotic acid may be negatively regulated by the intracellular amount of purine nucleotides. Lactobacillus bulgaricus could not grow in milk from which orotate was removed. This indicates that the activity of de novo pyrimidine synthesis is very low in this organism. When the intracellular amount of purine or pyrimidine base was limited, ribonucleic acid synthesis was markedly depressed, resulting in cell elongation, whereas deoxyribonucleic acid synthesis was not so much affected.     

Technical note: A new high-performance liquid chromatography purine assay for quantifying microbial flow

An HPLC method was developed to quantify the purines adenine and guanine and their metabolites xanthine and hypoxanthine in hydrolysates of isolated bacteria and omasal digesta and to assess the effect of using either purines only or purines plus metabolites as microbial markers for estimating microbial flow from the rumen. Individual purines and their metabolites were completely resolved on a C18 column using gradient elution with 2 mobile phases. Intraassay coefficient of variation ranged from 0.6 to 3.1%. Hydrolytic recovery of the 4 purine bases from their corresponding nucleosides averaged 101% (control), 103% (when added to bacterial isolates), and 104% (when added to omasal digesta). Mean concentrations of adenine, guanine, xanthine, and hypoxanthine were, respectively, 53, 58, 2.8, and 3.5 μmol/g of dry matter in omasal bacteria and 10, 12, 7.5, and 7.5 μmol/g of dry matter in omasal digesta, indicating that xanthine plus hypoxanthine represented 5% of total purines in bacterial hydrolysates but 41% of total purines in digesta hydrolysates. A significant negative relationship (R² = 0.53) between the sum of adenine and guanine and the sum of xanthine and hypoxanthine in digesta samples (but not bacterial isolates) indicated that 89% of the adenine and guanine originally present in ruminal microbes were recovered as xanthine and hypoxanthine. These results suggested that, when total purines are used as the microbial marker, both purines and their metabolites should be quantified and used to compute microbial nonammonia N and organic matter flows.     

Nucleic acids of species of Lactobacillus

Nucleic acid composition of 17 cultures representing the type or neotype strains of 15 named Lactobacillus species was studied. Nucleic acid characterization of these isolates was accompanied by a comparative study of conventional phenotypic reactions. The overall guanine plus cytosine mean deoxyribonucleic acid base composition ranged from 33 to 50% and genome sizes varied between 700 and 1500 X 10(6) daltons. Sporolactobacillus inulinus contained 2500 X 10(6) daltons of chromosomal deoxyribonucleic acid and was therefore similar in size to members of the genus Bacillus. Deoxyribonucleic acid hybridization at a temperature 15 degrees C below the thermal melting temperature confirmed the extreme molecular heterogeneity of many species. The genus can be divided into three major groups, each containing four or more species based on a combination of nucleic acid characteristics and conventional phenotypic reactions. Group I (33 to 39% guanine plus cytosine) contains Lactobacillus jensenii, Lactobacillus helveticus, Lactobacillus helveticus nov. ssp. jugurt, Lactobacillus acidophilus, Lactobacillus salivarius, and Lactobacillus sanfrancisco. Group II (42 to 48% guanine plus cytosine contains Lactobacillus plantarum, Lactobacillus rhamnosus, Lactobacillus casei, and Lactobacillus brevis, and Group III (48 to 50% guanine plus cytosine) contains Lactobacillus delbrueckii, Lactobacillus delbrueckii nov. ssp. bulgaricus, Lactobacillus delbrueckii nov. ssp. lactis, Lactobacillus delbrueckii nov. ssp. leichmanii, Lactobacillus fermentum, and Lactobacillus fermentum nov. ssp. cellobiosis.     

Nucleic Acid, Purine and Proximate Analyses of Mechanically Separated Beef and Veal

Deoxyribonucleic acid (DNA), ribonucleic acid (RNA), purine (adenine, guanine, hypoxanthine and xanthine) and proximate analyses of mechanically separated (MS) beef and veal were conducted to verify and evaluate changes in nucleic acid content which may result from mechanical separation. DNA and total nucleic acid levels were higher in both MS beef and veal whereas RNA levels were higher only in MS beef compared to hand deboned (HD) counterparts. Adenine, guanine and xanthine levels were higher in MS beef and veal, and hypoxanthine levels were lower compared to HD counterparts. Total purine content of MS beef did not differ from HD beef, whereas the purine content of MS veal was slightly higher than HD veal. It would appear that the addition of MS products to the diet would not significantly alter total purine consumption and hence should pose no risk to hyperuricemic individuals.     

挤压喷雾生产香菇粉工艺优化及产品理化性质

利用挤压喷雾技术生产香菇粉调味料。在单因素试验基础上进行正交试验优化挤压条件,通过对比挤压前后香菇粉的理化性质变化,研究香菇粉的使用条件、范围以及营养成分的变化规律。结果表明:最佳工艺为香菇原料初始水分含量50%、挤压温度150℃、模孔直径3 mm,此条件下制得的香菇粉香味物质I+G(I表示肌苷酸,G表示鸟苷酸)含量最高。挤压喷雾处理后的香菇粉脂肪含量下降,热性质稳定,表面更加致密光滑,粉状性质变佳且活性基团没有明显变化。挤压喷雾处理的香菇粉,在食品生产中可作为调味料和功能性添加剂使用。     

Effects of RNA, purines and pyrimidines on the development of Tribolium confusum (Duval)

Growth period and survival of Tribolium confusum were studied on a chemically defined diet alone or supplemented with RNA; guanine, adenine, cytosine, and uracil were also added to the basic diet either singly or in all possible combinations. The effect of supplementing a processed food (dehydrated tapioca pudding) with various levels of RNA on the growth and survival of the beetle was also studied.

Diets supplemented with RNA or a mixture of guanine + cytosine or all the four nucleotides were equally effective in stimulating growth and increasing survival of the beetle; guanine + adenine also improved the diet but not as well as the other three. In general, the purines guanine and adenine stimulated the growth and the pyrimidine cytosine acted as synergist while uracil had slight retardatory effect. The tapioca pudding, which is inadequate for supporting growth of the beetle was also improved by the addition of RNA.     

基于D152树脂吸附蛋白质结合SERS测定鱼肉中的鸟嘌呤含量

基于大孔树脂除杂与表面增强拉曼光谱的灵敏实时性,建立一种鱼肉中鸟嘌呤含量的快速检测方法。以D152树脂为填料,柱内直径1 cm,柱高20 cm,蛋白吸附率在83%～92%之间,鸟嘌呤滤除率大于95%。经单因素试验筛选和正交试验优化得出,上样流速为3 mL/min,洗脱液pH值为4.0以及洗脱流速为0.5 mL/min。基于银包金纳米颗粒（silver-coated gold nanoparticles,Au@Ag NPs）为表面增强基底,鸟嘌呤质量浓度在0.001～100 mg/L范围,鸟嘌呤质量浓度与拉曼强度呈线性关系,R2为0.969 9,检出限为0.1 mg/L。整个检测过程只需10 min,且无需复杂的样品处理。结果表明,D152树脂作为填料的层析柱能有效去除鱼肉中蛋白质等杂质的干扰,以Au@Ag NPs为基底的表面增强拉曼光谱技术的方法能够灵敏、快速的检测鱼肉中痕量嘌呤,检出限低于现有的高效液相色谱法,为开发设计水产品中嘌呤的快速检测方法提供研究基础。     

Oxidation of the antiviral drug acyclovir and its biodegradation product carboxy-acyclovir with ozone: kinetics and identification of oxidation products

The oxidation of the antiviral drug acyclovir (ACV) and its main biotransformation product carboxy-acyclovir (carboxy-ACV) by ozone was investigated. Both compounds have recently been detected in surface water, and carboxy-ACV has also been detected in drinking water. The experiments revealed a strong pH dependence of the oxidation of ACV and carboxy-ACV with reaction rate constants increasing by 4 orders of magnitude between the protonated, positively charged form (k(ox,PH(+)), ～2.5 × 10(2) M(-1) s(-1)) and the deprotonated, negatively charged form (k(ox,P(-)), 3.4 × 10(6) M(-1) s(-1)). At pH 8 a single oxidation product was formed which was identified via LC-LTQ-Orbitrap MS and NMR as N-(4-carbamoyl-2-imino-5-oxoimidazolidin)formamido-N-methoxyacetic acid (COFA). Using Vibrio fischeri , an acute bacterial toxicity was found for COFA while carboxy-ACV revealed no toxic effects. Ozonation experiments with guanine and guanosine at pH 8 led to the formation of the respective 2-imino-5-oxoimidazolidines, confirming that guanine derivatives such as carboxy-ACV are undergoing the same reactions during ozonation. Furthermore, COFA was detected in finished drinking water of a German waterworks after ozonation and subsequent activated carbon treatment.     

Electrochemical evaluation of total antioxidant capacity of beverages using a purine-biosensor

In this paper, it was evaluated the total antioxidant capacity (TAC) of beverages using an electrochemical biosensor. The biosensor consisted on the purine base (guanine or adenine) electro-immobilization on a glassy carbon electrode surface (GCE). Purine base damage was induced by the hydroxyl radical generated by Fenton-type reaction. Five antioxidants were applied to counteract the deleterious effects of the hydroxyl radical. The antioxidants used were ascorbic acid, gallic acid, caffeic acid, coumaric acid and resveratrol. These antioxidants have the ability to scavenger the hydroxyl radical and protect the guanine and adenine immobilized on the GCE surface. The interaction carried out between the purine-base immobilized and the free radical in the absence and presence of antioxidants was evaluated by means of changes in the guanine and adenine anodic peak obtained by square wave voltammetry (SWV). The results demonstrated that the purine-biosensors are suitable for rapid assessment of TAC in beverages.     

Purine Content of Raw and Roasted Chicken Broiler Meat

Adenine, guanine and hypoxanthine were determined in raw and roasted broiler parts. The levels of adenine and guanine increased slightly when the meat was cooked. These increases were attributed to moisture and fat losses by the tissues during roasting. The level of hypoxanthine remained constant or decreased in the tissues because some of the purine was removed with the cooking juices. The cooking juices were found to contain high levels of hypoxanthine and only trace amounts of adenine and guanine.     

食盐腌制对油炸罗非鱼风味的影响

为探究食盐腌制对油炸罗非鱼风味的影响,本研究以蛋白质、脂肪酸、游离氨基酸以及呈味核苷酸含量为考察指标,通过感官评定结合顶空固相微萃取-气质联用(HS-SPME-GC-MS)方法分析食盐腌制对油炸罗非鱼可挥发性风味物质以及风味组分间的影响。结果表明,腌制前后的油炸罗非鱼风味变化明显,感官评价结果显示经过食盐腌制处理,油炸罗非鱼油脂味、肉香味增强,土腥味减弱;HS-SPME-GC-MS检测结果显示,食盐腌制过后油炸罗非鱼的可挥发性气味物质中1-辛烯-3-醇相对含量降至4.17%,ROAV值为5.70、(E,E)-2,4-癸二烯醛含量升为5.12%,ROAV值为100,肉香味和油脂味明显增强,土腥味明显降低;食盐腌制处理后油炸罗非鱼蛋白质与脂肪的含量显著降低(P<0.05),游离氨基酸与脂肪酸的消耗量增加,分别为38.94 mg/100 g和2.37 mg/g,且呈味氨基酸的占比与不饱和脂肪酸的含量发生显著降低(P<0.05),提供鲜味的肌苷酸(Inosinic acid,IMP)和鸟苷酸(Guanylic acid,GMP)含量显著(P<0.05)降低。实验证明食盐腌制对油炸罗非鱼的风味物质及组分影响显著(P<0.05),显著降低蛋白质和脂肪的含量,降解形成的挥发性物质显著提高,提高罗非鱼的油脂香、鲜味和肉香味,降低其土腥味和青草味,明显改善罗非鱼的品质。     

Effect of Stewing on Purine Content of Broiler Tissues

The purine content of raw and stewed broiler tissues was evaluated. Stewed breast had slightly more adenine and stewed thigh had slightly more guanine than their raw counterparts. The analyses indicated that all of the purines were considerably higher in cooked skin than in raw skin. This obsrevation seems to be the result of extraction of other nitrogenous components (possibly collagen) from the skin. Hypoxanthine was the only purine which was studied that was extracted from the tissues in large amounts.