Electron spin resonance studies of free radicals in gamma-irradiated soybean paste.

Free radicals in gamma-irradiated soybean paste were investigated by electron spin resonance (ESR) spectroscopy to determine the effect of temperature (77-296 K) and moisture content (1-54%) of samples irradiated at high dose (1-40 kGy). The samples were kept in liquid nitrogen (77 K) during irradiation and subsequent ESR measurements. The spectra shown at 77 K consisted of the hydrogen atom lines at low and high field and complicated symmetric spectrum. By increasing the microwave power, the line shape of ESR spectra altered, which indicated the detection of different paramagnetic centers at different microwave powers. In saturation curves, it was possible to select four types of spectra components which were different in their relaxation times. By the different irradiation doses, the change in free radical concentration showed a curvilinearly increasing relationship with irradiation dose in wet samples, whereas a proportional relationship was observed with dried samples. This might indicate that the indirect process of free radical formation was involved with the existence of free water radicals in the wet samples.     

Mobility and distribution of water in cassava and potato starches by 1H and 2H NMR

Mobility and distribution of water in cassava (rainy and drought crops) and potato starches were studied by solid state and NMR relaxometry as a function of H(2)O and D(2)O contents ranging from 0 to 44% (dry basis). Measurements of relative mobility derived from (2)H solid state NMR were based on relative area and line shape analysis. The narrow peak (mobile component) started to show at 5% and increased with increasing D(2)O content. This increase in mobile fraction was accompanied by a line narrowing. The mobile fractions of deuterons reached a >98% level above 15% D(2)O, which is well below the water holding capacity of starch ( approximately 27%) and the previously assigned "glassy-rubbery transition point" (24.3%; Jouppila, K.; Roos, Y. H. The physical state of amorphous corn starch and its impact on crystallization. Carbohydr. Polym. 1997, 32, 95-104). This reconfirms the liquidlike nature of water in the so-called glassy state of starch granules. The plasticization effect of water on starch chains was observed at 14-17% for cassava and potato starches as indicated by the T(1) minimum. This, however, did not seem to relate to the difference observed in swelling among the starches studied.     

Analysis of gamma-irradiated melon, pumpkin, and sunflower seeds by electron paramagnetic resonance spectroscopy and gas chromatography-mass spectrometry

Seeds of melon (Citrullus lanatus var. sp.), pumpkin (Cucurbita moschata), and sunflower (Heliantus annus) were gamma-irradiated at 1, 3, 5, and 10 kGy and analyzed by electron paramagnetic resonance (EPR) and gas chromatography-mass spectrometry (GC-MS) according to EN1787:2000 and EN1785:2003, respectively. Distinguishable triplet signals due to the presence of induced cellulose radicals were found at 2.0010-2.0047 g in the EPR spectra. The gamma-irradiated radiolytic markers of 2-dodecylcyclobutanone (2-DCB) and 2-tetradecylcyclobutanone (2-TCB) were identified in all irradiated seed samples. Both the free radicals and the alkylcyclobutanones were found to increase with irradiation dose. In general, linear relationships between the amount of radicals and irradiation dosage could be established. Studies at an ambient temperature (20-25 degrees C) in a humidity-controlled environment showed a complete disappearance of the cellulosic peaks for irradiated samples upon 60 days of storage. Such instability behavior was considered to render the usefulness of using EPR alone in the determination of irradiated seed samples. On the other hand, 2-DCB and 2-TCB were also found to decompose rapidly (>85% loss after 120 days of storage), but the radiolytic markers remained quantifiable after 120 days of postirradiation storage. These results suggest that GC-MS is a versatile and complimentary technique for the confirmation of irradiation treatment to seeds.     

同步辐射(软X射线)对冬小麦的诱变效应及机理研究Ⅲ.辐照种子产生的自由基和辐照引起种胚DNA非按期合成的研究

应用电子自旋 (ESR)波谱术研究了同步辐射辐照种子产生的自由基及其与辐射损伤之间的关系。结果表明 ,同步辐射辐照的种子产生了大量的自由基 ,其种类、性质不同于γ射线 ,短寿命自由基 (小于 48h)产额高于γ射线 ,长寿命自由基 (大于48h)的稳定性大于γ射线。同步辐射自由基产额与辐照剂量呈线性关系。用3H TdR掺入法研究了受辐照种胚萌发早期DNA的合成动态 ,结果表明 ,同步辐射能够引起种胚DNA的损伤修复合成 ,其特点不同于γ射线。原冬 6号和北京41 1品种的DNA修复合成出现在种胚吸水后第一个正常的DNA合成峰之前 ,京冬8号品种出现在第一个小峰之后和 2个高峰之前     

Identification of irradiated cashew nut by electron paramagnetic resonance spectroscopy

Cashew nut samples were irradiated at gamma-radiation doses of 0.25, 0.5, 0.75, and 1 kGy, the permissible dose range for insect disinfestation of food commodities. A weak and short-lived triplet (g = 2.004 and hfcc = 30 G) along with an anisotropic signal (g perpendicular = 2.0069 and g parallel = 2.000) were produced immediately after irradiation. These signals were assigned to that of cellulose and CO 2 (-) radicals. However, the irradiated samples showed a dose-dependent increase of the central line (g = 2.0045 +/- 0.0002). The nature of the free radicals formed during conventional processing such as thermal treatment was investigated and showed an increase in intensity of the central line (g = 2.0045) similar to that of irradiation. Characteristics of the free radicals were studied by their relaxation and thermal behaviors. The present work explores the possibility to identify irradiated cashew nuts from nonirradiated ones by the thermal behaviors of the radicals beyond the period, when the characteristic electron paramagnetic resonance spectral lines of the cellulose free radicals have essentially disappeared. In addition, this study for the first time reports that relaxation behavior of the radicals could be a useful tool to distinguish between roasted and irradiated cashew nuts.     

Scavenging capacity of berry crops on superoxide radicals, hydrogen peroxide, hydroxyl radicals, and singlet oxygen

The antioxidant activities against superoxide radicals (O(2)(*)(-)), hydrogen peroxide (H(2)O(2)), hydroxyl radicals (OH(*)), and singlet oxygen ('O(2)) was evaluated in fruit juice from different cultivars of thornless blackberries (Rubus sp.), blueberries (Vaccinium spp.), cranberries (Vaccinium macrocarpon Aiton), raspberries (Rubus idaeus L. and Rubus occidentalis L.), and strawberries (Fragaria x ananassa Duch.). Among the different cultivars, juice of 'Hull Thornless' blackberry, 'Earliglow' strawberry, 'Early Black' cranberry, 'Jewel' raspberry, and 'Elliot' blueberry had the highest antioxidant capacity against superoxide radicals (O(2)(*)(-)), hydrogen peroxide (H(2)O(2)), hydroxyl radicals (OH(*)), and singlet oxygen ('O(2)). In general, blackberries had the highest antioxidant capacity inhibition of O(2)(*)(-), H(2)O(2), and OH(*). Strawberry was second best in the antioxidant capacity assay for these same free radicals. With regard to 'O(2) scavenging activity, strawberry had the highest value, while blackberry was second. Cranberries had the lowest inhibition of H(2)O(2) activity. Meanwhile, blueberries had the lowest antioxidant capacity against OH(*) and 'O(2). There were interesting and marked differences among the different antioxidants in their abilities to scavenge different reactive oxygen species. beta-Carotene had by far the highest scavenging activity against 'O(2) but had absolutely no effect on H(2)O(2). Ascorbic acid was the best at inhibiting H(2)O(2) free radical activity. For OH(*), there was a wide range of scavenging capacities from a high of 15.3% with alpha-tocopherol to a low of 0.88% with ascorbic acid. Glutathione had higher O(2)(*)(-) scavenging capacity compared to the other antioxidants.     

Extrusion Chemistry of Wheat Flour Proteins: I. Free Radical Formation

Electron paramagnetic resonance (EPR) spectroscopy was used to study free radical production in hard red wheat flours extruded according to a two-level fractional factorial experimental design (11 and 14% protein content, 160 and 185°C, 16 and 20% moisture, 300 and 500 rpm screw speed, and mass flow rate of 225 and 400 g/min). All spectra showed dominant broad singlets (g = 2.0053–2.0059) from nitrogen-centered radicals originating from heat-induced peptide scission and reactions of lipid radicals with side-chain amino groups. At 77 K, sulfur-oxyl or peroxyl radicals (g = 2.008–2.018), thiyl radicals (g = 2.025), and disulfide radical species (g = 2.032–2.035 and 2.05–2.06), resulting from intra- and intermolecular electron migration and shear-induced scission of disulfides, sometimes were present. The strongest EPR signals occurred under conditions of maximum free radical production and minimum opportunity for radical recombination: high protein flour (14%), high die temperature (180°C), and low moisture (16%). EPR signals correlated with sulfhydryl and disulfide (SH-SS) levels and physical properties of extrudates, indicating that free radicals are integrally involved in molecular changes that occur during extrusion.     

高压静电场对农作物种子生物学效应原发机制的探讨

该文选用一些农作物种子为代表，应用高压静电场对其进行处理，然后通过对其处理及对照组进行自由基含量的测定来探讨静电场处理农作物种子的生物效应的原发机制。实验选用大麦、甜菜种子，将种子随机分成2份，其中一份为对照组，另一份为处理组，将处理组置于平行板电极形成的匀强电场中进行处理。电场强度E分别为：E=2.5 kV/cm(大麦)、E=4.5 kV/cm(甜菜)，处理时间均为10 min。处理后将这些种子和对照组种子分别去皮研磨，过120目筛得粉末，用电子天平称取等量粉末装入样品细管，放入电子顺磁共振仪中进行自由基含量的测定，实验结果表明：一定强度的静电场能使农作物干种子的自由基含量显著提高。通过对实验结果的分析，该文提出静电场对作物种子的生物学效应的原发机制是：静电场是通过作用于种子内的水分子及一些生物大分子来提高种子内的自由基的含量而影响种子的活力的。     

Comparative proteomical analysis of zygotic embryo and endosperm from Coffea arabica seeds

During coffee seed development, proteins are predominantly deposited in cotyledons and in the endosperm. Reserve proteins of the 11S family are the most abundant globulins in coffee seeds, acting as a nitrogen source during roasting and guaranteeing flavor and aroma. The aim of the present study was to compare the protein profiles of endosperm and zygotic embryos of coffee seeds. Proteins were extracted from whole seed as well as from embryo and endosperm, separately. Total proteins were analyzed by two-dimensional electrophoresis (2-DE) followed by identification by mass spectrometry (MS). The most abundant spots observed in the gels of coffee seeds were excised, digested with trypsin, and identified by MS as subunits of the 11S globulin. Spots with identical pI and molecular masses were also observed in the protein profiles of coffee endosperm and embryo, indicating that 11S protein is also highly expressed in those tissues. Peptide sequence coverage of about 20% of the entire 11S globulin was obtained. Three other proteins were identified in the embryo and endosperm 2-DE profiles as a Cupin superfamily protein, an allergenic protein (Pru ar 1), exclusive to the endosperm 2D map, and a hypothetical protein, observed only in the zygotic embryo profile.     

Development of probe-based ultraweak chemiluminescence technique for the detection of a panel of four oxygen-derived free radicals and their applications in the assessment of radical-scavenging abilities of extracts and purified compounds from food and herbal preparations

We report here the development of a probe-based ultraweak chemiluminescence (uwCL) method capable of detecting a panel of four oxygen-derived free radicals (ODFRs) including superoxide (O2-), hydrogen peroxide (H2O2), hydroxyl radical (*OH), and peroxyl radical (ROO*) using different probes specific for these radicals performed by the same uwCL analyzer. The selected radical-generating systems and their corresponding uwCL-probing emitters were validated. These ODFR-detecting systems were subsequently utilized by us to assess the radical-scavenging ability (RSA) of a variety of extracts and purified constituents derived from foods and herbal preparations. Our approach for assessing RSA for these constituents is based on the suppression of uwCL generated by each ODFR, and the degrees of inhibition have been shown to be dose-dependent. For this reason, the estimation of IC50 for each testing compound can be obtained from the curve constructed based on the percent of inhibitions of uwCL versus the concentrations of the compound tested. To illustrate the practical applications of our devised methodology, data for comparative studies of RSA activities of fermented extracts of Cordeceps sinensis, purified methylgallate isolated from Toona sinesis, resveratrol purified from grape seeds, plus epimedin C from the aerial part of the Epimedium plant (yinyanghuo) are to be presented.     

The hypocholesterolemic activity of transgenic rice seed accumulating lactostatin, a bioactive peptide derived from bovine milk β-lactoglobulin

Lactostatin is a novel pentapeptide (IIAEK) derived from bovine milk β-lactoglobulin with greater hypocholesterolemic activity than β-sitosterol, the drug commonly used to treat hypercholesterolemia. We developed transgenic rice expressing lactostatin as a fusion protein with seed storage protein (SSP) glutelins under the control of three different endosperm-specific promoters. Lactostatin accumulated in transgenic rice seed at approximately 1.6 mg/g seeds (dry seeds) without any apparent influence on seed traits such as endogenous SSP expression levels or alterations in the intracellular structures of endosperm cells. Short-term (three day) oral administration of the glutelin fraction containing lactostatin (namely three times of 300 mg/kg body weight/day) extracted from transgenic rice seeds resulted in hypocholesterolemic activity in rats; namely, the serum low-density-lipoprotein (LDL) cholesterol level was significantly reduced accompanied by a significant increase in beneficial serum high-density-lipoprotein (HDL) cholesterol.     

胁迫对菜用大豆种子抗坏血酸-谷胱甘肽循环的影响

采用蛭石栽培,在100 mmol/L NaCl 胁迫下,对耐盐性不同的两个菜用大豆［Glycine max (L.)Merr.］品种种子的过氧化氢（H2O2）含量及抗坏血酸-谷胱甘肽（AsA-GSH）循环进行了研究。结果显示,NaCl胁迫显著增加了菜用大豆种子的H2O2含量,但耐盐品种 绿领特早的增幅低于盐敏感品种理想高产95-1。NaCl胁迫期间,绿领特早种子中抗坏血酸过氧化物酶（APX）、脱氢抗坏血酸还原酶（DHAR）、谷胱甘肽还原酶（GR）活性,AsA、GSH含量以及AsA/DHA值和GSH/GSSG值的增幅高于同期的理想高产95-1,或降幅低于同期的理想高产95-1; 脱氢抗坏血酸（DHA）、氧化型谷胱甘肽（GSSG）含量的增幅低于同期的理想高产95-1。表明 绿领特早种子在胁迫期间能够保持较高的AsA-GSH循环效率,可有效地抑制H2O2的积累,这可能是其耐盐性较强的重要原因之一。     

Evolution of the Levels of Free Radicals Generated on Wheat Flour and Wheat Bran by Electron Beam

This paper presents the experimental data regarding the levels of free radicals investigated by electron spin resonance (ESR) on wheat flour and wheat bran exposed to an electron beam. Samples were irradiated with an electron linear accelerator of 6 MeV and then stored at room temperature in air. ESR spectra of the samples indicated presence of free radicals induced by electron beam. Three types of free radicals have been detected in wheat flour, and signals typical for cellulose‐like radicals have been noticed for wheat bran, their concentration increasing with the irradiation dose. The postirradiation storage of both sample types caused radical decay with a two‐step pattern associated with fast decay of short‐life radicals followed by slow reduction of long‐life radicals. However, in the first step, the irradiated wheat bran exhibited more rapid decay of radicals than irradiated flour owing to their different complex matrixes, which led to paramagnetic structures of different origin after exposure to an electron beam. The ESR technique allows the detection of free radicals induced in wheat flour and bran by electron beam for a limited period after irradiation that could be shorter than the shelf life of the products.     

Profiling and characterization antioxidant activities in Anoectochilus formosanus hayata

Phytochemical characteristics and antioxidant activities of the crude and fractionated plant extracts of Anoectochilus formosanus were evaluated using five different assay systems. An acid-treatment (2 N HCl in 95% ethanol) was employed to treat a butanol fraction (BuOH), creating an acid-hydrolyzed BuOH fraction. The IC(50) values for DPPH radicals in the BuOH and acid-hydrolyzed BuOH fractions were 0.521 and 0.021 mg/mL, respectively. The acid-hydrolyzed BuOH exhibited approximately 5-fold higher activity in scavenging superoxide anion than catechin. The acid-hydrolyzed BuOH fraction also effectively protected phi x174 supercoiled DNA against strand cleavage induced by H(2)O(2) and reduced oxidative stress in HL-60 cells. Metabolite profiling showed that the aglycones of flavonoid glycosides in BuOH were produced after acid hydrolytic treatment, and this resulted in a significant increase in antioxidant activities of acid-hydrolyzed BuOH. One new diarylpentanoid, kinsenone, and three known flavonoid glycosides and their derivatives were identified for the first time from A. formosanus, with strong antioxidant properties.     

辐照加速黄酒陈化效果的研究

用适宜剂量辐照黄酒,经半年贮藏,达到陈化一年半的效果。辐照可使黄酒增香、味醇厚、爽口、鲜美。辐照酒的化学指标符合轻工部标准,色、香、味、格保持黄酒特有风味。辐照酒中游离氨基酸总量和人体必需氨基酸量有明显增加。卫生安全性试验表明,对健康无害,符合国家标准。辐照酒中H_2O_2含量无明显增加,未发现长寿命自由基,小白鼠急性LD_(50)>10g/kg体重,属实际无毒,微核实验为阴性。辐照陈化黄酒方法简便,成本低,具有很大的经济效益,有希望在生产中推广应用。     

Status of free radicals in Indian monsooned coffee beans gamma-irradiated for disinfestation

Free radicals in two cultivars of Indian monsooned coffee beans, gamma-irradiated for hygienic and quarantine purposes, were examined by entrapping the small amount of samples in potassium chloride powder in ESR quartz tubes. In contrast to a prominent free radical signal at g = 2.002, observed in spermoderm (silver skin) and cotyledon (whole seed without skin) parts of normal coffee beans, the same was not discernible in monsooned coffee bean parts of both cultivars. The ESR signal was found to be more prominent in the spermoderm than in the whole seed portion of the normal coffee beans. Common practices of roasting and powdering were found to generate quantitatively more free radicals in coffee beans than gamma-irradiation alone. Phenols, contributing maximally to observed free radical signals in coffee beans, were significantly different in normal and monsooned coffee beans. These observations on insignificant free radical population in irradiated monsooned coffee beans may be attributed to their inherent possession of high water activity, favoring decay of free radicals produced. Textural studies with monsooned coffee beans, before and after mild heat treatments, supported these findings.     

Distinctive antioxidant and antiinflammatory effects of flavonols

The antioxidant and antiinflammatory effects of flavonols have been suggested to be structure-related. Results revealed that selected flavonols, including fisetin (F), kaempferol (K), morin (MO), myricetin (MY), and quercetin (Q), exhibited distinctive free radical scavenging properties against different kinds of free radicals. The H donation (DPPH bleaching) potential was Q > F approximately equals MY > MO > K, indicating that the presence of a 3',4'-catechol moiety in the B ring correlated with high activity. The 4'-OH in the B ring was suggested to be important for reducing xanthing/xanthine oxidase-generated superoxide; while an additional OH moiety on the ortho sites (3' or 5') attenuated the effect as the observed inhibitory potency was K approximately equals MO > Q > F > MY. The relative inhibitory effect for Fenton-mediated hydroxyl radical was K approximately equals MO approximately equals Q > F > MY. This result implies the involvement of 4-keto, 5-OH region in Fe++ chelating and the negative effect of pyrogallol moiety in the B ring. Similar to the inhibitory activity against a N-formyl-methionyl-leucyl-phenylalanine (f-MLP)-stimulated oxidative burst in human polymorphonuclear neutrophils (PMN), our result showed that the structural peculiarity of the di-OH in the B ring obviously rendered F, Q, and MO more potent as ROS inhibitors than MY and K, which have tri- and mono-OH in the B ring, respectively. All of the previous data indicated that the structure prerequisite to reinforce the free radical scavenging activity varies with the type of free radical. We further analyzed the effects of flavonols on nitric oxide (NO) production in endotoxin-stimulated murine macrophages, RAW264.7 cells. Results showed that all flavonols (up to 10 microM) inhibited NO production without exerting detectable cytotoxicity. F, K, and Q dose-dependently repressed iNOS mRNA expression and prostaglandin E2 (PGE2) production, in part through an attenuating NF-kappaB signaling pathway. This result indicates that flavonols, despite structural similarity, have different antioxidant and antiinflammatory effects.     

低场核磁共振分析聚乙二醇对萌发期水稻种子水分吸收的影响

为研究聚乙二醇(PEG,polyethylene glycol)处理下水稻种子萌发过程中内部水分分布和变化规律,进而揭示水稻耐旱性在水分吸收规律上的重要特征。应用低场核磁共振的T2弛豫谱和质子密度加权成像分析了PEG处理下水稻种子萌发过程中的水分变化,研究了利用蒸馏水(对照)和质量分数分别为10%、20%PEG6000处理对两个水稻品种旱9710、辽星1发芽指标的影响,以及对两个水稻品种萌发0、6、22、48、72 h吸水量的影响,确定单位质量核磁信号幅值与水稻种子湿基含水率的回归函数关系。发芽指标检测结果显示:旱9710耐旱性高于辽星1。质子密度加权成像结果显示:在水分吸收初期,水分子直接通过种子表面裂缝进入种子体内,胚乳中的淀粉粒等物质开始吸水膨胀,种子体积增大。24 h后,种子内营养物质向种胚流动。PEG处理下,水稻种子吸水量明显减少,发芽速度明显降低,且PEG质量分数越高,发芽速度越慢。基于核磁共振理论及T2弛豫谱的多组分特征,当反演频率为10 000时,水稻种子萌发过程中的水分分为束缚水与自由水两部分。T2弛豫谱结果表明:在蒸馏水和质量分数分别为10%、20%PEG处理下,种子湿基含水率和核磁信号幅值均逐渐增长。PEG处理下,核磁信号幅值增长相比对照处理显著降低(P0.05)。PEG处理抑制了两个水稻品种种子对水分的吸收,PEG质量分数越高,抑制作用越强。PEG处理24 h后,耐旱性强的水稻品种吸水率相比对照处理的降低幅度小于耐旱性弱的品种。回归分析表明,3种处理下,核磁信号幅值和湿基含水率具有一致的线性关系(R2=0.983),由回归方程可以求得水稻种子萌发过程中各状态水分的含量。试验为研究水稻种子萌发过程对干旱胁迫的反应机制,开发种子水分微观活体无损检测技术等方面的研究提供理论支持和数据参考。     

Protective activities of stilbene glycosides from Acer mono leaves against H2O2-induced oxidative damage in primary cultured rat hepatocytes

In our previous study, we isolated two new hepatoprotective stilbene glycosides, 5-O-methyl-(E)-resveratrol 3-O-beta-D-glucopyranoside (MRA) and 5-O-methyl-(E)-resveratrol 3-O-beta-D-apiofuranosyl-(1-->6)-beta-D-glucopyranoside (MRAG), from the methanolic extract of Acer mono leaves. Thereby, we have attempted to elucidate the hepatoprotective mechanism of these compounds, focusing on antioxidative effects, using hydrogen peroxide (H2O2)-injured primary cultures of rat hepatocytes. Both MRA and MRAG showed potent hepatoprotective activities in pretreatment but showed little effects in posttreatment. In addition, they increased the glutathione (GSH) level in the normal control cultures and significantly prevented the depletion of GSH in H2O2-injured primary cultured rat hepatocytes. Moreover, these compounds significantly restored the level of GSH depleted by buthionine sulfoximine or diethylmaleate in the presence or absence of H2O2. Furthermore, these compounds preserved the activities of antioxidant enzymes such as superoxide dismutase, glutathione reductase, and glutathione peroxidase reduced by H2O2 insults. Meanwhile, MRA and MRAG showed moderate scavenging effects with IC50 values of 103.6 and 80.5 microM, respectively, as determined by 1,1-diphenyl-2-picryl-hydrazyl free radical scavenging activity. Taken together, these results suggest that MRG and MRAG exert significant hepatoprotective activities against H2O2-induced hepatotoxicity by maintaining the antioxidative defense system rather than scavenging free radicals.     

Binding of ferric iron to the cell walls and membranes of Bifidobacterium thermophilum: effect of free radicals.

Bifidobacterium thermophilum (ATCC 25866) was incubated with 100-120 microM (59)FeSO(4) and 300 microM excess of H(2)O(2) for up to 120 min in the presence and absence of glucose. Samples were withdrawn after 5, 30, 60, and 120 min. These were protoplasted and (59)Fe(III) measured in the supernatant fraction (cell walls) and protoplasts (cell membranes). These experiments were also repeated in the presence of 400 microM Al(III), which, in whole cells, caused an increase in Fe(III) binding. The amount of iron in the cell wall fraction was constant regardless of time of incubation, was unaffected by Al(III), and was reduced by approximately 20% by glucose. On the other hand, the amount of iron on the protoplasts increased with time and was affected by both Al(III) (upward) and glucose (downward). Scatchard plots indicated that the number of Fe(III) binding sites on the cell walls was 37.6 nmol/mg of dry cell weight at zero time, whereas that of cell membranes was (1)/(10) of that. It was concluded that Fe(III) binding by bifidobacterial cell walls was instantaneous and marginally dependent on free radical action. That of the cell membranes was time-dependent and was due to lipid peroxidation initiated by free radicals. Bifidobacteria can therefore function in the intestinal tract as probiotics by making Fe(OH)(3) unavailable to pathogens and by moderating free radical activity in the intestinal tract.