Inactivation of foodborne pathogenic and spoilage micro-organisms using ultraviolet-A light in combination with ferulic acid.

The low energy of UV-A (315-400 nm) is insufficient for disinfection. To improve UV-A disinfection technology, we evaluated the effect of ferulic acid (FA) addition on disinfection by UV-A light-emitting diode (LED) (350-385 nm) against various food spoilers and pathogens (seven bacteria and four fungi species). Photoantimicrobial assays were performed at FA concentrations below the MIC. The MIC of the isomerized FA, consisting of 93% cis-form and 7% trans-form, was very similar to that of the commercially available FA (trans-form). Irradiation with UV-A (1·0 J cm-2 ) in the presence of 100 mg l-1 FA resulted in enhanced reducing of all of the tested bacterial strains. A combination of UV-A (10 J cm-2 ) and 1000 mg l-1 FA resulted in enhanced reducing of Saccharomyces cerevisiae and one of the tested filamentous fungi. These results demonstrated that the combination of a short-term application of UV-A and FA at a low concentration yielded synergistic enhancement of antimicrobial activity, especially against bacteria. SIGNIFICANCE AND IMPACT OF THE STUDY: Microbial contamination is one of the most serious problems for foods, fruit and sugar thick juices. UV light is suitable for the nonthermal decontamination of food products by inactivating the contaminating micro-organisms. However, UV-A exposure is insufficient for disinfection. This study demonstrates that the combination of UV-A LED light (350-385 nm), which is not hazardous to human eyes and skin, and ferulic acid (FA), a known phytochemical and food additive, provides synergistic antimicrobial activity against foodborne pathogenic and spoilage micro-organisms. Therefore, FA addition to UV-A light treatment may be useful for improvement of UV-A disinfection technology to prevent food deterioration.

Effects of pressure and local anesthetic tetracaine on dipalmitoylphosphatidylcholine bilayers.

The temperature-pressure phase diagram of dipalmitoylphosphatidylcholine (DPPC) multilamellar vesicles was constructed in the presence of a local anesthetic tetracaine hydrochloride (TC-HCl). The phase-transition temperatures under various pressures were determined by the method of high-pressure light transmission. The temperature of the main transition from the ripple gel (P'(beta)) to the liquid crystal (L(alpha)) phase was depressed by the addition of TC-HCl and elevated by application of pressure up to 150 MPa. The temperature of the pretransition from the lamellar gel (L'(beta)) to the P'(beta) phase was also depressed by the addition of TC-HCl below ca. 10.0 mmol kg(-1) and elevated by the pressure below ca. 50 MPa. Therefore, pressure-anesthetic antagonism for both phase-transitions was confirmed. The pressure-induced interdigitated gel (L(beta)I) phase has been observed under high pressure above 100 MPa in the absence of TC-HCl. The L(beta)I phase is known to be induced also by a variety of small amphiphilic molecules such as ethanol, benzyl alcohol and TC-HCl. In the presence of TC-HCl ranging in concentration up to 20.0 mmol kg(-1), the L(beta)I phase instead of the P'(beta) phase appeared at higher pressure. Present results revealed that pressure facilitates, rather than antagonizes, the effect of TC-HCl on the occurrence of interdigitated gel phase. Furthermore, two regions of two phase coexistence were observed under high pressure in the presence of TC-HCl. One is probably a region of coexisting L(beta)I and L(alpha) phase, which was found between L(beta)I and L(alpha) phases under various pressures. The other is probably a region of coexisting L'(beta) and L(beta)I phase, which was observed in the presence of TC-HCl up to 10.0 mmol kg(-1) at the pressure above 40 MPa and at the temperature below ca. 35 degrees C.

Barotropic phase transitions of dioleoylphosphatidylcholine and stearoyl-oleoylphosphatidylcholine bilayer membranes.

In order to understand the effect of cis unsaturation on the thermotropic and barotropic phase behavior of phospholipid bilayer membranes, the phase transitions of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), 1-stearoyl-2-oleoyl-sn-glycero-3-phosphocholine (SOPC), 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) bilayer membranes were observed by high-pressure optical method. With respect to DOPC bilayer membrane, the so-called main transition between the liquid crystalline (Lalpha) and the lamellar gel (Lbeta) phases was observed in water at above 0 degrees C under high pressure, in addition to the transition between the Lalpha and the lamellar crystalline (L(C)) phases in 50% aqueous ethylene glycol. The pressure of main transition increased linearly with an increase in temperature. Extrapolation of temperature (T)-pressure (P) phase boundary to ambient pressure suggests the temperature of the main transition to be -40.3 degrees C, which has never been found by the DSC method. On the other hand, the temperature of L(C)/Lalpha phase transition in 50% aqueous ethylene glycol was found to be -12.0 degrees C at ambient pressure. The main transition temperatures for DSPC, SOPC and DOPC are 55.6, 6.7 and -40.3 degrees C, respectively, at ambient pressure. The substitution of cis unsaturated chain for saturated chains of DSPC brings about the depression of the main transition temperature by about 48 (+/-1) degrees C for each chain. The volume changes (deltaV) associated with the transitions were calculated from the transition enthalpy (deltaH) and the slope of T-P diagram (dT/dP) by means of the Clapeyron-Clausius equation. The value of deltaV for the main transition of SOPC bilayer membranes was reduced to half the volume change for DSPC bilayers, which means the introduction of the cis double bond in the acyl chain of lipids brings about the reduction of deltaV because of the disordered packing of unsaturated chains in the gel phase of lipid bilayer membranes.

Specific and non-specific binding of long-chain fatty acids to firefly luciferase: cutoff at octanoate.

Firefly luciferase emits a burst of light when the substrates luciferin and ATP are mixed in the presence of oxygen. We (I. Ueda, A. Suzuki, Biophys. J. 75 (1998) 1052-1057) reported that long-chain fatty acids are specific inhibitors of firefly luciferase in competition with luciferin in microM ranges. They increased the thermal transition temperature. In contrast, 1-alkanols of the same carbon chain length inhibited the enzyme non-competitively in mM ranges and decreased the transition temperature. The present study showed that the action of fatty acids switched from specific to non-specific when the carbon chain length was reduced below C8 (octanoate). The fatty acids longer than C10 inhibited the enzyme in microM ranges whereas those shorter than C8 required mM ranges to inhibit it. The longer fatty acids increased whereas shorter fatty acids decreased the transition temperature. The Hill coefficients of longer chain bindings were less than one whereas those of shorter chain were more than one. The shorter fatty acids interacted with the enzyme cooperatively at multiple sites. Binding of the longer fatty acids is limited. Fatty acids longer than C10 are high-affinity specific binders and followed Koshland's induced-fit model. Those shorter than C8 are low-affinity non-specific denaturants and followed Eyring's rate process model. These results contradict the general consensus that the size of the receptor cavity discriminates specific binders.

Barotropic phase transitions and pressure-induced interdigitation on bilayer membranes of phospholipids with varying acyl chain lengths.

The bilayer phase diagrams of a series of 1, 2-diacylphosphatidylcholines containing linear saturated acyl chain (C=13, 14, 15, 16, 17 and 18) were constructed by two kinds of high-pressure optical methods. One is the observation of isothermal barotropic phase transition and the other is the isobaric thermotropic phase transition. The temperature of the main transition from the ripple gel (Pbeta') phase to the liquid crystal (Lalpha) phase for each lipid was elevated by pressure. The slope of the temperature-pressure diagram, dT/dP, was in the range of 0.21-0. 23 K MPa-1 depending on the acyl chain length. The temperature of the pretransition from the lamellar gel (Lbeta') phase to the Pbeta' phase for each lipid was also elevated by pressure. The slope of phase boundary, dT/dP, for the pretransition was in the range of 0. 12-0.14 K MPa-1. Both temperatures of the main and pretransition under ambient pressure increased with an increase in acyl chain length. The chain length dependences of the pretransition and main transition temperatures describe smooth curves with no evidence of odd/even discontinuities. Pressure-induced interdigitated gel (LbetaI) phase was observed beyond 300 MPa for 14:0-PC, 175 MPa for 15:0-PC, 100 MPa for 16:0-PC, 80 MPa for 17:0-PC and 70 MPa for 18:0-PC, respectively. The minimum pressure for the interdigitation of lipid bilayer membranes decreased with an increase in acyl chain length in a manner of non-linear relation. The slopes of phase boundary between Lbeta' and LbetaI phases transformed from the negative slope to the positive slope as the pressure increases.

Effects of age and gender on the expression of brain-derived neurotrophic factor mRNA in rat retrosplenial cortex following administration of dizocilpine.

Using in situ hybridization, we studied the effects of age and gender on the expression of brain-derived neurotrophic factor (BDNF) mRNA and heat shock protein hsp-70 mRNA in the rat retrosplenial cortex following administration of the noncompetitive NMDA receptor antagonist (+)-MK-801 (dizocilpine). Male and female Sprague-Dawley rats (5 weeks, 12 weeks, or 10 months old) were given a single intraperitoneal injection of saline (1 ml/kg) or dizocilpine (0.3, 1.0, or 3.0 mg/kg). No expression of BDNF mRNA and hsp-70 mRNA was detected in the rat retrosplenial cortex after administration of saline (1 ml/kg, IP). Administration of dizocilpine (0.3, 1.0, or 3.0 mg/kg, IP) caused a marked induction of BDNF mRNA and hsp-70 mRNA in the retrosplenial cortex of male and female rats, in a dose-dependent manner. Female rats were more sensitive to the induction of BDNF mRNA and hsp-70 mRNA in the retrosplenial cortex by dizocilpine as compared to male rats. It was also found that adult (12 weeks old) and aged (10 months old) rats were more sensitive to the induction of hsp-70 mRNA and BDNF mRNA in the retrosplenial cortex by dizocilpine as compared to young (5 weeks old) rats. These results suggest that the age and gender differences observed in the expression of BDNF mRNA and hsp-70 mRNA in the retrosplenial cortex by dizocilpine may be associated with the differences in dizocilpine-induced neurotoxicity observed with gender and age within the same region.

Effects of trisodium nitrilotriacetate monohydrate, nitrilotriacetic acid and ammonium chloride on urinary bladder carcinogenesis in rats pretreated with N-bis(2-hydroxypropyl) nitrosamine.

The effects of trisodium nitrilotriacetate monohydrate (Na3 NTA.H2O) nitrilotriacetic acid (H3NTA) and ammonium chloride (NH4Cl) on two-stage urinary bladder carcinogenesis were examined. Carcinogenesis was initiated by administration of 0.2% N-bis (2-hydroxypropyl)-nitrosamine (DHPN) to male Wistar rats in the drinking water for 2 weeks, and then the animals were treated with basal diet containing Na3NTA.H2O, Na3NTA.H2O plus NH4Cl, H3NTA, H3NTA plus NH4Cl, or without these chemicals for 28 weeks. Na3NTA.H2O increased significantly the resultant incidence of neoplastic and preneoplastic lesions of the urinary bladder. Moreover, treatment with Na3NTA.H2O, without the initiation, itself induced papillary or nodular (PN)-hyperplasia. H3NTA produced only a slight increase in the incidence of preneoplastic urinary bladder lesions (PN-hyperplasia) in rats initiated by DHPN, and this was not statistically significant. Elevation of both pH and sodium ion concentration in the urine were correlated with promotion of tumor development. These data showed that Na3NTA.H2O was more effective than H3NTA with regard to promoting potential, and that changes in both urinary pH and concentration of sodium played important roles in enhancement of urinary bladder tumorigenesis by these chemicals.

Promoting effect of 2,4-diaminoanisole sulfate on rat thyroid carcinogenesis.

Two experiments were conducted to examine the effects of 2,4-diaminoanisole sulfate (2,4-DAAS) on thyroid function and carcinogenesis in male Wistar rats. In experiment 1, feeding with 2,4-DAAS resulted in significantly elevated levels of thyroid stimulating hormone (TSH), reduced thyroxine (T4) and triiodothyronine (T3) in the serum, although the latter had almost recovered to normal levels by week 6. However, skin application did not affect serum levels. In experiment 2, administration of 0.5% 2,4-DAAS in the diet for 19 weeks, a week after a single 210 mg/100 g body weight i.p. injection of N-bis(2-hydroxypropyl)nitrosamine (DHPN), significantly increased the incidence and numbers of preneoplastic lesions (focal hyperplasia), adenomas and carcinomas developing in the thyroid gland. Histologically, brown pigment was usually observed within follicular epithelial cells in non-tumorous regions, but not in the tumors themselves.

Increased expression of zif268 mRNA in rat retrosplenial cortex following administration of phencyclidine.

Phencyclidine (PCP) has been shown to cause neurotoxicity in rat retrosplenial cortex following a single administration, although the precise mechanism underlying PCP-induced neurotoxicity is unclear. Using in situ hybridization and immunohistochemistry, we studied the effects of PCP on expression of immediate early gene zif268 mRNA and zif268 protein in the rat brain. High constitutive levels of zif268 mRNA and zif268 immunoreactivity were observed in the brain of control rats. Administration of PCP (12.5, 25 or 50 mg/kg, i.p., 6 h) caused marked induction of zif268 mRNA in the rat retrosplenial cortex, in a dose-dependent manner. However, the basal levels of zif268 mRNA in the other regions of cerebral cortex were decreased by administration of PCP. Emulsion-autoradiographical study suggested that marked expression of zif268 mRNA was observed in the layers III and IV of retrosplenial cortex where the neurotoxicity of PCP was detected. Furthermore, zif268 immunoreactivity in the layer IV of retrosplenial cortex was not changed by administration of PCP (25 mg/kg, i.p., 5 h), but that in the other layers of retrosplenial cortex was reduced by PCP. These results suggest that immediate early gene zif268 may, in part, play a role in the neurotoxicity of NMDA receptor antagonists such as PCP.

A one-step sandwich enzyme immunoassay for human matrix metalloproteinase 8 (neutrophil collagenase) using monoclonal antibodies.

A one-step sandwich enzyme immunoassay (EIA) system for human matrix metalloproteinase 8 (MMP-8, neutrophil collagenase, EC 3.4.24.7) has been established with a pair of monoclonal antibodies prepared against the zymogen of MMP-8 purified from human neutrophils. MMP-8 in samples simultaneously reacted with both solid-phase and peroxidase-labeled antibodies. Sensitivity of this EIA system was 0.34 micrograms/l (5.7 pg/assay) and linearity was obtained between 0.5 and 500 micrograms/l (8.3-8300 pg/assay). The EIA system recognized both precursor and active forms of MMP-8 but not MMP-8 complexed with tissue inhibitors of metalloproteinases. There was no difference in the MMP-8 levels between the plasma samples from patients with rheumatoid arthritis or osteoarthritis and those from healthy subjects (median 6.2 micrograms/l, range 1.5-28 micrograms/l). However, the level in synovial fluids from patients with rheumatoid arthritis (median 345 micrograms/l, range 84-2860 micrograms/l) was shown to be higher than that from osteoarthritic patients. MMP-8 levels in human whole saliva from patients with periodontal diseases (median 282 micrograms/l, range 0-1420 micrograms/l) were also significantly higher than those from clinically healthy subjects (median 25 micrograms/l, range 0-100 micrograms/l). Immunoreactivity analyses showed that MMP-8 species in normal human plasma exists as a precursor but not as a complex form with tissue inhibitor of metalloproteinases (TIMP)-1 or TIMP-2.

Effect of local anesthetics on the bilayer membrane of dipalmitoylphosphatidylcholine: interdigitation of lipid bilayer and vesicle-micelle transition.

The phase transitions of dipalmitoylphosphatidylcholine (DPPC) bilayer membrane were observed by means of differential scanning calorimetry (DSC) as a function of the concentration of local anesthetics, dibucaine (DC x HCl), tetracaine (TC x HCl), lidocaine (LC x HCl) and procaine hydrochlorides (PC x HCl). LC x HCl and PC x HCl depressed monotonously the temperatures of the main- and pre-transition of DPPC bilayer membrane. The enthalpy changes of both transitions decreased slightly with an increase in anesthetic concentration up to 160 mmol kg(-1). In contrast, the addition of TC x HCl or DC x HCl, having the ability to form a micelle by itself, induced the complex phase behavior of DPPC bilayer membrane including the vesicle-to-micelle transition. The depression of both temperatures of the main- and pre-transition, which is accompanied with a decrease in enthalpy, was observed by the addition of TC x HCl up to 21 mmol kg(-1) or DC x HCl up to 11 mmol kg(-1). The pretransition disappeared when these concentrations of anesthetic were added, and the interdigitated gel phase appeared above these concentrations. The appearance of the interdigitated gel phase, instead of the ripple gel phase, brings about the stabilization of the gel phase by 1.8-2.4 kcal mol(-1). In the concentration range of 70-120 mmol kg(-1) TC x HCl (or 40-60 mmol kg(-1) DC x HCl), the enthalpy of the main transition exhibited a drastic decrease, resulting in the virtual disappearance of the main transition. This process includes the decrease in vesicle size with increasing anesthetic concentration, resulting in the mixed micelle of DPPC and anesthetics. Therefore, in this range of anesthetic concentration, the DPPC vesicle solubilized an anesthetic which coexists with the DPPC-anesthetic mixed micelle. Above the concentration of 120 mmol kg(-1) TC x HCl (or 60 mmol kg(-1) DC x HCl), there exists the DPPC-anesthetic mixed micelle. Two types of new transitions concerned with the mixed micelle of DPPC and micelle-forming anesthetics were observed by DSC.

Serum concentrations of basement membrane proteins in NIDDM as a prognostic marker for nephropathy.

Metabolic changes of basement membrane proteins, collagenous and non-collagenous, will be followed by morphological changes and develop into diabetic microangiopathy. In this study, serum concentrations of laminin and type IV collagen were measured in 87 cases o f non-insulin-dependent diabetes mellitus (NIDDM) and examined with the development of diabetic nephropathy and retinopathy. Serum type IV collagen was measured by two methods, assays of the 7S portion (7S-RIA) and of both the 7S and central triple helical domain (IV-EIA). IV-EIA showed more significant elevation with development of diabetic microangiopathy than 7S-RIA and laminin. After measurement of basement membrane proteins, 66 cases were followed clinically for 3 or 4 years. In 45 cases without worsening of diabetic nephropathy, IV-EIA and laminin concentrations in serum were significantly lower than those in 21 cases who showed worsening of diabetic nephropathy in the same period, Multivariate analysis revealed a good model including three dependent variables of serum values of laminin, IV-EIA and mean blood pressure for discriminating between the worsened group and the non-worsened group. High values of IV-EIA and laminin in serum may be useful in predicting the worsening of diabetic nephropathy.

Thermotropic and barotropic phase behavior of dihexadecylphosphatidylcholine bilayer membrane.

The temperature (T)-pressure (P) phase diagram of the ether-linked dihexadecylphosphatidylcholine (DHPC) multilamellar vesicles was constructed by the method of high-pressure optical density. The DHPC membrane at ambient pressure undergoes the pretransition (at 33.6 degrees C) from the interdigitated gel (L beta I) phase to the ripple gel (P' beta) phase, and succeedingly the main transition (at 44.4 degrees C) from the P' beta phase to the liquid crystal (L alpha) phase. Since the slope of the T-P diagram for the pretransition, 0.316 K MPa-1, is larger than that for the main transition, 0.242 K MPa-1, the phase boundary between P' beta and L beta I phases disappeared at high pressure above 130 MPa. A triple point among L beta I, P' beta and L alpha phases was found at 130 MPa and 74.5 degrees C. Difference in phase diagrams between the ether-linked and ester-linked phospholipid bilayer membranes has been elucidated.

Glutathione derivatives enhance adriamycin cytotoxicity in a human lung adenocarcinoma cell line.

We evaluated the effects of a panel of glutathione derivative (S-butyl, S-decyl, S-ethyl, S-heptyl, S-hexyl; S-methyl, S-nonyl, S-octyl, S-propyl and S-pentyl glutathiones) on glutathione-S-transferase activity in the cell lysates of a human lung cancer, PC-9. Glutathione derivatives inhibited glutathione-S-transferase activity in PC-9 cell lysates by up to 67%. When PC-9 cells were incubated with the IC50 concentration of adriamycin (200 nM) and with nontoxic concentrations (1 microM) of the glutathione derivatives, cytotoxicity ranged from -20% to +55% of the control levels. Enhancement of adriamycin toxicity by glutathione derivatives was significantly correlated with the inhibition of glutathione-S-transferase activity. S-decyl-glutathione, which was one of the most potent inhibitors of glutathione-S-transferase activity, significantly enhanced the adriamycin-induced antitumor effect in vivo. Findings suggest that some glutathione derivatives, including the S-decyl, S-octyl, and S-hexyl glutathiones, enhance adriamycin-induced cytotoxicity in part by inhibiting glutathione-S-transferase and that these agents may be useful as chemosensitizers for adriamycin therapy. In conclusion, the present results showed that some glutathione derivatives enhanced sensitivity of tumor cells to ADR by inhibiting GST activity. The use of BSO and EA as sensitizers to chemotherapy is currently being evaluated in clinical trials. The present data suggest that the use of GSH derivatives to modulate GST activity may improve the response to ADR.

Membrane-buffer partition coefficients of a local anesthetic tetracaine monitored by an anesthetic sensor; effects of temperature and pH.

Binding of a local anesthetic tetracaine (TC) to dimyristoylphosphatidylcholine (DMPC) bilayer membrane was studied by the potentiomerty with an ion-selective electrode sensitive to TC cation. DMPC membrane-buffer partition coefficient (K(app)) was determined in mole fraction unit as a function of pH for the lamellar gel (at 12 degrees C), ripple gel (at 20 degrees C), and liquid crystal (at 30 degrees C) phases. The partition coefficients of charged (K+) and uncharged TC (K0) into the DMPC membranes were estimated from the pH-dependence of K(app). The three states of DMPC membranes were more receptive to the uncharged TC than the charged species.

Non-Hodgkin lymphoma and coexisting primary cancers: a retrospective clinical analysis of 10 patients.

The simultaneous occurrence of non-Hodgkin lymphoma (NHL) and primary cancers is rare, and the treatment strategy for both malignancies is unclear. The authors analyzed the clinical records of 10 patients with NHL and coexisting primary cancers. All patients initially had symptoms of NHL, and all carcinomas were found at the initial workup of NHL by chance. The most common primary sites of coexisting cancers were the stomach (six patients) and the colon (two). Histologically, the majority of NHLs were intermediate grade, and all lesions were B-cell type. All primary cancers were adenocarcinoma. Initially, NHL was treated with radiotherapy or chemotherapy. Six primary cancers were resected surgically or endoscopically after the remission of NHL. The remaining four patients received no treatment for primary cancers because of advanced stages or early relapse of NHL. Three patients died of NHL, one died of cancer, and six were still alive, five without evidence of disease and one with disease. The authors conclude that early detection of a coexisting cancer and appropriate treatment after the remission of NHL may increase the possibility of a cure.

Safracins, new antitumor antibiotics. II. Physicochemical properties and chemical structures.

The chemical structures of safracins A and B are proposed to be 1 and 2 respectively on the basis of their physiocochemical properties and spectrometric studies.

Chronic bronchobronchiolitis obliterans associated with Stevens-Johnson syndrome.

We report a case of bronchobronchiolitis obliterans associated with Stevens-Johnson syndrome diagnosed using bronchography and bronchoscopy in a 25-year-old man with dyspnea. Eighteen months prior to admission, the diagnosis of Stevens-Johnson syndrome due to anticonvulsant therapy was made at another hospital. On admission, the patient was wheezing and had overinflation of the lungs, with marked obstructive pulmonary dysfunction. Ventilation and perfusion scintigrams revealed multiple ventilation and perfusion defects but not mismatch. Bronchography revealed central bronchiectasis and obstruction of the peripheral bronchi. Occlusion of the fifth right basal bronchus was observed with fiberoptic bronchoscopy. Bronchography and bronchoscopy provide useful information for the diagnosis of this disease.

Artificial SMA valve for treatment of urinary incontinence: upgrading of valve and introduction of transcutaneous transformer.

This paper is concerned with the development of an artificial urethral valve driven by shape memory alloy actuators, which is attached onto the urethra of a urinary incontinence sufferer for treating the involuntary micturition. Three types of compact cylindrical valves are assembled and their opening and closing functions are examined experimentally. The updated valve is heated and opened by using the transcutaneous energy transformer consisting of a pair of flexible spiral-formed copper wire coils. The experiment using the canine urinary canal verifies that the total system of the valve and the transformer works well as an artificial sphictor muscle and controls the urinary flow through the canal appropriately.

Personal exposure and health effect relationship for NO2 with urinary hydroxyproline to creatinine ratio as indicator.

Establishment of an exposure-effect relationship was attempted between personal nitrogen dioxide (NO2) exposure and urinary hydroxyproline to creatinine ratio of approximately 800 adult women, who were mothers of primary schoolchildren living in two communities around Tokyo. Daily average of the personal NO2 exposure (ENO2) was measured during wintertime by a newly developed personal monitor exposed for 24 hours. The hydroxyproline to creatinine ratio (HOP:C) in the urine sample collected early in the morning of the day for ENO2 measurement was used as a biochemical indicator of the health effect of NO2 exposure. The HOP:C was found to have significant correlation with ENO2 and number of cigarettes smoked actively and passively. ENO2, however, had no correlation with the intensity of the smoking levels; they might affect HOP:C independently. Stepwise multiple regression analysis revealed that HOP:C could be predicted by ENO2 and smoking habits at a high confidence level. The regression analysis of the active smokers' group indicated that a few cigarettes was enough to increase the HOP:C, while in the case of passive smoking, HOP:C increased proportionally to the number of cigarettes.