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To clarify changes of neutrophil functions, mental conditions and relationships among them, 19 male elite long-distance runners participated in this study for 6 months. Examinations, with informed consent, were carried out once a month. According to the results of physical characteristics, it was thought that training intensity was reduced after the main race, Hakone-Ekiden. Neutrophil functions were estimated by indices of reactive oxygen species production, determined by luminol- and lucigenin-dependent chemiluminescence (LmCL and LgCL, respectively) and cytochrome c reduction methods. The peak times (PT) in LmCL and LgCL (LgPT) were most prolonged in January and December, respectively. The peak heights (PH) in LmCL (LmPH) were enhanced in February. Decreased levels of negative categories in the profile of mood state (POMS) questionnaire and the total mood state (TMS) of POMS were observed in February without significance. Correlation analysis using measured values revealed significant negative correlation between LmPH and negative categories in POMS; however, these correlations were possibly a mere appearance, caused by personal differences. After eliminating personal differences, LgPT correlated positively to depression (p

Nitrogen dioxide (NO2) has been extensively studied for its immune modulating effects on pulmonary cells. Alveolar macrophages (AMs) play an important role in pulmonary immunity. The Brown-Norway (BN) rat has been studied as a high-risk model of allergic diseases. In this study, BN rats were exposed to NO2 from the embryonic or weanling period (EP or WP, respectively). To evaluate the effects of NO2 exposure on pulmonary immunity, the activity levels of rat AMs were assessed as reactive oxygen species-generating capacity, measured by a chemiluminescence (CL) technique, and as cytokine-producing ability. Except for 0.2 ppm of NO2 exposure, the CL responses of AMs obtained from the WP group at 12 weeks old were suppressed significantly. Changes of the cytokine-producing levels suggest that inflammatory reactions are terminated at 12 weeks in the EP group. Correlations between the CL responses and the cytokine levels reveal that NO2 exposure may modulate the direction of AM activation. The CL technique is thought to be useful to evaluate changes in AM activity. In this study, the results suggest that, using the high-risk model of allergic diseases, NO2 exposure from the weanling period has stronger effects on AM activity.

A metabolic study to evaluate nutritional balance, in which subjects have to eat all the foods supplied and maintain a set schedule, is thought to be stressful for the subjects. The effects of stress on the immune system have been studied extensively. However, there are no studies of non-specific immunological changes during a metabolic study. Serum opsonic activity (SOA) is a direct and suitable indicator of non-specific humoral immunity. In this study, we used luminol-dependent chemiluminescence (LmCL) to measure reactive oxygen species (ROS) generation from pooled human neutrophils as an indicator of SOA. We also measured serum immunoglobulin levels and plasma myeloperoxidase (MPO) activity. Eleven female college students took part in this 21 day metabolic study after giving their written informed consent. The results obtained suggest that the metabolic study has almost no effect on immunoglobulin levels. According to MPO levels, neutrophils in vivo may be deactivated to some minor extent. In contrast to these results, peak time (PT) and peak height (PH) of LmCL were changed significantly during the metabolic study. In conclusion, SOA increased during the 21 day metabolic study. There was no significant correlation between SOA and serum immunoglobulin levels on any of the study days.

A good deal of data are available on the bowel habits of pre-climacteric females during menstruation. Few studies have examined the same subject in females of climacteric age but who are still menstruating, so the present study was undertaken to examine the bowel habits in menstruating females in this age group. Subjects (n = 246) were residents of a city in northern Japan, aged from 45 to 55 years old and who were still menstruating. Their every-day state of defecation and fecal characteristics were assessed with regard to four parameters: bowel movement frequency, defecation state, fecal appearance and fecal consistency. Based on a perfect match to these four criteria, the subjects were assigned to two groups, the normal group and the constipation group, and changes were assessed by enquiring how their bowel habits differed immediately before and during menses compared with their usual state. Immediately before menstruation, in the constipation group feces became harder and looser in 22.1% and 13.7%, respectively, compared with 8.7% and 9.5% in the normal group. On the other hand, during menstruation in the constipation group, feces became harder and looser in 11.6% and 16.8%, respectively, compared with 5.7% and 8.9% in the normal group. In other words, the constipation group showed a greater change immediately before and during menstruation compared with the normal group, though the differences were not significant. Our data thus suggest that the changes in the bowel habits of women of climacteric age at menstruation are greater for those suffering from constipation than those who are not.

There have been many reports on the applications of low reactive level laser therapy (LLLT) for pain attenuation or pain removal. Our group has reported previously on the effects of in vitro low level laser irradiation (LLLI) particularly on the human neutrophil function using luminol-dependent chemiluminescence (LmCL) for measurement of reactive oxygen species (ROS) production from human neutrophils. However, the change in production by LLLI for each ROS e.g. superoxide, hydroxyl radicals, hydrogen peroxide, singlet oxygen, hypochlorite and so on are however not yet fully understood. In this study, we used luminol-dependent and lucigenin-dependent chemiluminescence (LmCL and LgCL, respectively) to measure the effects of LLLI on the ROS-production process of human neutrophils. Two soluble action stimuli, N-formyl-Met-Leu-Phe (fMLP) and phorbol myristate acetate (PMA) were used to avoid the possible influence of lag-time from recognition to uptake of particles at ROS production. When fMLP was used as the stimulus, the maximum intensity of the chemiluminescence response in LmCL increased but in LgCL it decreased following LLLI. When PMA was used as a stimulus, the times to reach the maximum intensity of the chemiluminescence response of LmCL and LgCL were shortened by LLLI but there was no effect on the maximum intensity of the chemiluminescence response of both. These results suggest that LLLI enhances the ROS production capability, and activates the conversion from superoxide to singlet oxygen and hypochlorite.

In order to evaluate the relationship between reactive oxygen species (ROS) production from human neutrophils and the health of the organism, we developed a new technique and apparatus to measure ROS, and studied the relationship between ROS production and many kinds of environmental factors, such as the circadian rhythm, various degrees of exercise loading and training, diet, alcohol, smoking, and other environmental factors. ROS production and serum opsonic activity did not significantly change at levels of up to 50-60% of VO2max, as advocated by the Japanese Ministry of Health, Labour and Welfare, but in the case of exercise overloading to more than 65% VO2max, ROS production increased. On the other hand, when neutrophils were exposed to environmental factors, e.g., ethanol, minerals, interferon, laser, ultraviolet light and so on, at a low density, ROS production from neutrophils was only slightly enhanced. However, when neutrophils were exposed to the same environmental factors at an excessive density, ROS production significantly decreased. Given that small increases in ROS production from neutrophils are beneficial to the organism, as many researchers have reported, ROS production at low density exposure to environmental factors may increase the autoimmunocompetency of the organism, in other words, it is good for the health. Furthermore, ROS production and serum opsonic activity showed an inverse relationship in many kinds of environment, indicating a mutual compensatory mechanism to maintain the homeostasis of the autoimmune system.

The number of bedridden elderly patients who need a liquid diet has been increasing. We evaluated the usefulness of a tube-fed liquid diet containing cellulose and oligosaccharide for elderly bedridden patients.Twenty-two elderly bedridden patients were given a liquid diet containing cellulose and lafinose for 4 wk. Parameters associated with blood and stool weight were measured during this period.Stool weight increased by 32.0 g in men and 8.8 g in women at 2 wk and remained at these weights up to 4 wk (P0.05 before and after 2 wk, 3 wk, and 4 wk in men, and P0.05 before and after 3 wk in women). Body weight increased in men (0.5 kg) and women (0.7 kg) 15 d after starting the diet and then remained at these values up to 29 d thereafter, and statistically significant differences were not seen. There were no changes in nutritional parameters such as total protein.No significant changes were observed in the general condition of elderly patients, with the exception of a positive increase in fecal weight, by 30-d supplementation of a liquid diet containing cellulose and lafinose.

A full marathon is the longest running race in official track events and is a form of acute exercise. However, no studies have examined the acute neutrophil function response to a competitive marathon race. Thirty-six male athletes who had just completed the 42.195 km course of the 50th Beppu-Oita Mainichi Marathon were enrolled in this study. Neutrophil oxidative burst activity, phagocytic activity and expression of CD11b and CD16 per cell were measured by flow cytometry immediately before and after the marathon. Total leukocyte/neutrophil counts increased significantly (p < 0.001), whereas total oxidative burst activity per neutrophil cell decreased significantly after the race (p < 0.001). Furthermore, total phagocytic activity per neutrophil cell also decreased after the race, although it was not significant (p = 0.08). Although CD11b expression per cell did not change, the expression of CD16 per cell significantly decreased (p < 0.001) after the race. In conclusion, a competitive marathon race decreased neutrophil functions (oxidative burst activity and phagocytic activity), which may be partly due to a decrease in CD16 expression. The increase in total neutrophil counts might reflect a compensatory response to counteract the decrease in neutrophil functions.

Chemiluminescence responses have been used for the evaluation of phagocyte function. In this study, to evaluate effects of training started after maturation on pulmonary immunity, the activity levels of rat alveolar macrophages (AMs) were assessed as reactive oxygen species (ROS) generating capacity, measured by lucigenin- and luminol-dependent chemiluminescence, using a parallel luminometer. One group of male Wistar rats started training at 11 weeks old and another group at 17 weeks old. The experimental period was 12 weeks, and about half of the rats were sacrificed after 6 weeks. The forced and voluntary exercises affect the mean levels of body weights and cell populations in the bronchoalveolar lavage fluid in younger animals; however, the voluntary exercise group in younger animals seemed to adapt after 12 weeks. By contrast, chemiluminescence responses in older rats observed after 6 weeks suggest that AMs are primed, and the maximum releasing activities of ROS are reduced. These changes in AM activity may be caused by the exercise and separation stresses and the rats may adapt to those stressors after 12 weeks. The chemiluminescent technique is thought to be useful to evaluate the changes of AM activity. Copyright © 2002 John Wiley & Sons, Ltd.

To obtain accurate estimates of daily nutrient intake, duplicate food samples should be analyzed chemically. Traditional methods of chemical analysis, however, are impracticable for surveys of numerous samples. This paper describes a new, rapid, and simple method of estimating dietary intake that utilizes a high speed and high performance carbon, hydrogen, and nitrogen autoanalyzer. In the present study, the applicability of the proposed method to surveys in the field was examined using duplicate samples consumed in one day. Protein weights were identical, because nitrogen values obtained by the autoanalyzer were used to calculate protein amounts in food samples. Fat weights estimated by the new method correlated significantly (n = 50; r = 0.979; y = 0.941x − 0.43; p

Evaluations of daily nutrient intakes with practical accuracy contribute not only to public but also to personal health. To obtain accurate estimations of nutrient intake, chemical analyses of a duplicate sample of all foods consumed are recommended. But these analytical methods are expensive, time consuming, and not practically applicable for field surveys dealing with numerous food samples. To solve this problem, a new rapid and simple method of estimating nutrients is developed here. Elemental compositions of cooked foods were examined using a high speed and high performance carbon, hydrogen, and nitrogen autoanalyzer, and the results showed good reproducibility. A significant correlation between Kjeldahl’s and the autoanalyzer methods was observed in the nitrogen measurement (n=20; r =0.999; p< 0.001), and very good agreement was observed between the methods. Therefore, the nitrogen amount obtained by the autoanalyzer was used for the estimation of the protein proportion in the cooked foods. The fat and carbohydrate proportions estimated by the new method correlated with the values obtained by the chemical method (p< 0.001 each). There were also good agreements of fat and carbohydrate proportions between the chemical and the new estimation methods. According to these results, the new, rapid and simple estimation method established in this study should be applicable to nutritional research.

To develop an index to measure alveolar macrophage activity, the fluorescent technique for detection of calcium flux was paid special attention. In this study, a parallel luminescence measuring system was remodelled for fluorescence measurement using a 96-well microplate. The fluorescence indicators widely used to measure cytosolic free calcium ion concentration require excitation at ultraviolet (UV) wavelengths. Instruments to produce UV wavelengths are expensive compared to those used to produce visible wavelengths, and UV wavelengths are potentially injurious to cells. To avoid these problems, Fluo 3 (excitation wavelength in the visible range) was used as the fluorescent dye for detecting calcium ions. The parallel luminometer was remodelled successfully for fluorescent measurement as assessed by the results obtained from the measurements of a common fluorescent dye, fluorescein. Concentrations of free calcium ions were measured using Fluo 3 at 37°C to consider the measurement of calcium flux in living cells. Although a linear relationship between concentrations of free calcium ions and fluorescence were observed, a diminution of fluorescence over time was also observed. To measure calcium flux in living cells, further instrumental and experimental improvements are thus needed. Copyright © 1999 John Wiley & Sons, Ltd.

The principal objective of this paper is to develop a simple and rapid method of estimating levels of fatigue so that chronic fatigue can be prevented. Long-distance runners belonging to a successful corporate team (Group A; 25 males) and representative runners at the prefectural level (Group B; 14 males) participated in this study. We examined the effects of strenuous physical exercise on serum enzyme activity and the fatigue level felt by the runners (subjective fatigue). The following parameters were measured on two consecutive mornings during a training period: physical characteristics, serum-biochemistry using the dry-chemistry method, and subjective fatigue determined using the questionnaire regarding subjective symptoms authorized by the Japan Association of Industrial Health and the Profile of Mood State (POMS). Group A was divided into A-Senior (17 males; highest performance level) and A-Freshman (8 males) subgroups according to the length of employment within the corporation (one year or more and less than one year, respectively). The levels of serum aspartate aminotransferase (AST) and creatine kinase (CK) were significantly lower in the A-Senior group than the other groups and this group displayed the “iceberg” POMS profile at both examinations. Some significant correlations between the elements of POMS and serum enzyme activity levels were observed among all three groups during both examinations. The decline of serum CK levels tended to accompany a decrease in “Fatigue” according to POMS among 9 of 17 members of the A-Senior group. This tendency between the A-Senior and the A-Freshman groups statistically differed according to theX - square analysis. Our results suggest that the effects of physical stress on serum enzyme activity levels and subjective fatigue are affected by performance levels. Physical fatigue seemed to be reflected by serum CK levels. Monitoring subjective fatigue while measuring serum enzyme activity levels using the dry-chemistry method immediately provides clinical value to players and coaches on site, and should therefore help to prevent a shift from “overreaching” to “overtraining”.

In this paper, we examine the effects of SPG, which is a well known BRM, both in vivo and in vitro on the neutrophilic ROS production and the serum opsonic activity by the chemiluminescence technique using luminol as a probe. To investigate the in vivo effects, SPG was administered to 12 healthy male volunteers and two phases of enhancement of the neutrophilic ROS production and the serum opsonic activity was observed. In vitro, the addition of SPG showed a dose-dependent suppression. To investigate the mechanisms in these contradictory effects of SPG, supernatants of a lymphocytes culture medium in the presence of SPG with or without mitogen were added to the neutrophils. The addition of supernatants at a lower concentration of SPG (0.01 mg/ml) with mitogens showed significant preventive effects on the neutrophilic ROS production for the duration of incubation. This suggests that cytokines derived from lymphocytes may contribute to the in vivo effects of SPG. SPG can play an important role in the host's defense against microbe infections by enhancing it's effect on non-specific immunity when administered in vivo.

The principal objective of this paper is basic research on the development of methods for the prevention of chronic fatigue. Overtraining was paid special attention to as one of the types of chronic fatigue condition. Six healthy male participants in a distance-running competition were recruited and used as subjects for this study with informed consent for all examinations. Subjects, that is, participants in the competition, tried to achieve their objective maximum running distance designated before the competition during one month, while in the same period performing their routine work duties. It was reported that the competition caused participants to exhibit an overtrained condition. In this paper, effects of endurance running during one month on blood properties and the fatigue level felt by the subject (subjective fatigue) were studied. Examinations carried out in this research were as follows; 1) physical examination was conducted 3 times during the study; 1 week before the competition, immediately before and 3 weeks after the end of the competition, 2) blood sampling was conducted 4 times; prior to the competition, 1 week after the beginning of and immediately before the end of the competition, and about 3 weeks after the competition, 3) subjective fatigue levels were monitored 8 times; 1 week before and one time in every week during the competition, and 1 week and again 3 weeks after the competition. Immediately before the end of the competition, when subjects might be expected to exhibit the overtrained condition, minor influences on fatigue levels were found. However, effects of the endurance run seemed to remain more than 3 weeks after the competition as indicated by changes of the population of white blood cells and blood hormone levels. The above mentioned results suggest that it is important to determine signs of overtraining at an early stage. To determine overtraining at an early stage, norepinephrine, testosterone and the ratio of testosterone to cortisol may be the most useful indices. These, as well as the element of “drowsiness and dullness” in the questionnaire on subjective symptoms of fatigue authorized by the Japan Association of Industrial Health and the elements of “anger”, “fatigue”, and “confusion” in the Profile of Mood State (POMS) questionnaire are thought to be the most important subjective indices.

It is important to measure superoxide generation in biological samples in research concerning reactive oxygen, because most reactive oxygen species biologically generated are derived from superoxides.This paper reports the establishment of a simultaneous multiple measurement method for detection of superoxides generated from phagocytes, human peripheral neutrophils and rat alveolar macrophages, using cvtochrome c reduction with stirring at 37°C. Using this method, the following results were obtained.1. For more objective presentation of obtained results, velocities of cytochrome c reduction were calculated by the linear regression method at each measurement point and new indicators of cytochrome c reduction, peak value (maximum reduction velocity) and peak time (time when peak value was observed), were introduced.2. Cytochrome c reduction was measured by dual wavelengths, 550nm and 492nm, under the conditions mentioned below.Final sample volume was 300 μl and phagocytes, 1.5 x 10(5) human neutrophils or 3.0 x 10(5) rat macrophages, were mixed with 2.0mg/ml of opsonized zymosan in the presence of 150 μM cytochrome c.3. In the case of human neutrophils, a small amount of cytochrome c reduction was induced by reactions with the microplate surface, whereas in the case of rat macrophages, which can phagocytize materials without opsonization, it was expected that substantial cytochrome c reduction would be observed. However, in comparison with the results of opsonized zymosan stimulation, peak value reached only 40% and peak time was significantly delayed.4. Peak value showed significant linear changes corresponding with changes of the opsonized zymosan concentration and phagocyte numbers when measurements were carried out 37°C.These results suggest that differences of phagocyte activity and serum opsonic activity, which are difficult to measure using previous methods, are clearly detected by the simultaneous measurement method and the indicators introduced in this paper.

To clarify the effects of forced or voluntary exercise on autonomic modulation of the cardiovascular system, we monitored changes in autonomic nervous activity in a mature rat by spectral analysis of the heart rate (HR) during a 10-week training period.Male Wistar rats implanted with a radio-telemetry system were divided into three groups at 18 weeks of age: (1) Control group (n = 8); (2) Voluntary group (n = 6), which were housed separately in a cage with a running wheel; (3) Forced group (n = 6), which were exercised on a treadmill (35 m/min, 15 min/day, 5 days/week). The electrocardiogram was analyzed by the maximum entropy method into two main oscillations, low-frequency (LF) and high-frequency (HF) oscillations, respectively. LF and HF are considered to be markers of both sympathetic and parasympathetic modulations and parasympathetic modulation, respectively.Average running distances of the Voluntary group were more than twofold higher than those of the Forced group. HR levels in the Forced group were lower than those in the Control group. LF and HF levels in the Control and the Forced groups were almost the same during the experiment, and those in the Voluntary group showed a tendency to decrease.The results in the Voluntary and the Forced groups suggest that cardiovascular adjustments are not simply caused by the quantity of exercise. In the Voluntary group, both sympathetic and parasympathetic activity may decrease with a predominance of sympathetic activity. Conversely, in the Forced group, the baroreflex may be hyper-activated by the undesired treadmill running and handling stress.

Although it generally thought thathabitual exercise protects an individual from infections, few careful scientific studies have been conducted. To clarify the influences of physical training on non-specific humoral immunity, both serum opsonic activity, which is a more direct indicator for the strength of non-specific humoral immunity to infections, and serum immunoglobulin and complement levels of 18 healthy male volunteers were assayed before and after a 10-week of training as indices of immuno defense. The serum levels of three immunoglobulins (IgG, IgA and IgM) and one complement (C3) were compared prior to and immediately after exercise both before and after training. Paired t-test revealed that before training exercise-induced increases in IgG and C3 were significant and after training increases in IgG, IgA, IgM and C3 were significant. But baseline (prior to exercise) levels of these immunoglobulins and complement were significantly suppressed during the training period. Serum opsonic activity was compared with each other in the same way as serum protein levels. The noutrophilic chemiluminescence Peak Height (PH), which is one of the indicators of serum opsonic activity, was significantly decreased immediately after exercise at the beginning stage of the training. After the training period, serum opsonic activity showed no noteworthy exercise-induced variations and baseline levels were slightly increased during the training period. These findings suggest that resistance and reactivity to the physical stress are improved and the non-specific humoral immunity, self-defense ability against infections, is considered to be improved by the training.

To clarify changes of neutrophil functions, mental conditions and relationships among them, 19 male elite long-distance runners participated in this study for 6 months. Examinations, with informed consent, were carried out once a month. According to the results of physical characteristics, it was thought that training intensity was reduced after the main race, Hakone-Ekiden. Neutrophil functions were estimated by indices of reactive oxygen species production, determined by luminol- and lucigenin-dependent chemiluminescence (LmCL and LgCL, respectively) and cytochrome c reduction methods. The peak times (PT) in LmCL and LgCL (LgPT) were most prolonged in January and December, respectively. The peak heights (PH) in LmCL (LmPH) were enhanced in February. Decreased levels of negative categories in the profile of mood state (POMS) questionnaire and the total mood state (TMS) of POMS were observed in February without significance. Correlation analysis using measured values revealed significant negative correlation between LmPH and negative categories in POMS; however, these correlations were possibly a mere appearance, caused by personal differences. After eliminating personal differences, LgPT correlated positively to depression (p0.05), anger (p0.05), fatigue (p0.01) and TMS (p0.05). These results suggest that the mean time from the recognition of foreign matter to the maximum production of superoxide from neutrophils is prolonged in the mentally suppressed conditions found under continuous physical training.

There have been many reports on the applications of low reactive level laser (LLL) therapy for pain attenuation or pain removal. Our group has reported previously on the effects of in vitro irradiation of LLL particularly on the phagocytic activity of human neutrophils, using luminol-dependent chemiluminescence (LmCL) for measurement of reactive oxygen species (ROS) production from human neutrophils. But the mechanisms of the attenuation of phagocytic activity of neutrophils by LLL irradiation is not yet full understood. In this study, we used luminol-dependent and lucigenin-dependent chemiluminescence (LgCL) for detection of affected ROS producing process of human neutrophils by LLL irradiation, Two soluble action stimuli, N-formyl-Met-Leu-Phe (fMLP) and phorbol myristate acetate (PMA) were used to avoid the possible influence of lag-time from recognition to uptake of particles at the ROS production. In case of using fMLP as a stimulus, the maximum luminescence intensity of LmCL was increased but LgCL luminescence was decreased by LLL irradiation. When PMA was used as a stimulus, the times to reach the maximum luminescence intensity of LmCL and LgCL were shortened by LLL irradiation but there was no effect on the maximum luminescence intensity of both. These results suggest that LLL irradiation enhances the ROS production activity of human neutrophils by the activation of the superoxide converting system, the active element in which is mainly myeloperoxidase. LLL irradiation enabled a more rapid activation of the superoxide production system, NADPH-oxidase.

In a summer training camp, strenuous physical exercises are carried out to improve endurance ability. The effects of the training stress on the immune system have been studied extensively. However, less attention has been paid to non-specific immunological changes. Serum opsonic activity (SOA) is a more direct and suitable indicator of non-specific humoral immunity. In this study, we used the luminal-dependent chemiluminescence (LmCL) to measure reactive oxygen species generation from pooled human neutrophils as an indicator of SOA. We also measured plasma myeloperoxidase (MPO) and plasma cytokine levels. Twenty-two female college runners took part in this study after giving their written informed consent. There was no significant difference in the physical characteristics and serum enzyme levels. However, significant differences were observed in the changes of blood property and plasma cytokine levels. According to MPO levels, neutrophils in vivo may be deactivated during the camp. Positive correlations between fluctuations of pro- and anti-inflammatory cytokine levels were observed in this study. According to the results of peak time and peak height of LmCL, SOA seems to increase at first qualitatively and then quantitatively during the summer camp. The increased SOA level may compensate for the decrease in neutrophil activity. Copyright © 2009 John Wiley & Sons, Ltd.

The ion chromatographic method is a rapid and reliable technique for the simultaneous determination of inorganic anion concentrations in serum. We are investigating the effects of various diseases on the serum levels of the following inorganic anions: phosphate, bromide, nitrate, and sulphate. In this paper, we tested the intra- and intermeasurement reproducibility of the ion chromatographic method using pooled normal human serum. We found a good intrameasurement reproducibility for all of the above inorganic anions. The intermeasurement reproducibility was good for three of the inorganic anions but not for nitrate. We measured the serum levels of these inorganic anions in 241 inhabitants in one rural area. Using sera of normal inhabitants chosen from the above group, we used the mean value plus or minus two standard deviations as the normal range of each inorganic anion. Our values are in agreement with previously published reports. We found that the serum levels of nitrate and sulphate tended to be increased in inhabitants with liver or kidney disfunction respectively.

Many metal materials are used in clinical medicine and the number used is increasing. However, there are a few papers concerning the biological effects, especially effects of clinically used metals on cellular immunity. In this paper, silver (Ag), nickel (Ni), cobalt (Co), and chromium (Cr), clinically commonly used metals, the effects of which on neutrophilic activity have already reported using the luminol-dependent chemiluminescence (LDCL) method, were examined to determine their influence on the reactive oxygen species (ROS) generating capacity of human neutrophils and on a serum opsonic activity. Silver suppressed the ROS generating capacity of neutrophils in a dose-dependent manner but did not affect the serum opsonic activity. Nickel also suppressed the ROS generating capacity of neutrophils but, at some concentrations, caused the serum opsonic activity to increase. In contrast, Cobalt increased both the ROS generating capacity of neutrophils and the serum opsonic activity. Like silver, chromium did not affect the serum opsonic activity and suppressed the ROS generating capacity of neutrophils. However, the suppression of ROS generating by chromium occurred differently from that of silver. In the cases of nickel and cobalt, effects of these metals on hemolyzing activity of serum complements (CH-50) were measured and consumption of complements was observed at the same metal concentration at which serum opsonic activity was elevated. In this paper, we report that some metals influence not only the ROS generating capacity of neutrophils but also the serum opsonic activity, results thought to be important for more details studies of the effects of metals on cellular immunity.

We have already reported a neutrophil separation method for the multiple simultaneous measurement of neutrophil chemiluminescence. However, when the reported separation method was used, at least 4 ml of venous blood was needed to collect enough neutrophils for chemiluminescent measurement. Because of this blood volume, there is a limitation on applications of the multiple simultaneous method for neonates, infants, and in some clinical situations. To expand the application of this neutrophil chemiluminescence measurement into clinical and health science areas, we have developed a new method for separation of neutrophils from a relatively small amount of blood (500 microliters). In addition, the influences of remaining red blood cells and hemoglobin levels in the neutrophil fraction on the chemiluminescence were examined to determine the necessity for elimination procedures. The new separation method used a capillary tube (length, 130 mm; outside diameter, 5 mm; thickness, 0.8 mm) with density gradient reagents (Histopaque 1077 and 1119). After centrifugal separation (500 g, 30 min), the neutrophil fraction was isolated with 93.1 +/- 4.7% purity and 60.6 +/- 11.1% yield. This purity and yield were comparable to or better than those with the previously reported method, while levels of remaining red blood cells and hemoglobin were about the same. Remaining red blood cells and hemoglobin in the neutrophil fraction acted on the chemiluminescence as a quencher. For the correct estimation of neutrophil chemiluminescence, elimination of remaining red blood cells and hemoglobin in the neutrophil fraction is necessary. This new neutrophil separation method is a very useful method, especially for cases in which available blood amounts are limited.

The purpose of this study was to examine the effect of umpires' intake of water during a summertime baseball game, on serum and urinary biochemical elements, body weight and physiological factors. Twenty-eight umpires were classified into two groups. Group W was composed of 14 umpires who had water while those who did not were group C. Group W had 250 ml of water that was cooled to about 5 degrees C after the 5th inning of the game. The game was played in the following environment: atmosphere temperature was 31 degrees C; wet-bulb temperature was 26 degrees C; blackglove temperature was 40 degrees C; the velocity of the wind was 1.18 m/sec in bright sunshine on a hot and moist day. The results obtained were as follows. In group W, more body weight was lost than in group C, but statistically there was no significant difference. Urinary volume in group W became smaller than in group C, but statistically there was no significant difference. Water in blood increased in group W and decreased in group C but statistically there was significant difference. In both groups, oral temperature decreased, but only in group C was there a statistically significant difference. Although the intake of water increased the volume of perspiration, it did not dilute the concentrated blood, suppress the rise of body temperature, or result in a rise of low blood pressure.

To investigate the in vivo effect of antimicrobial agent on neutrophil chemiluminescence and serum opsonic activity, cefbuperazone sodium (CBPZ) was administered to healthy male volunteers via an intravenous drip. A peripheral blood sample (10 ml) was collected 7 times: prior to injection, then 1, 3, 6, 9, 12 and 24 h after injection. Neutrophil chemiluminescence and serum opsonic activity was measured by the simultaneous multiple measurement system based on luminol-dependent chemiluminescence and indicated by peak height and peak time. Neutrophils were activated at hour 1 from the point of view of peak height and statistical significance was observed. The fluctuation of serum opsonic activity was small and statistical significance was not observed. CBPZ has an in vivo effect on neutrophils which was not supposed from in vitro study.

多量に摂取し, 栄養学的バランスや生体に大きな影響を与えている砂糖やアルコールの広範囲の生体影響をみる目的で, 肥満をおこしにくいFisher系雄ラットを用い, 5% ethanolと15% sucroseの9か月間にわたる慢性投与実験を行ない, 次の結果を得た。1) ethanol投与群, sucrose投与群ともに飲料水中エネルギー量分だけ摂取飼料量を減らす傾向を示し, 調査した投与5か月目, 投与9か月目ともにcontrolとの差がほとんどないほど厳密に摂取エネルギー量を揃えた。またFisher系ラットはsucroseを好み, 5%ethanol投与群が蒸留水と同程度の飲水量であったのに対し, 5% sucrose投与群は2倍以上の飲水量を示した。2) ethanol投与群, sucrose投与群ともに血圧の上昇は示さなかった。3) ヘモグロビン, ヘマトクリットおよびSMACによる20種類の臨床検査を行なったところ, 総ビリルビンとγ-GTPは検出限界以下であった。それ以外の項目でethanol投与群はcontrol群に比べGPTとClの有意な低下とアルカリフォスファターゼ, BUN, Fe, Kの有意な増加を示した。一方, sucrose投与群はcontrol群に比べヘモグロビン, ヘマトクリット, A/G比, GOT, GPT, アルカリフォスファターゼ, 尿酸, 中性脂肪, カルシウムの有意な低下を示した。以上の結果からethanol投与で肝臓と腎臓に障害を示す傾向が, sucrose投与で貧血と全身の活性の低下の傾向が示された。

Even at present there are few experiments dealing with the levels of inorganic anions present in human serum; only chloride and phosphate are determined routinely. The fact that much less attention has been paid to other anions is due to a lack of rapid or convenient analytical methods. A suitable, though novel, method for the determination of anions is ion chromatography. After an appropriate dilution, the sample is directly injected into the ion chromatography without further pretreatment and plural anions are determined simultaneously.In this paper, the authors report on ion chromatography measurements; i. e., on (1) the accuracies of measurements, (2) comparisons with colorimetric assays, and (3) an approach to the quantitative analysis of inorganic anions present in human serum.Standard solutions including equal amounts of 7 anion species fluoride, chloride, nitrite, phosphate, bromide, nitrate, and sulphate, were made. These anions were accurately measured simultaneously, ranging from about 100mg/l to 0.1mg/l. Using river and hot spring water in Oita prefecture, good results were obtained from a comparative study of traditional colorimetric assays and the ion chromatographic methods. The concentrations of phosphate, bromide, nitrate, and sulphate in a standard human serum were determined and these agreed with the values found by other methods.The ion chromatographic method is a rapid and reliable technique for the simultaneous determination of these anions in a small volume of serum. At present the authors are engaged in investigating the effects of various diseases on the levels of inorganic anions.

This paper is intended to give the information we need about personal reactivity to transient loads for the object of estimating human physical fitness. For this purpose, both serum immunoglobulin levels and neutrophilic phagocytosis were assayed as indices to self-defense activities. To determine the later described reproducibility of experimental results, two independent experiments were made on two different groups-one consisting of twelve and the other of fourteen male voluntary students of our college-exercised in the playgroud. The serum levels of three immunoglobulins (IgG, IgA and IgM) and two complements (C3 and C4), prior to and immediately after exercise, were compared with each other. They rose without exception after exercise. Paired T-test revealed that in the group of twelve students, rises in IgG, IgA, C3, α-1-antitrypsin and transferrin were significant at P

The structure, 3-hydroxy-2,3-didehydro-β,χ-caroten-4-one, assigned to tedanin, the main carotenoid pigment of the marine sponge “Tedania digitata (O. Schmidt),” has been confirmed by total synthesis starting with β,χ-carotene. An isomer, 3-hydroxy-2,3-didehydro-β,φ-caroten-4-one, was also synthesized from β,φ-carotene.

The structure, 3-hydroxy-2,3-didehydro-β,χ-caroten-4-one, assigned to tedanin, the main carotenoid pigment of the marine sponge “Tedania digitata (O. Schmidt),” has been confirmed by total synthesis starting with β,χ-carotene. An isomer, 3-hydroxy-2,3-didehydro-β,φ-caroten-4-one, was also synthesized from β,φ-carotene.

Microsomal superoxide anion ( O − 2 ) production was detected using the chemiluminigenic probe, bis-N-Methylacridinium nitrate (lucigenin). Superoxide dismutase (SOD) inhibited 55% of the light emission but in the presence of a detergent (Triton X100) SOD inhibited the light emission by 94%. Lucigenin chemiluminescence from rat liver microsomes supplemented with NADPH was found to be selective and sensitive in detecting the O − 2 production. Treatment of rats with poly IC and LPS resulted in a decrease of the hepatic microsomal cytochrome P450 content by 44% and 37% respectively. The decrease in the cytochrome P450 contents was accompanied by a decrease in LgC1 from the hepatic microsomal fractions by 61% for the poly IC and by 51% for the LPS treated rats. This is the first report to demonstrate that decreased P450 in the presence of normal amounts of cytochrome P450(c) reductase produce correspondingly less O − 2 from the microsomes.

The purpose of this study was to investigate the effects of physical loads, measured physiologically and biochemically, on baseball umpires during a baseball game in hot weather in the summer.These baseball games were held under the high temperature of about 30°C, a high humidity of 70% and high levels of solar radiation with a black-globe temperature of 45°C.The results obtained were as follows.1) The decreases in umpires' body weights immediately after the game were significant without exception, being about 1.19kg (1.7%). This could have been due to perspiration during the baseball game.2) Blood pressures, systolic and diastolic, were observed to have decreased significantly after the game.3) Total protein, albumin, total cholesterol, blood urea nitrogen, uric acid and creatinine in blood were observed to have significantly increased after the game.4) The serum electrolytes sodium, chloride, calcium and phosphorus showed a significant increase after the game, but potassium was observed to have significantly decreased then.5) Protein and creatinine in the urine increased significantly, but the entire urine volume decreased significantly due to perspiration.6) The average energy consumption of the umpire during a baseball game was about half as much as that of a player.The total amount of physical exercise by the former was ‘mild’ in degree as compared with that of the latter.The physical load produced by the umpiring labor presumably resulted mainly from the heat stress brought about by exposure to the summer heat for a long time.7) The physical loads were different depending on the position of the umpire, the umpire-in-chief having the greatest load, followed in descending order by the second-base umpire, the first-base umpire and the third-base umpire.

We have developed a microplate photon counting system based on a cooled charge-coupled device (Lumi Box U-800 II) jointly with Maikurotekku Nition Company (Chiba, Japan). The system makes it possible to quantify chemiluminescence (CL) in a 96-well microplate automatically and simultaneously in a single experiment. We studied the measurement conditions for a luminol-dependent CL assay from neutrophils stimulated with opsonized zymosan (OZ) using this system. Parameters examined included the effect of OZ dose per well, mixing speed, mixing time and detection time on CL responses. The results indicated that this system allows the measurement of CL from phagocytes on a large number of samples using small amounts of sample and regents.