Development of a DNA Array for the Simultaneous Detection and Identification of Sugar Thick Juice Bacterial Contaminants

Despite the use of generally accepted good storage practices, sugar thick juice degradation caused by microbiological contamination occasionally occurs, causing considerable financial loss. In this study, a DNA array was developed for simultaneous detection and identification of the most prominent microflora present during thick juice storage, which may cause degradation of the thick juice. Specific oligonucleotides were developed for several bacterial taxa, including the genera Bacillus, Kocuria, Staphylococcus and Tetragenococcus and the species Aerococcus viridans, Leuconostoc mesenteroides and Tetragenococcus halophilus. The DNA array was validated using both pure cultures and industrial samples. In addition, comparisons were made between the developed array, PCR assays specifically targeting the thick juice contaminants and classical microbial platings. The array was found to be reliable and sensitive enough to detect and identify the target bacteria. In addition, the array was used to monitor the target microbial populations in thick juice during long-term storage and degradation. Results are discussed in relation to DNA stability in thick juice.     

Present knowledge of the bacterial microflora in the extreme environment of sugar thick juice

The diversity of the bacterial population in sugar thick juice, an intermediate product in the production of beet sugar, which exhibits an extreme, osmophilic environment with a water activity value (a_w) less than 0.86, was assessed with both culture-dependent and -independent 16S ribosomal RNA (rRNA) gene-based analyses. In comparison with previous studies, the number of different thick juice bacterial species increased from 29 to 72. Remarkably, a limited, gram-positive, culturable flora, encompassing species of Bacillus, Staphylococcus and mainly Tetragenococcus dominated thick juice during storage, while a more heterogeneous and unculturable fraction of Acinetobacter, Sporolactobacillus and Thermus species could be detected in freshly produced thick juice. Notably, almost all bacteria detected in the thick juice were also detected in the air, emphasising the importance of further investigation and assessment of strategies to reduce (air) contamination during processing and storage. The discovery of the contamination source may be used for the development of management strategies for thick juice degradation resulting from microbial activity.     

传统发酵豆酱中嗜盐四联球菌的分离鉴定及增鲜菌株筛选

为了获得高产鲜味肽嗜盐四联球菌（Tetragenococcus halophilus）优良菌株,从辽宁省7个不同地区的96份自然发酵豆酱样品中分离出嗜盐四联球菌疑似菌株83株,通过提取各菌株DNA、16S rDNA序列测定和同源性对比分析,其中47株菌为嗜盐四联球菌,分别测定47株菌的产多肽能力、产γ-谷氨酰转肽酶能力、产蛋白酶能力、电子舌风味值分析等指标,结合因子分析以及因子得分综合评价,筛选得到LY9-1产鲜味肽潜力最佳,通过电子舌结果也证明,LY9-1发酵液鲜味值最高,为15.05±0.02,产蛋白酶活力高达（85.45±0.03）U/mL,产γ-谷氨酰转肽酶能力（44.23±0.03）U/m L,产多肽（17.55±0.13）mg/mL,推测该菌株具有较强的增鲜潜力,可为进一步研究增鲜机理以及在发酵食品增鲜中的产业化应用提供理论支持。     

嗜盐四联球菌SNTH-1产多肽发酵条件优化及其呈味、抗氧化特性

该研究以产多肽能力较强的嗜盐四联球菌（Tetragenococcus halophilus）SNTH-1为试验菌株,大豆蛋白为发酵基质,通过单因素试验、Plackett-Burman试验、最陡爬坡试验及Box-Behnken响应面试验对其产多肽培养条件进行优化,并对其呈味及抗氧化特性进行研究。结果表明,嗜盐四联球菌SNTH-1产多肽的最优培养条件为：发酵时间40 h、盐度5.2%、发酵温度37 ℃、接种量3%、初始pH值8.1,在此优化条件下,多肽含量为（32.96±0.02）mg/mL。菌株SNTH-1所产多肽的鲜味、厚味、甜味、咸味、苦味的风味强度较优化前有所提高,清除1,1-二苯基-2-三硝基苯肼（DPPH）自由基的半抑制浓度（IC50）值为0.22 mg/mL。     

Changes in anthocyanins in arils of chitosan-coated pomegranate (Punica granatum L. cv. Rabbab-e-Neyriz) fruit during cold storage

Edible coatings as chitosan treatments (0%, 1% and 2%) were applied to ‘Rabbab-e-Neyriz’ pomegranate (Punica granatum L.). The effect of chitosan coating on individual anthocyanins and colour parameters of the juice during storage at 2 °C or 5 °C was examined. Six predominant anthocyanins were identified in the juice, with up to 935 mg/L total anthocyanins at the time of harvest. Cyanidin 3,5-diglucoside (402 mg/L) was the major pigment. The total anthocyanin content and chroma decreased with storage time in all applied treatments, although lightness and hue angle increased. These changes were reduced with chitosan treatments and at lower storage temperature (2 °C as compared to 5 °C). Based on the obtained results, the diglucoside anthocyanins were more stable than the monoglucosides. Chitosan coating followed by cold storage delayed anthocyanin degradation and prevented colour deterioration in the pomegranate arils.     

Vitamin C Content in Sea Buckthorn Berries (Hippophaë rhamnoides L. ssp. rhamnoides) and Related Products: A Kinetic Study on Storage Stability and the Determination of Processing Effects

ABSTRACT: The primary vitamin in sea buckthorn berries is vitamin C containing values of approximately 400 mg/100 g. Processing effects were investigated during juice and concentrate production from sea buckthorn berries (Hippophaë rhamnoides) and storage stability of juices was determined for up to 7 d using berries and juices from 2 different growing areas. During industrial juice production the technological processing of the berries caused a loss of about 5% to 11% total ascorbic acid (TAA) in the generated juice. The production of the concentrated juice resulted in 50% depletion of TAA. Sea buckthorn berries and juice were stored at 6, 25, and 40 °C for up to 7 d to investigate the temperature effects on TAA during storage. Analysis of kinetic data suggested that the degradation follows a 1st‐order model. The results of the experiments showed that storage of sea buckthorn juices for 7 d at cold temperature (6 °C) already resulted in a degradation of TAA of about 11% to 12%.     

The effects of freezing and thawing on the survival of Escherichia coli O157:H7 in apple juice

Unpasteurized apple juice, adjusted to pH 3.6 to 7.0 was inoculated (10(7) CFU/ml) with single strains of E. coli O157:H7 to evaluate the effect of frozen storage on the viability of this organism. Samples were stored under frozen conditions (-20+/-2 degrees C) for up to 16 days. Cell populations were determined at regular intervals by plating onto tryptic soy agar with added pyruvate (TSAP) or onto sorbitol MacConkey agar (SMA). Populations in the neutralized juice remained unchanged during frozen storage. Populations in non-neutralized juice decreased by 1-3 log10 CFU/ml depending on the strain tested and the pH of the juice. The greatest population decrease was observed with the first freeze/thaw cycle of frozen storage (24 h) and a slow decline in survival occurred thereafter. Injury was observed after 2 weeks of storage when juice pH was at or below pH 4.2. When samples were subjected to multiple freeze/thaw cycles, loss of viability and injury increased with each freeze/thaw cycle.     

Effect of heterologous expression of molecular chaperone DnaK from Tetragenococcus halophilus on salinity adaptation of Escherichia coli

Molecular chaperone DnaK of halophilic Tetragenococcus halophilus JCM5888 was characterized under salinity conditions both in vitro and in vivo. The dnaK gene was cloned into an expression vector and transformed into Escherichia coli. The DnaK protein obtained from the recombinant E. coli showed a significantly higher refolding activity of denatured lactate dehydrogenase than that from non-halophilic Lactococcus lactis under NaCl concentrations higher than 1 M. E. coli without the overexpression of DnaK exhibited a growth profile with a prolonged lag phase and suppressed maximum cell density in Luria-Bertani medium containing 5% (0.86 M) NaCl. On the contrary, the overexpression of T. halophilus DnaK greatly shortened this prolonged lag phase with no effect on maximum growth, while that of L. lactis DnaK decreased maximum growth. The amount of protein aggregates was increased by salt stress in the E. coli cells, while this aggregation was greatly suppressed by the overexpression of T, halophilus DnaK. These results suggest that heterologous overexpression of T. halophilus DnaK, via its chaperone activity, promotes salinity adaptation of E. coli.     

包装材料对贮藏期荔枝汁品质及非酶褐变的影响

研究了聚酯(PET)瓶、聚乳酸(PLA)瓶和聚酯/铝箔/聚乙烯(PET/Al/PE)复合袋装荔枝汁在25℃避光贮藏时的品质变化和非酶褐变情况,采用偏最小二乘法回归(PLSR)分析包装材料对荔枝汁非酶褐变的影响。结果表明:贮藏过程,PLA瓶装荔枝汁的L-抗坏血酸(L-AA)发生快速降解,PET瓶和复合袋中的L-AA降解缓慢;贮藏初期,溶解氧浓度(DOC)均急剧下降,贮藏后期,除PLA瓶中的DOC快速上升外,其它包装材料中的DOC均平缓下降并趋于稳定。包装材料阻隔性能对L-AA含量和DOC有显著(p<0.05)影响。蔗糖不断水解,果糖和葡萄糖含量在贮藏初期快速地增加,然后缓慢下降。氨基酸总量和总酚含量不断下降。可溶性固形物(TSS)含量缓慢上升,pH缓慢下降。5-HMF含量和褐变指数不断上升。包装材料阻隔性能使得荔枝汁中的溶解氧浓度差异,溶解氧能引起L-AA的降解和酚类物质氧化聚合,导致荔枝汁发生非酶褐变。     

Determination of processing effects and of storage stability on vitamin K1 (Phylloquinone) in Sea Buckthorn Berries (Hippophaë rhamnoides L. ssp. rhamnoides) and related products

ABSTRACT:  Phylloquinone (vitamin K₁) is the primary dietary source of vitamin K. Processing effects and stability of phylloquinone were investigated during juice and concentrate production from sea buckthorn ( Hippophaë rhamnoides ) using berries from 2 different growing areas. During industrial juice production the technological processing of the berries caused a loss of about 36% to 54% phylloquinone in the generated juice. The following processing steps leading to the concentrated juice resulted in a complete depletion of phylloquinone. Sea buckthorn berries and juice were stored at 6, 25, and 40 °C for up to 7 d to determine the temperature effects on phylloquinone during storage. Content of vitamin K₁ in sea buckthorn berries was affected by storage time and storage temperature. Storage of freshly harvested berries resulted in a significant increase ( P < 0.01) of phylloquinone ranging from 21% up to 186% (wet weight). The juices showed almost identical significant degradation ( P < 0.01) of phylloquinone of about 18% to 32% at 6, 25, and 40 °C indicating that intensity of decomposition is independent of temperature (6 to 40 °C) and storage time in the range of consumer storage conditions.     

贮藏过程中溶解氧对荔枝汁非酶褐变的影响

本文研究了荔枝汁在不同初始溶解氧浓度及有顶空氧存在的贮藏条件下的非酶褐变情况,采用PLSR进行分析揭示不同溶解氧浓度对荔枝汁非酶褐变的影响规律。结果表明:在0~3 d的贮藏期内,随着DOC的快速下降,低溶解氧和中溶解氧荔枝汁的L-AA进行有氧降解;在3 d~17 w的贮藏期内,DOC下降缓慢,趋于稳定,L-AA进行有氧降解和无氧降解,并以无氧降解为主。随着贮藏前期DOC和顶空氧浓度的快速下降,高溶解氧荔枝汁的L-AA持续进行有氧降解并在第4 w时基本全部降解完毕。三种溶解氧浓度的荔枝汁中的蔗糖均不断降解,但果糖和葡萄糖含量呈现先快速增加后减少的趋势。5-HMF和褐变指数不断地增加,溶解氧浓度越高,增加的越快。表儿茶素不断的降解,而芦丁含量比较稳定。在不同溶解氧浓度下,荔枝汁的非酶褐变是以L-AA降解及酚类物质氧化聚合为主,美拉德反应为辅。     

Comparing equivalent thermal, high pressure and pulsed electric field processes for mild pasteurization of orange juice. Part I: Impact on overall quality attributes

Mild heat pasteurization, high pressure processing (HP) and pulsed electric field (PEF) processing of freshly squeezed orange juice were comparatively evaluated examining their impact on microbial load and quality parameters immediately after processing and during two months of storage. Microbial counts for treated juices were reduced beyond detectable levels immediately after processing and up to 2 months of refrigerated storage. Quality parameters such as pH, dry matter content and brix were not significantly different when comparing juices immediately after treatment and were, for all treatments, constant during storage time. Quality parameters related to pectinmethylesterase (PME) inactivation, like cloud stability and viscosity, were dependent on the specific treatments that were applied. Mild heat pasteurization was found to result in the most stable orange juice. Results for HP are nearly comparable to PEF except on cloud degradation, where a lower degradation rate was found for HP. For PEF, residual enzyme activity was clearly responsible for changes in viscosity and cloud stability during storage.

Industrial relevance

Development of mild processing technologies with a minimal impact on fruit juice can be considered as a true alternative of fresh fruit. The present work presents a fair comparison of mild heat treated, high pressure (HP) and pulsed electric field (PEF) processed orange juice as an alternative for thermal pasteurization. Orange juices were monitored during two months of storage.     

EFFECT OF PACKAGING MATERIALS ON COLOR,VITAMIN C AND SENSORY QUALITY OF REFRIGERATED MANDARIN JUICE

ABSTRACT

Refrigerated mandarin juice was packed in four different containers, three cartons with different composition and one polyethylene terephthalate transparent bottle, and was stored at 4C for 90 days. During the storage of these juices, changes in the headspace gas composition, vitamin C, and CIE L*, a* and b* color coordinates were evaluated. In addition, a consumer panel evaluated the sensory color, fresh mandarin flavor and presence of off‐flavors in the juices. Experimental data indicated that the deterioration of mandarin juices (ascorbic acid degradation and darkening of color) was triggered by the rise in oxygen in the headspace of the storage containers. The type of container played a predominant role in determining the juice quality, with carton containers with an inner layer of aluminum foil providing the juices with the best quality throughout their storage.

PRACTICAL APPLICATIONS

Results from this study will provide manufacturers of mandarin juice with information dealing with the storage and quality of juices packed in different containers. In this way, if manufacturers want to use transparent polyethylene terephthalate (PET) bottles showing the color and appearance of the juice, they will be aware that the shelf life of the juice will be much shorter than in packed cartons; this reduction will be from more than 90 to 36 days (PET bottle). On the other hand, if manufacturers want to use carton for their packaging, they will be aware that using a container with a thick inner layer of aluminum foil will maintain the quality of the juice for a longer time (over 90 days) compared with a shelf life of about 54 days from cartons with an inner layer of ethylene vinyl alcohol copolymers.     

黑加仑果汁中花色苷的贮藏稳定性

以黑加仑果汁为原料,研究不同温度、不同可溶性固形物含量和抗氧化剂（抗坏血酸）存在条件下贮藏过程中果汁花色苷的化学稳定性、降解动力学及颜色变化。结果表明：果汁中花色苷在4 ℃、可溶性固形物含量9°Brix条件下较稳定,随着贮藏温度升高和可溶性固形物含量升高,花色苷残留率越低。贮藏过程中花色苷的降解符合一级反应动力学,反应速率常数k越大,半衰期t1/2越小,活化能Ea越低。添加50 mg/L抗坏血酸使果汁花色苷降解速率常数k明显增大,半衰期t1/2和降解活化能Ea明显减小,因此抗坏血酸对花色苷残留率的减少有促进作用。在贮藏过程中随着时间的推移,花色苷不断的减少,果汁发生褐变,亮度和红色度逐渐降低,黄色度逐渐增加,而且抗坏血酸对贮藏过程中花色苷的颜色变化具有促进作用。     

The effect of storage on the antioxidant activity of reconstituted orange juice which had been pasteurized by high pressure or heat

The effect of storage on the antioxidant activity of pasteurized reconstituted orange juice was studied. Either heat or high pressure was used for pasteurization and total antioxidant activity and ascorbic acid loss were measured during storage at different temperatures (0, 5, 10, 15 °C). The total antioxidant activity of orange juice, calculated as the sum of the activities of different antioxidant compounds, decreased during storage. This was mainly because of loss of ascorbic acid. However, rates of degradation of ascorbic acid were lower for orange juice treated with high pressure and this led to a better retention of its antioxidant activity when compared with juice pasteurized in a conventional way.     

甘氨酸甜菜碱和分子伴侣dnaK在嗜盐四联球菌耐盐机制中的作用

能耐高盐的嗜盐四联球菌CICC 10469是从传统的酱料中分离得到.为研究耐盐特性与相容性溶质积累和分子伴侣dnaK特性的关系,使用核磁共振来检测其主要的相容性溶质.不同盐浓度对耐盐的嗜盐四联球菌和不耐盐的乳酸乳球菌MG 1363的生长的影响通过测量OD600来获得.盐浓度对分子伴侣dnaK表达的影响通过实时荧光定量RT-PCR来检测.相容性溶质和可溶性蛋白随盐浓度的变化通过HPLC和酶标仪来检测.结果显示嗜盐四联球菌以甘氨酸甜菜碱作为主要的相容性溶质.适合的盐浓度能够促进菌体的生长,在一定盐浓度范围内,甘氨酸甜菜碱,可溶性蛋白和分子伴侣dnaK随盐浓度的增加而升高,然而超过一定盐浓度范围后,表达量将会减少.这表明细菌能够根据外部环境做出适当的调整以维持细胞的正常生理功能.而乳酸乳球菌MG1363很少有相容性溶质和分子伴侣dnaK以耐受外部盐浓度.这表明相容性溶质和分子伴侣dnak在嗜盐四联球菌生长中起重要的作用.总之,中度嗜盐菌的主要耐盐机制是通过快速合成和释放甘氨酸甜菜碱及分子伴侣dnaK高活性表达来完成.     

Effect of Storage on Anthocyanin Degradation in Black Mulberry Juice and Concentrates

Anthocyanins present in fruits and vegetables are receiving increased attention because of their potential antioxidant activity but are very susceptible to degradation during processing and storage. Effect of storage on kinetics of anthocyanin degradation and hydroxymethylfurfural (HMF) formation in black mulberry juice and concentrate was determined during 8 months of storage at temperatures of 5°, 20°, 30°, and 40 °C. The monomeric anthocyanin degradation was in accordance with the first-order reaction kinetics and the activation energies of anthocyanin degradation in black mulberry juice and concentrate were found as 56.48 and 49.75 kJ mol^?1, respectively. HMF formation in black mulberry juice and concentrate increased linearly with storage time and temperature and followed zero-order reactions. The activation energies of HMF formation in black mulberry juice and concentrate were found as 75.70 and 104.11 kJ mol^?1, respectively. The losses of antioxidant activity for black mulberry juice and concentrate during storage at different temperatures were in the ranges of 4.87–16.01 and 4.47–33.57 %, respectively. Antioxidant activity in black mulberry juice and concentrate was correlated with total monomeric anthocyanins.     

Bergamot: A source of natural antioxidants for functionalized fruit juices

Bergamot is a common Italian citrus fruit, cultivated almost exclusively to produce essential oils; the juice is considered a waste product, which represents a serious environmental and economic problem for the industries. The aim of this study was to re-evaluate bergamot juice through its chemical characterization and its use to enrich and fortify fruit juices. To investigate this, apples and apricots were used for the laboratory-scale production of fruit juice, following both the traditional industrial recipe and those with the addition of bergamot juice at 10% or 20%, together with or in order to replace the synthetic additives normally used in the industrial process (ascorbic acid and citric acid). The ascorbic acid content and the antioxidant activity were measured during the different steps of juice production and after storage at 37 °C for 15 days to evaluate juice shelf-life. Apricot and apple juices fortified with bergamot juice showed a significant increase in their antioxidant properties and a decreased reduction in ascorbic acid content after the typical production steps. All of the results obtained support the hypothesis that the addition of bergamot juice to juices preserves their ascorbic acid content from thermal degradation and contributes to enhance the antioxidant activity, ensuring a product much richer in antioxidants and ascorbic acid. A preliminary consumer test encouraged the production of bergamot fortified fruit juices. Finally, this is the first time that isorhoifolin and rutin have been detected in bergamot juice.     

Influence of storage temperature and ascorbic acid addition on pomegranate juice

Our recent work shows that in pomegranate juice the destructive effect of ascorbic acid on anthocyanins is not well correlated with changes in the observed colour. However, the addition of vitamin C to this juice has no additional benefit, since ascorbic acid degradation is very rapid and a slightly orange hue results, after a period of storage, in those juices to which ascorbic acid was added. Data are also included on the influence of storage temperature on juice colour stability, indicating that this parameter is more specific for colour alteration and anthocyanin degradation. © 2001 Society of Chemical Industry     

Juice matrix composition and ascorbic acid fortification effects on the phytochemical,antioxidant and pigment stability of açai (Euterpe oleracea Mart.)

The effects of juice matrix composition on the phytochemical stability of açai (Euterpe oleracea Mart.) were evaluated by contrasting natural clarified juice systems with isolated polyphenolic and anthocyanin fractions, in the presence or absence of ascorbic acid (500 mg/l) under accelerated storage conditions (37 °C). Polyphenolic (anthocyanin and non-anthocyanin polyphenolics) and anthocyanin fractions were isolated using C18 Sep-Pak columns and then re-dissolved in the original aqueous juice matrix (unbound fraction) or in a citric acid buffer (pH 3.5). The isolation of anthocyanins from the açai matrix improved their colour stability but a greater retention of non-anthocyanin polyphenolics and antioxidant properties was favoured by the initial juice composition. The presence of non-anthocyanin polyphenolics exerted a protective effect against ascorbic acid oxidation and enhanced polyphenolic and antioxidant stability in isolates fortified with ascorbic acid. However, all isolates obtained from açai experienced significant colour, polyphenolic, and antioxidant losses during storage and this indicated that optimization of early stages of industrial processing, storage and distribution are necessary to retain the functional properties of açai-containing products.