Protoporphyrin IX is an intermediate molecule in the heme biosynthetic pathway. Intra- and extracellular concentrations of protoporphyrin IX in the wild type strain, Myxococcus xanthus DK1622 were measured by reverse phase HPLC. The amount of intracellular protoporphyrin IX continuously increased and reached 6.4 picomoles/mg of protein at the stationary phase. Extracellular protoporphyrin IX began to be detected from the mid-exponential phase. The culture supernatant that was collected in the stationary phase contained approximately 3.0 picomoles of proto-porphyrin IX per mg of protein. Spores formed by nutrient depletion contained about 6.5 picomole protoporphyrin IX/mg of protein. The synthesis of protoporphyrin IX and cell growth were strongly inhibited by addition of succinylacetone to a final concentration of $500\muM$. Succinylacetone, however did not appear to interfere developmental processes. Normal developmental behaviors including aggregation and spore formation was achieved even if succinylacetone was added in a medium. Photolysis among cells grown on a starvation medium supplemented with succinylacetone was also observed. These results indicate that protoporphyrin IX may be important to M. ,xanthus vegetative growth, but not critical to development processes.
For studying chlorophyll biosynthetic heterogeneity of plants, it is necessary to establish a technique for microassay of a putative monovinyl and divinyl protoporphyrin IX. Precise determination of the amounts of monovinyl and divinyl protoporphyrin IX is difficult with optical electronic spectroscopy even at 77$\circ$C. Such a problem could be solved by conversion of protoporphyrin IX to protoporphyrin IX dimethylester with diazomethane and subsequent Mg insertion into protoporphyrin IX dimethylester by reaction with a Grignard solution. The proportion of monovinyl and divinyl Mg-protoporphyrin IX dimethylester formed was measured instead of direct measuring that of protoporphyrin IX by low-temperature spectrofluorometry. The relative proportions of monovinyl and divinyl of protoporphyrin IX, Mg-protoporphyrin IX, and Mgprotoporphyrin IX dimethylester have not changed during the chemical conversion steps. This analysis system could be useful for the study of the monovinyl and divinyl chlorophyll biosynthetic routes in plants.
In this study, the origin of the monovinyl chlorophyll a carboxylic biosynthetic route was investigated in barley (Hordeum vulgare L.) and com (Zea mays L.). Protoporphyrin IX accumulated in vivo or in vitro was found to be all of the divinyl form. Furthermore, the conversion of divinyl protoporphyrin IX to monovinyl protoporphyrin IX in vitro was not observed. In contrast, the biosynthesis and accumulation of monovinyl Mg-protoporphyrin IX and its methyl ester occurred in etiolated leaves and divinyl Mg-protoporphyrin IX was convertible to monovinyl Mg-protoporphyrin IX in vitro. These results suggest that the monovinyl chlorophyll ${\alpha}$ carboxylic biosynthetic route in plants may originate from the divinyl Mg-protoporphyrin IX pool.
The UV-vis absorption spectrum of protoporphyrin IX shows a very sharp and strong absorption maximum peak at 398 nm in acetonitrile-water mixture solution (1:1 v/v). When divalent metal ions such as $Cu^{2+}$, $Zn^{2+}$, and $Ca^{2+}$ ion were added to protoporphyrin IX, metal protoporphyrin IX complexes were thereby produced. Cu-protoporphyrin IX complexes have the largest formation constant ($K_f$) among them. The fluorescence intensity of protoporphyrin IX was diminished by the presence of $Cu^{2+}$, $Zn^{2+}$, $Ca^{2+}$, $Mn^{2+}$, and $Ni^{2+}$ ions as quenchers. However, $Mg^{2+}$, $Mn^{2+}$, and $Ni^{2+}$ ions are hardly combined with protoporphyrin IX. $Mg^{2+}$ ion does not take part in the fluorescence quenching process of protoporphyrin IX in acetonitrile-water mixture solution. According to the Stern-Volmer plots, fluorescence quenching by $Cu^{2+}$, $Zn^{2+}$, and $Ca^{2+}$ ions involves static quenching, which is due to complex formation. On the contrary, dynamic quenching has a large influence on the overall quenching process, when $Mn^{2+}$ and $Ni^{2+}$ ions were added to protoporphyrin IX in acetonitrile-water mixture solution.
The relative state of human iron storage may be ascertained more reliably through determination of the serum iron, iron binding capacity, transferrin saturation and absorption of radioactive iron in conjunction with studies of red cell morphology than from the study of red cell morphology alone. Recent investigations have shown that there is an increase in red cell protoporphyrin concentration in iron deficiency anemia. The significance of the red cell protoporphyrin has been discussed greatly during the years since its discovery. Two of the main factors which appear to influence the amaunt of protoporphyrin are increased erythropoiesis and factors interfering with the utilization of iron in the synthesis of hemoglobin, and iron deficiency. Recently Heller et al. have described a simplified method for blood protoporphyrin assay and this technique could be used assess nutritional iron status, wherein even minor insufficiencies are detectable as increased protoporphyrin concentrations. Based on the evaluation of the relationship between nutritional iron status and red cell protoporphyrin as an index suitable for the detection of the iron deficiency is described in this paper. RESULTS 1. Hemoglobin Concentrations and Anthropometric Measurements. The mean and standard deviations of the various anthropometric measurements of different age and sex groups are shown in table 1. There measurements have been compared with the Korean Standard. In the absence of local standards for arm circumference and skin-fold thickness over triceps, they have been compared with the standard from Jelliffe. Table 2,3, and 4 give anthropometric measurements and frequency (%) of anemia in children surveyed. The mean height of the children studid was 10 to 20 percent; below the Korean Standard. The distribution of height below 80 percent of the Standard was 21.2 percent, however, among anemic group this percentage was 27.7 percent. In general, the mean weight of the children was 10 to 15 percent below the Korean Standard. The percentage of children with weight less than 80 percent of the Standard was about 35 percent. But in the anemic group of the children, this percentage was 44 percent. The mean arm circumference was about 15 percent lower than the Jelliffe's standard. 61.2 percent of the children had values of arm circumference below 80 percent of the standard. Children with low hemoglobin levels, this percentage was 80 percent. The mean skinfold thickness over the triceps of the children studied was about 25 Percent lower than the Jelliffe's standard and 61.2 percent of the children had the value less than 80 percent of the standard. Among anemic children, this percentage was 70.8%. As may be seen from table 5, the mean hemoglobin concentration of the total group was 11.3g/100ml. Hemoglobin concentration was less than 11.0g/100ml. in 65(36.5%) of the 178 children. The degree of anemia in most of these children was mild with a hemoglobin level of less than 8.0g/100ml. found in only one child. In general, the prevalence of anemia was high in female children than male and decreased its frequency with increasing age. Relatively close relationship was observed between hemoglobin level and anthrophometric measurements especially high between arm circumference and skinfold thickness and hemoglobin but very low in height and low in weight and hemoglobin level, estimated by chi-square value. II. Serum iron, Transferrin saturation (1) Serum iron, and transferrin saturation Serum iron, transferrin saturation and red cell protoporphyrin concentrations were estimated in sub-sample of 84 children from 1 to 6 years and 24 older children between 7 and 13 years of age. The findings are presented in table 6. The mean serum iron concentration of the total group was 59ug/100ml. However, the level incrased with age from 36.6ug/100ml. (1-3years) to 80.8ug/100ml. (7-13 years). 60 percent of these children had a serum iron level less than 50ug/10ml. in the 1-3 years age group and 31.4 percent for 4-6 years group. These contrast with the finding of 12.5 percent anemic children in the 7-13 years age group. The mean transferrin saturation for the total group was 18.1 percent and frequency of anemia by transferrin saturation was observed same pattern as serum iron concentration. (2) Red cell protoporphyrin concentrations. (a) Red cell protoporphrin levels of children: Red cell protoporphyrin and other biochemical data are shown in table 4. The mean concentration in red cell of all children was fround 46.3ug/100ml. RBC. and differences with age groups were observed; in the age group 1-3 years, the mean concentration was $59.5{\pm}32.14$ ug/100ml. RBC; 4-6 years $44.1{\pm}22.57$ ug/100ml. RBC. and 7-13 years, $39.0{\pm}13.56$ ug/100ml. RBC. (b) Normal protoporphyrin values in adults: It was observed that in 10 normal adult males studied here the level of protoporphyrin in red cell ranged from 18 to 54 ug/100ml. RBC. and the mean concentration was $47.5{\sim}14.47$ ug/100ml. RBC. Other biochemical determination made on the same subjects are presented in table 8. (c) Red tell protoporphyrin concentration of occupational blood donors: The results of analyses for red cell protoporphyrin as well as serum iron, transferrin saturation and hemoglobin in the 76 blood donors are presented in table 7 and 8. In this experiment, donors were selected at random, however, most of them bled repeatedly because of poor economic situation, I doubt. Table 9 shows the distribution of red cell protoporphyrin concentration and hemoglobin concentration of occupational donors. The mean hemoglobin value for the total was 11.9 g/100 ml. When iron deficiency anemia is defined as a transferrin saturation below 15%, prevalence of anemia was 47.4 percent and the mean serum iron was 27.1ug/100ml. and red cell protoporphyrin, 168.3ug/100ml. RBC. However, mean serum iron and protoporphyrin concentration of above 15% transferrin saturation were 11.6 ug/100 ml. and 58.8 ug/100 ml. RBC. respectively. The mean Protoporphyrin concentration of non-anemic (above 15% transferrin saturation) donors was slightly higher than the results of normal adult males.
Magnesium-protoporphyrin IX (Mg-PPn), which is formed through chelation of protoporphyrin IX (PPn) with Mg ion by Mg chelatase, is the first intermediate for the (bacterio)chlorophyll biosynthetic pathway. Interestingly, Mg-PPn provides peroxidase activity (approximately $4{\times}10^{-2}units/{\mu}M$) detoxifying $H_2O_2$ in the presence of electron donor(s). The peroxidase activity was not detected unless PPn was chelated with Mg ion. Mg-PPn was found freely diffusible through the membrane of Escherichia coli and Vibrio vulnificus, protecting the cells from $H_2O_2$. Furthermore, unlike photosensitizers such as tetracycline and PPn, Mg-PPn did not show any phototoxicity, but rather it protected cell from ultraviolet (UV)-A-induced stress. Thus, the exogenous Mg-PPn could be used as an antioxidant and a UV block to protect cells from $H_2O_2$ stress and UV-induced damage.
Journal of Korean Society of Occupational and Environmental Hygiene
/
v.3
no.2
/
pp.141-151
/
1993
Blood samples obtained from 200 adults who had visited the "S" general hospital were analyzed to compare the zinc protoporphyrin (ZPP) levels quantified by high performance liquid chromatograph (HPLC) and by hematofluorometer (HF) to investigate the methodological difference if any and the relationship between the levels of blood lead and ZPP among no-lead exposed adults. Also investigated were the distribution of ZPP and protoporphyrin IX (PPIX) concentrations, the establishment of normal levels of blood ZPP and blood lead, and the contribution of age and sex factors to these values. These subjects had no previous occupational exposure to lead. The results obtained were as follows : 1. The mean values of blood lead for male and female subjects were $9.46{\pm}2.44{\mu}g/dl$ and $8.09{\pm}2.17{\mu}g/dl$, respectively. The difference observed in the mean concentrations between male and female subjects was statistically very significant. 2. The mean values of blood ZPP by HPLC for male and female subjects were $15.94{\pm}4.55{\mu}g/dl$ and $22.26{\pm}6.61{\mu}g/dl$, respectively. The difference observed in the mean concentrations between male and female subjects was statistically not significant. The mean values of blood PPIX by HPLC for male and female subjects were $2.51{\pm}1.78{\mu}g/dl$ and $2.81{\pm}1.56{\mu}g/dl$, respectively. The difference observed in the mean concentrations between male and female subjects was statistically not significant. 3. The mean values of blood ZPP by HF for male and female subjects were $28.44{\pm}7.11{\mu}g/dl$ and $37.77{\pm}8.04{\mu}g/dl$, respectively. The difference observed in the mean concentrations between male and female subjects was statistically very significant. 4. No statistically significant correlation was found between the levels of blood ZPP and blood lead. 5. The ratio of ZPP and protoporphyrin IX (PPIX) concentration to erythrocyte protoporphyrin (EP, EP=ZPP+PPIX) concentration was 87.4% and 12.6%, respectively. 6. A statistically very significant correlation was found between the ZPP concentrations determined by HPLC and the values by HF (r=0.7565). The ZPP concentraitons quantified by HF were 1.75 times as high as the values obtained by HPLC. 7. The blood ZPP concentrations quantified by HPLC, HF, and spectrofluorometer (SF) from the blood samples obtained from 14 lead-exposed workers and from 16 no-lead exposed adults showed wide variations. The ZPP concentrations by HF were the highest followed by the levels obtained by SF and by HPLC. In the exposed group, no statistically significant difference was found among three methods of quantifying blood ZPP levels. In the no-lead exposed group, however, statistically significant difference was observed among these methods. The ZPP concentrations by HF were about twice as high as those of by HPLC or by SF. Among three methods of quantifying blood ZPP (HPLC, SF and HF), the results revealed significant difference. Therefore it is suggested that objective methods of quantifying blood ZPP and a system of correcting different ZPP levels be developed by the ministry of Labor.
Journal of Korean Society of Occupational and Environmental Hygiene
/
v.3
no.1
/
pp.62-67
/
1993
This study aims at investigating the relationships between the urinary mercury concentration and blood zinc-protoportphyrin, serum cholinestrase activity, making 149 workers exposed to mercury vapor and 68 workers who were not exposed to mercury among the workers in a flurorescent lamp manufactureing factory an object of this investigation. The results are as follows ; 1. In an exposed group the number of those whose urinary mercury concentration showed over $100{\mu}g/l$ was 21 persons (14.3%) among 147 workers. The average urinary mercury concentration was $52.1{\pm}46.1{\mu}g/l$($1.8-361.2{\mu}g/l$), which proved to be higher than the average concentration in a control group. 2. In an exposed group, the average concentration of blood zinc-protoporphyrin was $27.8{\pm}12.5{\mu}g/dl$($12.2-101.5{\mu}g/dl$), which proved to be somewhat higher than the average concentration in a control group. But it did not show a significant difference. 3. In an exposed group, the average concentration of serum cholinesterase activity showed $1936.7{\pm}341.0IU/l$(1,120,0-2,8750IU/l), which proved to be lower than the average concentration in a control group. 4. The relational coefficient between urinary mercury concentration and blool zinc-protoporphyrin, serum cholinesterase activity of the whole workers exposed to mercury showed little difference. While the relational coefficient between the urinary mercury concentration and blood zinc-protoporphyrin of the workers whose urinary mercury concentration showed over $100{\mu}g/l$ was relatively high, which was 0.62.
The effects of S-23142{N-(4-chloro-2-fluoro-5-propargyloxyphenyl)-3, 4, 5, 6-tetrahydrophtalimide}, on protoporphyrin IX(PPIX) biosynthesis in Spinacia oleracea L, leaf in vivo and in vitro condition were investigated by reversed-phase HPLC with fluorescence detector. The stroma and the membrane fraction of spinach chloroplast were isolated by osmotic regulation. The conversion of ${\delta}$-aminolevulinic acid(ALA) to PPIX occured more in the stroma than in the membrane fraction. It suggested that the enzymes that catalyse PPIX biosynthesis from ALA were localized in the stroma. Also, the synthesized PPIX content from ALA was completely inhibited by $10^{-8}M$ of S-23412 or $10^{-7}M$ of acifluorfen in the stroma but not in the membrane fractions. Therefore, these results suggested that the target site of S-23142 and acifluorfen may exist in the stroma fraction of spinach chloroplast.
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