Studies on Enzymatic Liquefaction and Saccharification of Starch

It was confirmed that turbid substances which remained in saccharified liquid after digestion of starch slurry with bacterial alpha-amylase and glucoamylase were insoluble starch particles and that these materials were formed during heat treatment of the slurry in liquefaction process.

In iodine staining reaction, the suspension of these materials showed only a slightly dirty blue color; even after boiling of this suspension for 30 min at 135°C, this staining reaction was not complete and the maximum color intensity could be obtained after treatment with 1 n caustic soda solution.

It is assumed from the iodine color reaction, that the main component of these materials was glucose polymer of straight chain, like that of amylose, and that this would be strongly associated with each other to form the compact molecule.

The method for determination of these insoluble starch particles in the starch slurry was established and by this method the contents of these materials were determined in some types of starches.     

Protein aggregation and degradation during iodine labeling and its consequences for protein adsorption to biomaterials

Protein adsorption on modified and unmodified polymer surfaces investigated through radiolabeling experiments showed a tendency for higher than expected albumin and immunoglobulin G (IgG) adsorption. Possible enhanced protein aggregation and degradation caused by the iodine labeling method used were analyzed through chromatography and spectroscopy techniques. Results show that the iodine labeling method using chloramine-T (CAT) as an oxidizing agent can cause both enhanced aggregation and fragmentation of proteins. Albumin shows an enhanced tendency to aggregate after iodine labeling using the CAT method, and higher amounts of fragmentation are observed for CAT-labeled IgG molecules relative to unlabeled IgG molecules as well as to IgG molecules labeled using the Iodo-Gen method. These results show that the widely applied method of radioisotope labeling for quantitative assessment of protein adsorption should be used with caution and preferably should be validated by a label-free methodology for each combination of radiolabel and protein. The results obtained in this study can be used to optimize investigation of protein adsorption on surfaces of materials for biomedical devices.     

Investigation of the effect of amylose/iodine complexation on the conformation of amylose in aqueous solution

Using a potato amylose fraction of 8 × 10⁵, molecular-weight viscosity studies were carried out at 25°C on solutions containing 0.176–0.042% polymer, 8.67 mM KI, 1% ethanol, and different concentrations of iodine. By a novel extrapolation method, the intrinsic viscosities of the amylose/iodine complex were determined under various conditions of iodine binding (0–0.133 g I₂/g amylose). Contrary to the view long held in this research area, it was found that the intrinsic viscosity of amylose solutions decreases significantly upon complex formation with iodine. Taking into account the results of our previous kinetic studies, the present findings are interpreted in terms of an amylose model characterized by loose, extended helical regions which are interrupted by short disordered regions. It is proposed that the intrinsic viscosity decrease observed is due to a shortening of the linear dimension of the polymer chain. This conformation change is apparently caused by the contraction of loose helical regions of the amylose macromolecule due to the entrapment of iodine (and perhaps other) atoms inside the helical cavities.     

海藻中有机碘的研究Ⅰ.海藻中有机碘含量测定

采用国际上目前通用的差减法来计算海藻中的有机碘,即首先测定海藻中的总碘和无机碘,其差减值为有机碘.碘的测定采用了碘离子选择电极法和中子活化法作为对照。在此基础上,还测定了新鲜海带中碘的含量、分布以及有机碘和无机碘的比例。研究结果表明,海带中碘的平均含量占鲜重的0.133％,其中88％的碘是以碘离子的形式存在,有机碘只占总碘的12％,同时海带不同部位碘的含量不同,叶部外缘含碘较多,是叶中部的2倍左右,尤其叶尖部的含量达到鲜重的0.183％。而有机碘的含量分布规律则不同,有机碘的含量在靠近根部的位置较高,为鲜重的13.9％。这种分布特点可能与海带的生命活动规律有关。     

The interaction of lipids with iodine in monomolecular films

The interactions of iodine with oxidized cholesterol, dipalmitoyllecithin and egg lecithin were studied by the monolayer method. It was found that iodine is incorporated in the hydrophobic region of the film, unsaturated bonds being essential for the process. No interaction with hydrophilic groups could be detected. The iodide ion had no effect on the incorporation. The amount of incorporated iodine was the largest for egg lecithin, and less for oxidized cholesterol, while dipalmitoyllecithin showed no measurable effect. This tendency is in accordance with the charge-transfer complexing abilities of the lipids and the effect of iodine on the conduction of hydrated lipid samples. Our results thus favour the electronic conduction mechanism in iodine-doped bilayer systems.     

Iodine uptake and loss-can frequent strenuous exercise induce iodine deficiency?

Most of the daily dietary iodine intake (approximately 90 %) will be excreted in the urine; measurement of urinary iodine excretion is thus routinely used as an index of dietary iodine intake. However, urinary excretion is not the only means of iodine loss. Subjects such as athletes or those participating in vigorous exercise can lose a considerable amount of iodine in sweat, depending on environmental factors such as temperature and humidity. In areas of lower to moderate dietary iodine intake, loss in sweat can equal that in urine. Although electrolyte loss in sweat is well-recognized and replacement strategies are adopted, there is less recognition of potential iodine loss. Crude calculations reveal that if sweat iodide losses are not replaced, dietary stores could be depleted in an athlete undergoing a regular training regime. The significance of these losses could be increased in areas where dietary iodine intake is lower in the summer months. Although there is little doubt that excessive sweating can induce a relative iodine deficiency state, there is no case as yet for iodine supplementation in those that take vigorous exercise. However, sustained iodine loss may have implications for thyroid status and possibly consequences for athletic performance.     

Activation of iodine into a free-radical intermediate by superoxide: a physiologically significant step in the iodination of tyrosine

A pivotal biochemical event in the thyroid physiology is identified unravelling a superoxide anion radical-mediated activation of iodine into an active I.- form, which could be the intermediate that is incorporated onto tyrosine. This active iodine species gives fairly stable spin-adducts with PBN that could be characterized using EPR spectroscopy. Thus, a long-lasting puzzle regarding the iodine intermediate formed before iodination of tyrosine seems to be solved.     

The Mechanism of the Gram Reaction. II. The Function of Iodine in the Gram Stain

Fifty-five reagents were studied as to their ability to replace iodine in the Gram stain. None gave results as good as iodine. Eight gave usable Gram preparations, and forty-seven gave negative results. Omission of the counterstain resulted in increasing to thirty-three the number of reagents giving differentiation, but this, was not considered a true Gram differentiation. Many oxidizing agents were shown not to be substitutes for iodine; therefore the function of iodine must be more than to serve as an oxidizing agent. Many reagents which formed precipitates with the dye could not replace iodine; therefore factors other than precipitate formation must be involved. However, all agents which were good substitutes for iodine were both good oxidizing and dye precipitating agents. Experiments involving the study of cell membrane permeability showed that Gram-positive cells were less permeable to iodine in alcoholic solution than Gram-negative cells. This difference could not be demonstrated for iodine in aqueous solution. It was concluded that iodine served to form a dye-iodine precipitate (or complex) in the cell. Since Gram-positive cells were less permeable to iodine in alcohol than Gram-negative cells, this resulted in a slower dissolving out of this complex from Gram-positive cells during de-colorization and hence a slower decolorization time. The relative solubilities of dye precipitates in alcohol and in aqueous safranin solution were also indicated as an important factor influencing decolorization. Dyes which formed highly soluble precipitates with iodine could not be used in the Gram stain. It is not proposed that the mechanism of the Gram stain is entirely one of membrane permeability; chemical factors are undoubtedly important and will be discussed in a later paper. However, it is proposed that the chemical and physical factors are closely interrelated in the Gram stain mechanism.     

Biodistribution and Ct-Imaging Characteristics of Iopromide-Carrying Liposomes in Rats

Abstract

Biodistribution and computed tomography (CT) imaging characteristics of iopromide-carrying liposomes were investigated in healthy or tumor-bearing rats. The mean diameter of the liposomes obtained by the ethanol evaporation method was approximately 0.5 urn and the encapsulation amounted to 32 %. In the biodistribution study a significant accumulation of the liposomal contrast medium was observed at both examined doses (250 and 1000 mg total iodine/kg b.w.) in liver and spleen. A dose-dependent enrichment in these organs could be demonstrated. Increasing the iodine and thus lipid dose resulted in a marked increase in blood iodine concentration for prolonged time periods due to saturation of liver uptake. Increasing the injection rate 20-fold at a dose of 250 mg iodine/kg b.w. did not significantly (p > 0.05) change the biodistribution behaviour. In the CT study in healthy rats doses in the range from 100 to 2000 mg iodine/kg were investigated/There was an increase in density in liver and spleen in all animals immediately after the intravenous injection. At lower doses the increase in liver density reached the maximum level within a few minutes and remained almost unchanged during the whole study period (24 h). At doses above 500 mg total iodine or lipid/kg b.w. saturation of liver phagocytosis could be observed. In tumor-bearing rats (Novikoff hepatoma) 150 and 200 mg iodine/kg resulted in a significant increase in density difference between liver and tumor. Liver lesions down to a size of about 5 mm could be clearly delineated over the whole study period (up to 30 minutes).     

A rapid,sensitive, simple plate assay for detection of microbial alginate lyase activity

Screening of microorganisms capable of producing alginate lyase enzyme is commonly carried out by investigating their abilities to grow on alginate-containing solid media plates and occurrence of a clearance zone after flooding the plates with agents such as 10% (w/v) cetyl pyridinium chloride (CPC), which can form complexes with alginate. Although the CPC method is good, advantageous, and routinely used, the agar in the media interferes with the action of CPC, which makes judgment about clearance zones very difficult. In addition, this method takes a minimum of 30 min to obtain the zone of hydrolysis after flooding and the hydrolyzed area is not sharply discernible. An improved plate assay is reported herein for the detection of extracellular alginate lyase production by microorganisms. In this method, alginate-containing agar plates are flooded with Gram's iodine instead of CPC. Gram's iodine forms a bluish black complex with alginate but not with hydrolyzed alginate, giving sharp, distinct zones around the alginate lyase producing microbial colonies within 2–3 min. Gram's iodine method was found to be more effective than the CPC method in terms of visualization and measurement of zone size. The alginate-lyase-activity area indicated using the Gram's iodine method was found to be larger than that indicated by the CPC method. Both methods (CPC and Gram's iodine) showed the largest alginate lyase activity area for Saccharophagus degradans (ATCC 43961) followed by Microbulbifer mangrovi (KCTC 23483), Bacillus cereus (KF801505) and Paracoccus sp. LL1 (KP288668) grown on minimal sea salt medium. The rate of growth and metabolite production in alginate-containing minimal sea salt liquid medium, followed trends similar to that of the zone activity areas for the four bacteria under study. These results suggested that the assay developed in this study of Gram's iodine could be useful to predict the potential of microorganisms to produce alginate lyase. The method also worked well for screening and identification of alginate lyase producers and non-producers from environmental samples on common laboratory media. They did this by clearly showing the presence or absence of clearance zones around the microbial colonies grown. This new method is rapid, efficient, and could easily be performed for screening a large number of microbial cultures. This is the first report on the use of Gram's iodine for the detection of alginate lyase production by microorganisms using plate assay.     

Changes in Blood-Brain Barrier Nutrient Transport in the Offspring of Iodine-Deficient Rats and Their Preventability

Thyroid hormones affect the structure and function of biological membranes. Whether or not they affect the Blood-Brain Barrier nutrient transport, the rate limiting membrane transport regulating nutrient supply to brain is to be established yet. That the impaired brain development and function seen in iodine deficiency could be due to such an effect has been assessed in situ by the brain uptake index (BUI) method in Wistar/NIN rat pups born to dams subjected to dietary iodine deficiency/rehabilitation for different times. Compared to controls (C), there was a significant decrease in the BUI values of 2-Deoxy-D-Glucose (2-DG) and L-leucine (Leu) in the pups (D1) born to dams chronically fed low iodine test (LIT) diet through their active growth and subsequent pregnancy and lactation. Surprisingly transport of L-Tyrosine (Tyr) and sucrose (the background marker) was not altered, nor was the BBB transport of all these nutrients affected by feeding LIT diet during the mothers' gestation (D2) and lactation (D3) only. The hypothyroidism in D1 pups was only moderate and preventable by rehabilitation of mothers with control diet from conception (R1) or parturition (R2), as were the changes in BBB nutrient transport. The results suggest that chronic material dietary iodine deficiency impairs BBB nutrient transport in the offspring and this could be prevented by their rehabilitation with iodine.     

Iodine deficiency activates antioxidant genes and causes DNA damage in the thyroid gland of rats and mice

Because thyroid nodules are frequent in areas with iodine deficiency the aim of this study was to characterise molecular events during iodine deficiency that could explain mutagenesis and nodule formation. We therefore studied gene expression of catalytic enzymes prominent for H(2)O(2) detoxification and antioxidative defence, quantified DNA oxidation and damage as well as spontaneous mutation rates (SMR) in mice and rats fed an iodine controlled diet. Antioxidative enzymes such as superoxide dismutase 3, glutathione peroxidase 4 and the peroxiredoxins 3 and 5 showed increased mRNA expression, which indicates increased radical burden that could be the cause of additional oxidized base adducts found in thyroidal genomic DNA in our experiments of iodine deficiency. Furthermore, the uracil content of thyroid DNA was significantly higher in the iodine-deficient compared to the control group. While SMR is very high in the normal thyroid gland it is not changed in experimental iodine deficiency. Our data suggest that iodine restriction causes oxidative stress and DNA modifications. A higher uracil content of the thyroid DNA could be a precondition for C-->T transitions often detected as somatic mutations in nodular thyroid tissue. However, the absence of increased SMR would argue for more efficient DNA repair in response to iodine restriction.     

Quantification of total iodine in intact granular starches of different botanical origin exposed to iodine vapor at various water activities

Iodine has been used as an effective tool for studying both the structure and composition of dispersed starch and starch granules. In addition to being employed to assess relative amylose contents for starch samples, it has been used to look at the molecular mobility of the glucose polymers within intact starch granules based on exposure to iodine vapor equilibrated at different water activities. Starches of different botanical origin including corn, high amylose corn, waxy corn, potato, waxy potato, tapioca, wheat, rice, waxy rice, chick pea and mung bean were equilibrated to 0.33, 0.75, 0.97 water activities, exposed to iodine vapor and then absorbance spectra and LAB color were determined. In addition, a new iodine quantification method sensitive to <0.1% iodine (w/w) was employed to measure bound iodine within intact granular starch. Amylose content, particle size distribution of granules, and the density of the starch were also determined to explore whether high levels of long linear glucose chains and the surface area-to-volume ratio were important factors relating to the granular iodine binding. Results showed, in all cases, starches complexed more iodine as water content increased and waxy starches bound less iodine than their normal starch counterparts. However, much more bound iodine could be measured chemically with waxy starches than was expected based on colorimetric determination. Surface area appeared to be a factor as smaller rice and waxy rice starch granules complexed more iodine, while the larger potato and waxy potato granules complexed less than would be expected based on measured amylose contents. Corn, high amylose corn, and wheat, known to have starch granules with extensive surface pores, bound higher levels of iodine suggesting pores and channels may be an important factor giving iodine vapor greater access to bind within the granules. Exposing iodine vapor to moisture-equilibrated native starches is an effective tool to explore starch granule architecture.     

Increment of Iodine Content in Vegetable Plants by Applying Iodized Fertilizer and the Residual Characteristics of Iodine in Soil

As a new attempt to control iodine deficiency disorder (IDD), we explored a method of iodine supplementation by raising the iodine content in vegetables. When grown in the soil supplemented with iodized fertilizer, the three experimental plant species (cucumber, aubergine, and radish) show increasing iodine levels in both leaf and fruit/rhizome tissues as the iodine content added in soil increases. Excessive iodine added to soil can be toxic to plants, whereas the tolerance limit to excessive iodine varies in the three plant species tested. The migration and volatilization of iodine in soil is correlated with the properties of the soil used. The residual iodine in soil increases as the iodine added to soil increases. The diatomite in the iodized fertilizer helps to increase the durability of the iodized fertilizer. This study potentially provides a safe and organic iodine supplementation method to control IDD.     

Chemical and enzymatic modification of proteins in the 30 S ribosome of Escherichia coli

Methods are described for the iodination of ribosomal proteins by iodine monochloride and potassium iodide and bovine lactoperoxidase. Ribosomes that were maximally iodinated did not synthesize polyphenylalanine. About one-half of the tyrosine residues could be iodinated with iodine monochloride in the intact ribosome with no change in the sedimentation properties of the particle. When proteins were extracted and dissolved in 5 m-urea, all of the tyrosine residues could be iodinated with iodine monoehloride.     

A spectrophotometric assay for lipid peroxides in serum lipoproteins using a commercially available reagent

A method is described for measuring lipid peroxides by means of the color reagent of a commercially available test kit for cholesterol estimation. In principle, this assay makes use of the oxidative capacity of lipid peroxides to convert iodide to iodine, which can be measured photometrically at 365 nm. Calibration curves were obtained using peroxides such as H2O2, t-butyl hydroperoxide, and cumene hydroperoxide. A stoichiometric relationship was observed between the amount of organic peroxides assayed and the concentration of iodine produced. Concentrations of lipid peroxides as small as 1 nmol/ml could be measured. The ability to estimate lipid peroxides of isolated low density lipoprotein was demonstrated.     

Size and CT density of iodine-containing ethosomal vesicles obtained by membrane extrusion: Potential for use as CT contrast agents

Computed tomography (CT) is the primary non-invasive imaging technique used for most patients with suspected liver disease. In order to improve liver-specific imaging properties and prevent toxic effects in patients with compromised renal function, we investigated the encapsulation of iodine within ethosomal vesicles. As a first step in the development of novel contrast agents using ethosomes for CT imaging applications, iodine was entrapped within ethosomes and iodine-containing ethosomes of the desired size were obtained by extrusion using a polycarbonate membrane with a defined pore size. Ethosomes containing iodine showed a relatively high CT density, which decreased when they were extruded, due to the rupture and re-formation of the lipid bilayer of the ethosome. However, when a solution with a high iodine concentration was used as a dispersion media during the extrusion process, the decrease in CT density could be prevented. In addition, ethosomes containing iodine were taken up efficiently by macrophages, which are abundant in the liver, and these ethosomes exhibited no cellular toxicity. These results demonstrate that iodine could be entrapped within ethosomal vesicles, giving the ethosomes a relatively high CT density, and that the extrusion technique used in this study could conveniently and reproducibly produce ethosomal vesicles with a desired size. Therefore, ethosomes containing iodine, as prepared in this study, have potential as contrast agents with applications in CT imaging.     

碘与甲状腺疾病

碘是人体必需的微量元素,是合成甲状腺激素（thyroid hormone,TH）的主要原料。人体内的碘主要从饮水及食物中获取。碘的摄入缺乏或过量,不仅对TH的合成及分泌有至关重要的影响,而且与甲状腺形态及多种甲状腺疾病的发生、发展及转归密切相关。加碘盐的推广有效预防了碘缺乏可能引起的相关疾病;而碘过量导致的甲状腺疾病谱和发病率的急剧变化,也引起各界高度重视。流行病学调查表明,碘的摄入量与甲状腺功能亢进、自身免疫性甲状腺炎、甲状腺功能减退、甲状腺肿大和甲状腺癌等疾病的发病率密切相关。针对碘的相关生物学问题,对近年来的基础研究和人群研究进行综述,希望借此抛砖引玉,引起相关部门的重视,提高人民群众对碘营养的科学认识水平。     

The use of C14-labelled substrate in histochemical demonstration of different forms of phosphorylase

Active and total phosphorylase activity, using labelled C¹⁴-glucose-1-phosphate as the substrate, is demonstrated by histoautoradiographic method. This method can demonstrate the polysaccharide synthesizedin vitro by phosphorylase without intervention from the unlabelled pre-existing glycogen. C¹⁴-glucose can not replace C¹⁴-glucose-1-phosphate as substrate. The distribution of phosphorylase in tissue sections, except in cases of very low activity, is similar to that obtained by customary dilute Lugol's iodine staining method. The relative difference of intensity between active and total phosphorylase, as revealed by iodine staining, is also reflected by histoautoradiographic method. Histoautoradiographic method has several advantages over the iodine staining method. This method is more sensitive for demonstration of very low phosphorylase activity which may escape detection by iodine staining. Branching enzyme activity, especially when it favors synthesis of glycogen type of polysaccharide instead of amylopectin type, can be better detected by this method. Active phosphorylase substrate medium can be used to demonstrate this activity in plant tissues, where the presence of pre-existing starch often prohibits the use of iodine staining method. Stripping film method for autoradiography is recommended for the study of this enzyme activity.     

Fractionation of iodinated particles and mitochondria from thyroid by zonal centrifugation and a study of their heterogeneity

1. The subcellular particles of horse and rat thyroids were fractionated in a B XIV zonal rotor on a non-linear gradient of Ficoll after labelling with radioactive iodine in vitro (horse) or in vivo (rat). In the horse, the resulting fractions were analysed for radioactive iodine, protein and enzymes representative of certain subcellular particles. In the rat, iodine turnover and thyrotrophin stimulation were studied. 2. The population of iodinated particles could be subdivided into three main classes, characterized by differences in beta-galactosidase and acid phosphatase content and position in the gradient. The presence of a fourth class of particles is suggested. 3. It is concluded that iodinated particles isolated from the thyroid are essentially secondary lysosomes. Their heterogeneity is established with respect to their position in the gradient, their content of acid hydrolases and their iodine turnover. 4. The iodine pools of these secondary lysosomes are increased by thyrotrophin without any change in their number. 5. Their functional significance is discussed. 6. The distribution of mitochondria as judged by succinate dehydrogenase was also studied. The succinate dehydrogenase was spread throughout the gradient with a maximum of activity (40%) in the upper layer of the gradient. Separation of mitochondria from lysosomes by this method was not successful.