Novel procedure for the extraction and concentration of carotenoid-containing chromoplasts from selected plant systems

Natural sources of carotenoids for nutraceutical use are desired by the food industry as a result of the increased production of convenience and other highly processed foods. As new physiological roles are discovered for some of the minor carotenoids that are found in only small amounts in present sources, the need for discovery of new sources will amplify. Thus, a method is needed that will effectively and gently concentrate carotenoids from potential new sources for subsequent identification and analysis. A procedure is presented by which carotenoid-containing tissue chromoplasts can be extracted and subsequently concentrated by precipitation, all in an aqueous milieu. The chromoplasts are extracted and solubilized with 0.3% sodium dodecyl sulfate (SDS) in water. The addition of a nominally equal volume of acetonitrile to the chromoplasts in SDS immediately precipitates the chromoplasts out of solution with generally >90% recovery. Carotenoids contained in the concentrated, still-intact chromoplasts can then be solubilized by organic solvent extraction for subsequent analysis. This methodology offers a means to effectively and gently concentrate carotenoids from fruit tissues where yields are often low (e.g., yellow watermelon).     

Isolation and composition of chromoplasts from tomatoes

The fruit of the tomato plant is composed of elongated tomato cells filled with organelles called chromoplasts (plastids). These plastids scattered throughout the cell are rich in nutrients, particularly protein (33%) and lipids (20%). They can be released from the cells by rupture of their cell membranes and then isolated. Plastids and their cell contents can be utilized by the food-processing industry for the preparation of special food products. This study was designed to examine the macronutrient content of isolated tomato plastids and, therefore, determine its potential nutritional value. Use of tomato plastids in pasta sauces and rice dishes, salsa, and extrusion products would increase the nutritional value of the product. Because glucose has been removed in the process of plastid isolation, tomato plastids are useful in the diets of diabetics and cardiovascular patients, as well as for patients in need of weight reduction. Composition comparison of tomato plastid is made with tomato paste, from which glucose has not been removed. Many people require low-sugar products for medical reasons (diabetics and those with cardiovascular disease) and others for weight loss. Therefore, tomato chromoplasts having high protein and lipid contents and low sugar content may be useful in meeting these particular human needs.     

The effects of frozen storage conditions on lycopene stability in watermelon tissue

The purpose of this investigation was to evaluate the rate of deterioration of lycopene in watermelon tissue during frozen storage, because little is known about the stability of watermelon tissue lycopene under cold storage conditions. Heart tissue from each of nine individual watermelons was stored at -20 or -80 degrees C as either small chunks or puree and periodically sampled over a year's time. Initial freeze-thaw experiments indicated that a small percentage of lycopene, approximately 4-6%, degraded during an initial freeze-thaw. Analyses of the samples showed a loss of approximately 30-40% lycopene over a year's storage at -20 degrees C and a loss of approximately 5-10% over the same period at -80 degrees C. Lycopene was slightly more stable in pureed compared with diced watermelon tissue at -20 degrees C, but not at -80 degrees C. The kinetic data were best fitted by application of two simultaneous, first-order decay processes. HPLC analysis of the samples after a year's storage suggested that beta-carotene was more stable during storage at -20 degrees C than was lycopene.     

Carotenoid changes of intact watermelons after storage

Watermelon contains lycopene, a red carotenoid pigment that has strong antioxidant properties. The lycopene content of watermelon is substantial, contributing 8-20 mg per 180 g serving. There are no reports on carotenoid changes in whole watermelon during storage. Three types of watermelon, open-pollinated seeded, hybrid seeded, and seedless types, were stored at 5, 13, and 21 degrees C for 14 days and flesh color, composition, and carotenoid content were compared to those of fruit not stored. Watermelons stored at 21 degrees C had increased pH, chroma, and carotenoid content compared to fresh fruit. Compared to fresh fruit, watermelons stored at 21 degrees C gained 11-40% in lycopene and 50-139% in beta-carotene, whereas fruit held at 13 degrees C changed little in carotenoid content. These results indicate that carotenoid biosynthesis in watermelons can be affected by temperature and storage.     

Carotenoid content of 50 watermelon cultivars

The lycopene content of 50 commercial cultivars of seeded and seedless red-fleshed watermelons was determined. Scanning colorimetric and spectrophotometric assays of total lycopene were used to separate watermelon cultivars into low (<50 mg/kg fw), average (50-70 mg/kg fw), high (70-90 mg/kg fw), and very high (>90 mg/kg fw). Cultivars varied greatly in lycopene content, ranging from 33 to 100 mg/kg. Most of the seeded hybrid cultivars had average lycopene contents. Sixteen of the 33 seedless types had lycopene contents in the high and very high ranges. All-trans-lycopene was the predominant carotenoid (84-97%) in all watermelon cultivars measured by high-performance liquid chromatography, but the germplasm differed in the relative amounts of cis-lycopene, beta-carotene, and phytofluene. Red-fleshed watermelon genotypes vary extensively in carotenoid content and offer opportunities for developing watermelons with specifically enhanced carotenoids.     

Carotenoid pigmentation affects the volatile composition of tomato and watermelon fruits, as revealed by comparative genetic analyses

Tomato near-isogenic lines differing in fruit carotenogenesis genes accumulated different aroma volatiles, in a strikingly similar fashion as compared to watermelon cultivars differing in fruit color. The major volatile norisoprenoids present in lycopene-containing tomatoes and watermelons were noncyclic, such as geranial, neral, 6-methyl-5-hepten-2-one, 2,6-dimethylhept-5-1-al, 2,3-epoxygeranial, (E,E)-pseudoionone, geranyl acetone, and farnesyl acetone, seemingly derived from lycopene and other noncyclic tetraterpenoids. Beta-ionone, dihydroactinodiolide, and beta-cyclocitral were prominent in both tomato and watermelon fruits containing beta-carotene. Alpha-ionone was detected only in an orange-fleshed tomato mutant that accumulates delta-carotene. A yellow flesh (r) mutant tomato bearing a nonfunctional psy1 gene and the yellow-fleshed watermelon Early Moonbeam, almost devoid of carotenoid fruit pigments, also lacked norisoprenoid derivatives and geranial. This study provides evidence, based on comparative genetics, that carotenoid pigmentation patterns have profound effects on the norisoprene and monoterpene aroma volatile compositions of tomato and watermelon and that in these fruits geranial (trans-citral) is apparently derived from lycopene in vivo.     

Factors affecting lycopene oxidation in oil-in-water emulsions

Evidence that dietary lycopene decreases the risk for a number of health conditions has generated new opportunities for the addition of lycopene to functional foods. This work examined the potential of oil-in-water emulsions as a lycopene delivery system for foods. Oil-in-water emulsions containing lycopene were prepared using different kinds of surfactant (cationic, anionic, and nonionic) and oil types (corn oil, stripped corn oil, and hexadecane). The formation of fatty acid oxidation products and the degradation of lycopene and tocopherol were then monitored. Fatty acids and lycopene had greater stability in oil-in-water emulsions stabilized by cationic dodecyltrimethylammonium bromide (DTAB) or nonionic polyoxyethylene (23) lauryl ether than by anionic sodium dodecyl sulfate (SDS). Oxidative stability in the corn oil-in-water emulsions stabilized by SDS was in the following order: tocopherolhexadecane>tocopherol-stripped corn oil. Lycopene degradation rates were similar in emulsions with and without fatty acids, suggesting that lycopene loss was independent of the presence of fatty acids. These results suggest that the stability of lycopene in oil-in-water emulsions could be inhibited by altering the emulsion droplet interface and by the presence of tocopherols.     

Effect of different dextrose equivalent of maltodextrin on the interactions with anionic surfactant in an isothermal titration calorimetry study

Isothermal titration calorimetry (ITC) was used to study interactions between an anionic surfactant (sodium dodecyl sulfate, SDS) and maltodextrins with different dextrose equivalents (DE) in a buffer solution (pH 7.0, 10 mM NaCl, 20 mM Trizma, 30.0 degrees C). The interaction between SDS and maltodextrin was exothermic, which was attributed to incorporation of the hydrocarbon tail of the surfactant into a helical coil formed by the maltodextrin molecules. ITC measurements indicated that the number of SDS molecules bound per gram of maltodextrin increased with decreasing maltodextrin DE, i.e., increasing molecular weight. It was proposed that SDS only binds to maltodextrin molecules that have a DE greater than 10 glucose units.     

拮抗菌强化的生物有机肥对西瓜枯萎病的防治作用

由西瓜专化型尖孢镰刀菌(Fusarium oxysporum f.sp.niveum)引起的西瓜枯萎病是导致西瓜生产毁灭性损失的土传病害,当前对该病尚无有效的防治措施.为了探索该病的生物防治效果,本研究从土壤中分离筛选西瓜枯萎病的拮抗菌,制成生物有机肥,通过温室盆栽试验检验防病效果,并对与拮抗相关的拮抗菌葡聚糖酶进行分子生物学检测.从不同土壤中分离纯化到对西瓜枯萎病菌有潜在拮抗作用的细菌172株,通过平板对峙法筛选出抑菌率在60％以上的拮抗细菌13株,从中挑选出2株抑菌率最高的菌株Cy5和CR38,分别用其与已腐熟的有机肥制成生物有机肥BIO5和BIO38.盆栽试验结果表明,BIO5在防病和促进西瓜生长方面表现优于BIO38.与对照相比,BIO5和BIO38对西瓜枯萎病的相对防治率分别为75％和25％.BIO5处理植株的株高、地上部鲜重、地上部干重分别比对照增加64.8％、63.0％和50％.施用生物有机肥还能显著改变根际土壤的微生物组成.BIO5处理根际土壤的细菌和枯草芽孢杆菌数量分别比对照增加48.5％和61.1％,真菌和尖孢镰刀菌的数量比对照分别下降52.1％和70.2％.分子生物学分析表明,菌株Cy5属于Paenibacillus jamilae菌株,并含有类似于P.polymyxa的β-1,3-1,4-葡聚糖酶.本研究结果说明,拮抗菌强化的生物有机肥对西瓜枯萎病有防治潜力.     

Increasing Slowly Digestible Starch Content of Normal and Waxy Maize Starches and Properties of Starch Products

Changes in the digestibility and the properties of the starch isolated from normal and waxy maize kernels after heat‐moisture treatment (HMT) followed by different temperature cycling (TC) or isothermal holding (IH) conditions were investigated. Moist maize kernels were heated at 80°C for 2 hr. The HMT maize kernels were subjected to various conditions designed to accelerate retrogradation of the starch within endosperm cells. Two methods were used to accelerate crystallization: TC with a low temperature of –24°C for 1 hr and a high temperature of 20, 30, or 50°C for 2, 4, or 24 hr for 1, 2, or 4 cycles, and IH at 4, 20, 30, or 50°C for 24 hr. The starch granules were then isolated from the treated kernels. The starch isolated from HMT normal maize kernels treated by TC using –24°C for 1 hr and 30°C for 2 hr for 2 cycles gave the greatest SDS content (24%) and starch yield (54%). The starch isolated from HMT waxy maize kernels treated by TC using –24°C for 1 hr and 30°C for 24 hr for 1 cycle had an SDS content of 19% and starch yield of 43%. The results suggest that TC after HMT changes the internal structure of maize starch granules in a way that results in the formation of SDS (and RS). They also suggest that thermal treatment of maize kernels is more effective in producing SDS than is the same treatment of isolated starch. All starch samples isolated from treated normal maize kernels exhibited lower peak viscosities, breakdown, and final viscosities and higher pasting temperatures than did the control (untreated normal maize starch). Although peak viscosities and breakdown of the starch isolated from treated waxy maize kernels were similar to those of the control (untreated waxy maize starch), their pasting temperatures were higher. The starch isolated from treated normal and waxy maize kernels with the highest SDS contents (described above) were further examined by DSC, X‐ray diffraction, and polarized light microscopy. Onset and peak temperatures of gelatinization of both samples were higher than those of the controls. Both retained the typical A‐type diffraction pattern of the parent starches. The relative crystallinity of the starch from the treated normal maize kernels was higher than that of the control, while the relative crystallinity of the starch from the treated waxy maize kernels was not significantly different from that of the control. Both treated starches exhibited birefringence, but the granule sizes of both starches, when placed in water, were slightly larger than those of the controls.     

Chromoplast morphology and beta-carotene accumulation during postharvest ripening of Mango Cv. 'Tommy Atkins'

Accumulation of beta-carotene and trans-cis isomerization of ripening mango mesocarp were investigated as to concomitant ultrastructural changes. Proceeding postharvest ripening was shown by relevant starch degradation, tissue softening, and a rising sugar/acid ratio, resulting in a linear decrease (R (2) = 0.89) of a ripening index (RPI(KS)) with increasing ripening time. A modest accumulation of all-trans-beta-carotene and its cis isomers resulted in a slight pigmentation of the mango chromoplasts, because ambient temperatures of 18.2-19.5 degrees C provided suboptimal ripening conditions, affecting color development and beta-carotene biosynthesis. The ultrastructures of chromoplasts from mango mesocarp and carrot roots were comparatively studied by means of light and transmission electron microscopy. Irrespective of the ripening stage, mango chromoplasts showed numerous plastoglobuli varying in size and electron density. They comprised the main part of carotenoids, thus supporting the partial solubilization of the pigments in lipid droplets. However, because different pigment-carrying tubular membrane structures were also observed, mango chromoplasts were assigned to the globular and reticulotubular types, whereas the crystalline type was confirmed for carrot chromoplasts. The large portions of naturally occurring cis-beta-carotene in mango fruits contrasted with the predominance of the all-trans isomer characteristic of carrots, indicating that the nature of the structure where carotenoids are deposited and the physical state of the pigments are crucial for the stability of the all-trans configuration.     

Characterization of interactions between chitosan and an anionic surfactant

Chitosan is a cationic biopolymer that has many potential applications in the food industry because of its unique nutritional and physiochemical properties. Many of these properties depend on its ability to interact with anionic surface-active molecules, such as phospholipids, surfactants, and bile acids. The purpose of this study was to characterize the interaction between chitosan and a model anionic surfactant (sodium dodecyl sulfate, SDS) using isothermal titration calorimetry (ITC), surfactant-selective electrode (SSE), and turbidity measurements. ITC and SSE indicated that SDS bound strongly to chitosan via a highly exothermic interaction. The turbidity measurements indicated that chitosan formed insoluble complexes with SDS that strongly scattered light. The chitosan bound approximately 4 mM of SDS per 0.1 wt % chitosan before becoming saturated with surfactant. The SDS-chitosan interaction was weakened appreciably by the presence of 100 mM NaCl, which suggested that it was electrostatic in origin. This study provides information about the origin and characteristics of molecular interactions between chitosan and anionic surface-active lipids that may be useful for the rational design of chitosan-based food ingredients with specific nutritional and functional characteristics, e.g., cholesterol lowering or fat replacement.     

Extraction and Activation of Wheat Polyphenol Oxidase by Detergents: Biochemistry and Applications

Polyphenol oxidases (PPOs) from several plant species, including wheat, have been implicated in undesirable brown discolorations of food products. It has been demonstrated that these enzymes are often present in a latent form or are membrane‐associated, necessitating detergent or other treatments to obtain fully active preparations. Here, the influence of different detergents on wheat meal and flour PPOs was investigated. Extraction in presence of 50 mM SDS led to a 5‐ to 15‐fold increase in PPO activity, making quantitative assays in flour from low‐PPO lines more robust. Among a series of additional nonionic, anionic, and cationic detergents tested, only n ‐lauroylsarcosine increased extractable PPO activity to a degree comparable to that of SDS. Additional experiments suggested that a large fraction of wheat meal PPOs may be membrane‐associated and that SDS is able to activate PPOs extracted from high‐activity but not from low‐activity wheat lines. PPO activities assayed after SDS extraction of meal and flour were highly correlated with each other and with activity determined in whole (intact) kernels in absence of SDS. Correlation coefficients between PPO activities measured with all these methods and noodle brightness were about equal, indicating that activities assayed after SDS extraction are useful for germplasm screening and quality prediction.     

Combined pressure-temperature effects on carotenoid retention and bioaccessibility in tomato juice

This study highlights the changes in lycopene and β-carotene retention in tomato juice subjected to combined pressure-temperature (P-T) treatments ((high-pressure processing (HPP; 500-700 MPa, 30 °C), pressure-assisted thermal processing (PATP; 500-700 MPa, 100 °C), and thermal processing (TP; 0.1 MPa, 100 °C)) for up to 10 min. Processing treatments utilized raw (untreated) and hot break (～93 °C, 60 s) tomato juice as controls. Changes in bioaccessibility of these carotenoids as a result of processing were also studied. Microscopy was applied to better understand processing-induced microscopic changes. TP did not alter the lycopene content of the tomato juice. HPP and PATP treatments resulted in up to 12% increases in lycopene extractability. all-trans-β-Carotene showed significant degradation (p < 0.05) as a function of pressure, temperature, and time. Its retention in processed samples varied between 60 and 95% of levels originally present in the control. Regardless of the processing conditions used, <0.5% lycopene appeared in the form of micelles (<0.5% bioaccessibility). Electron microscopy images showed more prominent lycopene crystals in HPP and PATP processed juice than in thermally processed juice. However, lycopene crystals did appear to be enveloped regardless of the processing conditions used. The processed juice (HPP, PATP, TP) showed significantly higher (p < 0.05) all-trans-β-carotene micellarization as compared to the raw unprocessed juice (control). Interestingly, hot break juice subjected to combined P-T treatments showed 15-30% more all-trans-β-carotene micellarization than the raw juice subjected to combined P-T treatments. This study demonstrates that combined pressure-heat treatments increase lycopene extractability. However, the in vitro bioaccessibility of carotenoids was not significantly different among the treatments (TP, PATP, HPP) investigated.     

不同嫁接砧木对旱砂田西瓜生长及品质和产量的影响

近年来宁夏旱砂西瓜产区嫁接栽培迅速发展,但砧木品种繁杂。为筛选出适宜宁夏的旱砂西瓜嫁接砧木品种,选用10个南瓜砧木品种,观察了不同砧木对旱砂田西瓜生长、果实品质及产量的影响。结果表明,采用宁砧1号、宝根103、金城雪峰嫁接的旱砂西瓜折合产量较高,分别为42 470.37 、41 877.78、41 844.44 kg/hm2。宁砧1号嫁接的旱砂西瓜果皮硬度、果肉硬度、果实纵径最高,皖砧2号、宝根103、思状7号嫁接的旱砂西瓜果皮较厚,金城雪峰嫁接的旱砂西瓜果实中心可溶性固形物含量及总糖含量最高,宝根103嫁接的旱砂西瓜果实边缘可溶性固形物含量及总酸含量均高于其他砧木,宁砧1号嫁接的旱砂西瓜果实Vc含量均显著高于其他砧木。综合考虑认为,砧木品种金城雪峰、宝根103、宁砧1号对提高旱砂西瓜品质和产量的效果最好。     

Cellular uptake of carotenoid-loaded oil-in-water emulsions in colon carcinoma cells in vitro

Oil-in-water emulsions allow the preparation of lipophilic compounds such as carotenoids in the liquid form. Here, the effect of a combination of some emulsifiers, such as two whey protein isolates (BiPro and BioZate), sucrose laurate (L-1695), and polyoxyethylene-20-sorbitan-monolaurate (Tween 20), on the stability of lycopene and astaxanthin in emulsions, droplet size, and cellular uptake of these carotenoids has been investigated. The degradation of lycopene was slightly more pronounced than that of astaxanthin in all emulsions. The concentration of lycopene and astaxanthin decreased by about 30% and 20%, respectively, in all emulsions after 3 weeks of storage in the dark at 4 degrees C. The kind of emulsifiers or their combinations have played an important role in the cellular uptake by the colon carcinoma cells line HT-29 and Caco-2.     

西瓜枯萎病生防菌枯草芽孢杆菌B11菌株高产拮抗物质的诱变选育

从西瓜根围分离得到的一株有效抑制西瓜枯萎病菌的枯草芽孢杆菌B 11菌株(B acillus subtilisstra in B 11)。为获得高产拮抗物质的诱变菌株,采用紫外线诱变方法对B 11菌株进行诱变,结果获得抑菌活性比野生菌B 11菌株强的6株诱变菌株,其中G 17菌株的抑菌活性及粗拮抗物质含量均高于B 11菌株,且遗传性较稳定;再以G 17菌株为供试菌株,用亚硝基胍(NTG)对其进行化学诱变,结果获得抑菌活性较强的4株诱变菌株,其中BV 22菌株对西瓜枯萎病菌的抑菌活性最高,其抑菌活性比G 17和B 11菌株分别提高了36.88%和80.00%;其粗拮抗物质的含量分别提高了30.15%和62.53%。研究结果说明,用物理诱变B 11菌株后获得的诱变菌株再用化学方法进行诱变,可选育出高产拮抗物质的诱变菌株。     

西瓜根际促生菌筛选及生物育苗基质研制

通过从西瓜根际分离筛选具根际定殖能力的植物根际促生菌,将其保活添加至普通育苗基质研制生物育苗基质,以确保功能菌株能够在苗期定殖根际,进而在移栽后发挥促生功能。结果表明,分离获得一株同时具有产吲哚乙酸(IAA)和NH_3,且对尖孢镰刀菌和茄科劳尔氏菌均有拮抗作用的植物根际促生菌(PGPR)菌株N23;在三季育苗试验中,与普通基质处理(CK)相比,添加菌株N23的生物育苗基质所育种苗,在多项苗期生长指标上均表现出稳定的促生作用;盆栽试验表明,除叶绿素相对含量测量值(SPAD)外,生物基质所育西瓜种苗的其他检测指标均显著高于对照(普通育苗基质所育种苗,下同);田间试验表明,生物基质所育种苗西瓜、黄瓜、辣椒和番茄种苗移苗后,在苗期植株株高和茎粗均显著优于对照,在产量上均增产10%以上。结合形态、生理生化特征和16S rDNA基因序列分析,初步鉴定菌株N23为芽孢杆菌属细菌(Bacillus sp.)。综上,利用芽孢杆菌N23研制的生物育苗基质能够有效促进所育不同作物种苗质量,增强移栽后作物的生长和田间产量。因此,本研究能够为根际有益微生物的应用提供新的思路,为生物育苗基质的研制提供理论支撑。     

Chemical and cellular antioxidant activity of phytochemicals purified from olive mill waste waters

The isolation and identification of a phytocomplex from olive mill waste waters (OMWW) was achieved. The isolated phytocomplex is made up of the following three phenolic compounds: hydroxytyrosol (3,4-DHPEA), tyrosol (p-HPEA) and the dialdehydic form of decarboxymethyl elenolic acid, linked with (3,4-dihydroxyphenyl)ethanol (3,4-DHPEA-EDA). The purification of this phytocomplex was reached by partial dehydration of the OMWW, followed by liquid-liquid extraction with ethyl acetate and middle pressure liquid chromatography (MPLC) on a Sephadex LH-20 column. The phytocomplex accounted for 6% of the total phenolic content of the OMWW. The phytocomplex and individual compounds were tested for antioxidant capacity by the oxygen radical absorbance capacity (ORAC) method. The ORAC phytocomplex produced 10,000 ORAC units/g dry weight, whereas the cellular antioxidant activity, measured by the cellular antioxidant activity in red blood cell (CAA-RBC) method, demonstrated that the phytocomplex and all of the components are able to permeate the cell membrane thus exhibiting antioxidant activity inside the red blood cells. Our phytocomplex could be employed in the formulation of fortified foods and nutraceuticals, with the goal to obtain substantial health protective effects due to the suitable combination of the component molecules.