Curcuma longa extract reduces serum inflammatory markers and postprandial hyperglycemia in healthy but borderline participants with overweight and glycemia in the normal/prediabetes range: a randomized, double-blind, and placebo-controlled trial.

The spice turmeric, which has the Latin name Curcuma longa (C. longa), has various physiological effects. This study evaluated the effects of a hot water mixture with supercritical carbon dioxide C. longa extracts, CLE, and the potential active components of C. longa, turmeronols A and B and bisacurone on inflammation and glucose metabolism. First, we investigated the effect of CLE and the potential active components of C. longa on lipopolysaccharide-induced inflammation in RAW264.7 macrophages. We found a significant decrease in the production of interleukin (IL)-1ß, IL-6, tumor necrosis factor (TNF)-α, and nitric oxide with CLE, turmeronol A, and bisacurone, Significant inhibition of each of these substances was also observed, except for TNF-α with turmeronol B. The second part of our work was a 12-week randomized, double-blind, placebo-controlled study in healthy but borderline adults aged 40 to 69 years with overweight and normal/prediabetes glycemia. We compared blood inflammatory and glycometabolic markers in the CLE (n = 55) and placebo groups (n = 55). We found significantly lower serum high-sensitivity C-reactive protein and hemoglobin A1c levels in the CLE group. This group also showed significant improvements in postprandial hyperglycemia and insulin sensitivity indices. Our findings indicate that CLE may reduce low-grade inflammation and thus improve insulin sensitivity and postprandial hyperglycemia. Clinical trial registration: https://center6.umin.ac.jp/cgi-open-bin/ctr/ctr_view.cgi?recptno=R000051492, UMIN-CTR, UMIN000045106.

Turmeronols (A and B) from Curcuma longa have anti-inflammatory effects in lipopolysaccharide-stimulated BV-2 microglial cells by reducing NF-κB signaling.

Turmeronols (A and B), bisabolane-type sesquiterpenoids found in turmeric, reduce inflammation outside the brain in animals; however, their effects on neuroinflammation, a common pathology of various neurodegenerative diseases, are not understood. Inflammatory mediators produced by microglial cells play a key role in neuroinflammation, so this study evaluated the anti-inflammatory effects of turmeronols in BV-2 microglial cells stimulated with lipopolysaccharide (LPS). Pretreatment with turmeronol A or B significantly inhibited LPS-induced nitric oxide (NO) production; mRNA expression of inducible NO synthase; production of interleukin (IL)-1ß, IL-6, and tumor necrosis factor α and upregulation of their mRNA expression; phosphorylation of nuclear factor-κB (NF-κB) p65 proteins and inhibitor of NF-κB kinase (IKK); and nuclear translocation of NF-κB. These results suggest that these turmeronols may prevent the production of inflammatory mediators by inhibiting the IKK/NF-κB signaling pathway in activated microglial cells and can potentially treat neuroinflammation associated with microglial activation.

A Hot Water Extract of Curcuma longa L. Improves Fasting Serum Glucose Levels in Participants with Low-Grade Inflammation: Reanalysis of Data from Two Randomized, Double-Blind, Placebo-Controlled Trials.

The dietary spice Curcuma longa L. (C. longa), also known as turmeric, has various biological effects. A hot water extract of C. longa was shown to have anti-inflammatory activities in preclinical and clinical studies. Chronic low-grade inflammation is associated with the disruption of glucose homeostasis, but the effect of C. longa extract on glucose metabolism in humans is poorly understood. Therefore, we investigated the effect of C. longa extracts on serum glucose levels in the presence of low-grade inflammation. We reanalyzed our published data from two randomized, double-blind, placebo-controlled trials in overweight participants aged 50 to 69 years and performed a stratified analysis using the inflammatory marker high-sensitivity C-reactive protein (hsCRP). In both studies, participants took a test food with a hot water extract of C. longa (C. longa extract group, n = 45 per study) or without C. longa extract (placebo group, n = 45 per study) daily for 12 weeks, and we measured the levels of serum hsCRP and fasting serum glucose. The mean baseline hsCRP value was used to stratify participants into two subgroups: a low-hsCRP subgroup (baseline mean hsCRP < 0.098 mg/dL) and a high-hsCRP subgroup (baseline mean hsCRP ≥ 0.098 mg/dL). In the low-hsCRP subgroup, we found no significant difference in fasting serum glucose levels between the two groups in either study, but in the high-hsCRP subgroup, the C. longa extract group had significantly lower levels of serum hsCRP (p < 0.05) and fasting serum glucose (p < 0.05) than the placebo group in both studies. In conclusion, a hot water extract of C. longa may help to improve systemic glucose metabolism in people with chronic low-grade inflammation.

Curcuma longa extract improves serum inflammatory markers and mental health in healthy participants who are overweight: a randomized, double-blind, placebo-controlled trial.

BACKGROUND: The dietary spice Curcuma longa, also known as turmeric, has various biological effects. Both a water extract and a supercritical carbon dioxide extract of C. longa showed anti-inflammatory activities in animal studies. However, the anti-inflammatory effect in humans of a mixture of these two C. longa extracts (CLE) is poorly understood. Therefore, we investigated the effect of CLE containing anti-inflammatory turmeronols on chronic inflammation and general health. METHODS: We performed a randomized, double-blind, placebo-controlled study in healthy subjects aged 50 to 69 years with overweight. Participants took two capsules containing CLE (CLE group, n = 45) or two placebo capsules (placebo group, n = 45) daily for 12 weeks, and serum inflammatory markers were measured. Participants also completed two questionnaires: the Medical Outcomes Study (MOS) 36-Item Short-Form Health Survey (SF-36) and the Profile of Mood States (POMS) scale. Treatment effects were analyzed by two way analysis of variance followed by a t test (significance level, p <  0.05). RESULTS: After the intervention, the CLE group had a significantly lower body weight (p <  0.05) and body mass index (p < 0.05) than the placebo group and significantly lower serum levels of C-reactive protein (p < 0.05) and complement component 3 (p < 0.05). In addition, the CLE group showed significant improvement of the MOS SF-36 mental health score (p < 0.05) and POMS anger-hostility score (p < 0.05). CONCLUSION: CLE may ameliorate chronic low-grade inflammation and thus help to improve mental health and mood disturbance. TRIAL REGISTRATION: UMIN-CTR, UMIN000037370. Registered 14 July 2019, https://upload.umin.ac.jp/cgi-open-bin/ctr/ctr_view.cgi?recptno=R000042607.

Correction: Turmeronol A and turmeronol B from Curcuma longa prevent inflammatory mediator production by lipopolysaccharide-stimulated RAW264.7 macrophages, partially via reduced NF-κB signaling.

Correction for 'Turmeronol A and turmeronol B from Curcuma longa prevent inflammatory mediator production by lipopolysaccharide-stimulated RAW264.7 macrophages, partially via reduced NF-κB signaling' by Chinatsu Okuda-Hanafusa et al., Food Funct., 2019, 10, 5779-5788. DOI: 10.1039/C9FO00336C.

Hot Water Extract of Curcuma longa L. Improves Serum Inflammatory Markers and General Health in Subjects with Overweight or Prehypertension/Mild Hypertension: A Randomized, Double-Blind, Placebo-Controlled Trial.

To investigate the effect of a hot water extract of C. longa L. (WEC) containing anti-inflammatory agents, bisacurone, and turmeronol on chronic inflammation, a randomized double-blind placebo-controlled study was conducted in middle-aged and elderly subjects aged 50-69 years with overweight or prehypertension/mild hypertension. The subjects consumed 900 mg WEC tablets, containing 400 µg bisacurone, 80 µg turmeronol A and 20 µg turmeronol B (WEC group: n = 45), or placebo tablets without WEC (placebo group: n = 45) daily for 12 weeks. Serum inflammatory and metabolic markers were measured. The subjects also completed the MOS 36-item short-form health survey (SF-36) and the Profile of Mood States scale (POMS). In the WEC group, the serum levels of C-reactive protein, tumor necrosis factor-α, interleukin-6, and soluble vascular cell adhesion molecule-1 decreased significantly. Compared with the placebo group, the WEC group had significantly lower serum levels of glucose, hemoglobin A1c, and triglycerides, as well as higher serum levels of high-density lipoprotein cholesterol. The WEC group also showed significant improvement of SF-36 scores (for general health, vitality, mental health, and mental summary component) and POMS scores for positive mood states (vigor-activity and friendliness). In conclusion, WEC may ameliorate chronic low-grade inflammation, thus contributing to the improvement of associated metabolic disorders and general health.

Turmeronol A and turmeronol B from Curcuma longa prevent inflammatory mediator production by lipopolysaccharide-stimulated RAW264.7 macrophages, partially via reduced NF-κB signaling.

Chronic inflammation depends on inflammatory mediators produced by activated macrophages and is the common pathological basis for various diseases. Turmeronol is a sesquiterpenoid found in the spice turmeric (Curcuma longa), which is known to have anti-inflammatory activity. To elucidate the anti-inflammatory mechanism of turmeronol, we investigated the influence of turmeronol A and turmeronol B in mouse macrophages (RAW264.7 cells) stimulated with lipopolysaccharide (LPS). Pretreatment of RAW264.7 cells with either turmeronol A or B significantly inhibited LPS-induced production of prostaglandin E2 and nitric oxide, as well as expression of mRNAs for the corresponding synthetic enzymes. In addition, the turmeronols significantly inhibited LPS-induced upregulation of interleukin-1ß, interleukin-6, and tumor necrosis factor-α at the mRNA and protein levels. Both turmeronols also inhibited nuclear translocation of nuclear factor κB (NF-κB), with a similar time course to the NF-κB inhibitor pyrrolidine dithiocarbamate, but not curcumin (another NF-κB inhibitor). Thus, both turmeronols prevented activation of macrophages and inflammatory mediator production, possibly by suppressing activation of NF-κB, and therefore have potential for use in preventing chronic inflammatory diseases.

Effects of hot water extract of Curcuma longa on human epidermal keratinocytes in vitro and skin conditions in healthy participants: A randomized, double-blind, placebo-controlled trial.

BACKGROUND: Curcuma longa has been reported to have anti-inflammatory effects. Skin inflammation impairs skin functions. OBJECTIVES: Our aim was to investigate the effect of a hot water extract of C longa (WEC) on skin conditions in cell studies using keratinocytes and in clinical trials. METHODS: We measured proinflammatory cytokine levels in ultraviolet B-irradiated keratinocytes in the presence or absence of WEC. The effects of WEC on hyaluronan production in keratinocytes were also determined. In a randomized, double-blind, placebo-controlled trial, 47 healthy participants were assigned to 8-week intervention groups with daily intakes of WEC with or without curcumin or a placebo. The water content and transepidermal water loss in the face and minimal erythema dose on the back after ultraviolet B irradiation were evaluated every 4 weeks. RESULTS: Hot water extract of C longa significantly inhibited increases in ultraviolet B-induced tumor necrosis factor α and interleukin 1ß at the mRNA and protein levels. WEC also significantly increased hyaluronan production from nonstimulated keratinocytes. In the randomized, double-blind, placebo-controlled trial, increases from baseline in the water content of the face were significantly greater at weeks 4 and 8 in the WEC group, but not in the WEC + curcumin group, than in the placebo group. There were no significant differences in transepidermal water loss and minimal erythema dose among the groups. CONCLUSIONS: The cell studies confirmed that WEC has anti-inflammatory effects and augments hyaluronan production in the skin. The results of clinical trials suggest that WEC may be useful for moisturizing facial skin. TRIAL REGISTRATION: UMIN Clinical Trials Registry 000028510. Retrospectively registered.

Inhibitory effect of the compounds from the water extract of Curcuma longa on the production of PGE2 and NO in a macrophage cell line stimulated by LPS.

We wished to search for the compounds contributing to the anti-inflammatory effects of the water extract of Curcuma longa (WEC). WEC was fractioned and the fractions were evaluated with regard to their inhibitory effect on the production of nitric oxide (NO) from the macrophage cell line stimulated by lipopolysaccharide. Compounds in the active fractions were isolated and identified. One isolated compound was identified as new: (6S)-2-methyl-5-hydroxy-6-(3-hydroxy-4-methylphenyl)-2-heptene-4-one (1). Four isolated compounds were identified as known: (6S)-2-methyl-6-(4-hydroxyphenyl)-2-heptene-4-one (4), bisabolone-4-one (5), curcumenone (6), and turmeronol A (8). Three isolated compounds were not identified their stereostructures but their planar structures: 2-methyl-6-(4-hydroxymethyl-phenyl)-2-heptene-4-one (2), 2-methyl-6-(2,3-epoxy-4-methyl-4-cyclohexene)-2-heptene (3), and 4-methylene-5-hydroxybisabola-2,10-diene-9-one (7). Compounds 1, 4, 7 and 8 inhibited production of prostaglandin E2 and NO. Others inhibited NO production only. These results (at least in part) show the active compounds contributing to the anti-inflammatory effects of WEC, and may be useful for elucidating its various beneficial physiologic effects.

Effect of a water extract of Curcuma longa on emotional states in healthy participants.

Physiological symptoms of mood disturbances, such as fatigue or anxiety, are closely related to inflammation in the central nervous system or the whole body. Curcuma longa is widely used as a dietary spice and has been reported to have anti-inflammatory activity. To investigate the effect of a water extract of C. longa (WEC) on emotional states, a randomized, double-blind, placebo-controlled, parallel-group study was conducted with healthy participants. Forty-eight participants were randomly assigned to receive five tablets containing 150 mg WEC and 0.40 mg bisacurone (L-WEC group), five tablets containing 900 mg WEC and 2.40 mg bisacurone (H-WEC group), or matching placebo tablets (placebo group) daily for 8 weeks. Participant emotional states were measured every 4 weeks using the Profile of Mood States (POMS). The changes from week 0 to week 8 in the fatigue score of the POMS were significantly lower in the L-WEC group than in the placebo group. This result suggests that daily intake of 150 mg WEC may positively influence emotional fatigue, and further investigation focused on emotional fatigue is needed.

A hot water extract of turmeric (Curcuma longa) suppresses acute ethanol-induced liver injury in mice by inhibiting hepatic oxidative stress and inflammatory cytokine production.

Turmeric (Curcuma longa) is a widely used spice that has various biological effects, and aqueous extracts of turmeric exhibit potent antioxidant activity and anti-inflammatory activity. Bisacurone, a component of turmeric extract, is known to have similar effects. Oxidative stress and inflammatory cytokines play an important role in ethanol-induced liver injury. This study was performed to evaluate the influence of a hot water extract of C. longa (WEC) or bisacurone on acute ethanol-induced liver injury. C57BL/6 mice were orally administered WEC (20 mg/kg body weight; BW) or bisacurone (60 µg/kg BW) at 30 min before a single dose of ethanol was given by oral administration (3·0 g/kg BW). Plasma levels of aspartate aminotransferase and alanine aminotransferase were markedly increased in ethanol-treated mice, while the increase of these enzymes was significantly suppressed by prior administration of WEC. The increase of alanine aminotransferase was also significantly suppressed by pretreatment with bisacurone. Compared with control mice, animals given WEC had higher hepatic tissue levels of superoxide dismutase and glutathione, as well as lower hepatic tissue levels of thiobarbituric acid-reactive substances, TNF-α protein and IL-6 mRNA. These results suggest that oral administration of WEC may have a protective effect against ethanol-induced liver injury by suppressing hepatic oxidation and inflammation, at least partly through the effects of bisacurone.

Oral intake of a combination of glucosyl hesperidin and caffeine elicits an anti-obesity effect in healthy, moderately obese subjects: a randomized double-blind placebo-controlled trial.

BACKGROUND: We have previously shown that a combination of glucosyl hesperidin (G-hesperidin) plus caffeine reduces accumulation of body fat, whereas G-hesperidin or caffeine alone shows little effect on high-fat diet-induced obesity in mice. The aim of this study is to evaluate the anti-obesity effect of G-hesperidin plus caffeine on body fat and serum TG in healthy subjects with moderately high body mass index (BMI) and serum TG. Since we considered that there are individual differences in caffeine sensitivity, we conducted dose-finding study of caffeine combined with G-hesperidin. METHODS: Seventy-five healthy subjects with moderately high BMI (24-30 kg/m(2)) and serum TG (100-250 mg/dl) were divided and assigned to 12-week intervention with daily intakes of 500 mg of G-hesperidin with or without 25, 50, or 75 mg of caffeine, or placebo in a randomized double-blind placebo-controlled design . RESULTS: After intervention, decreases in abdominal fat area (AFA), especially subcutaneous fat area (SFA), were significantly greater in the G-hesperidin with 50-mg caffeine group (AFA:-8.4 ± 21.9 v.s. 16.3 ± 34.1 cm(2); p < 0.05, SFA: -9.3 ± 17.1 v.s. 11.2 ± 18.3 cm(2); p < 0.01) and in the G-hesperidin with 75-mg caffeine group (AFA:-17.0 ± 31.4 v.s. 16.3 ± 34.1 cm(2); p < 0.01, SFA: -12.4 ± 18.7 v.s. 11.2 ± 18.3 cm(2); p < 0.01) than in the placebo group. Fat-decreasing effects of G-hesperidin were enhanced dose-dependently by caffeine addition. BMI decreases were significantly greater in the G-hesperidin with 75-mg caffeine group than in the placebo group (-0.56 ± 0.74 v.s. -0.02 ± 0.58 kg/m(2); p < 0.05). G-hesperidin with/without caffeine had no effect on serum TG (p > 0.05 v.s. placebo). CONCLUSIONS: These data suggested that a combination of 500-mg G-hesperidin with 50- or 75-mg caffeine may be useful for the prevention or treatment of obesity. TRIAL REGISTRATION: UMIN Clinical Trials Registry 000019241 .

A hot water extract of Curcuma longa inhibits adhesion molecule protein expression and monocyte adhesion to TNF-α-stimulated human endothelial cells.

The recruitment of arterial leukocytes to endothelial cells is an important step in the progression of various inflammatory diseases. Therefore, its modulation is thought to be a prospective target for the prevention or treatment of such diseases. Adhesion molecules on endothelial cells are induced by proinflammatory cytokines, including tumor necrosis factor-α (TNF-α), and contribute to the recruitment of leukocytes. In the present study, we investigated the effect of hot water extract of Curcuma longa (WEC) on the protein expression of adhesion molecules, monocyte adhesion induced by TNF-α in human umbilical vascular endothelial cells (HUVECs). Treatment of HUVECs with WEC significantly suppressed both TNF-α-induced protein expression of adhesion molecules and monocyte adhesion. WEC also suppressed phosphorylation and degradation of nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, alpha (IκBα) induced by TNF-α in HUVECs, suggesting that WEC inhibits the NF-κB signaling pathway.

A combination of glucosyl hesperidin and caffeine exhibits an anti-obesity effect by inhibition of hepatic lipogenesis in mice.

To develop an anti-obesity agent, we examined the combination effect of glucosyl hesperidin (G-hesperidin) and caffeine on obesity in mice. High-fat diet-induced obese KK mice were fed a low-fat diet with or without G-hesperidin, caffeine, or their combination for 2 weeks. Decreases in body weight and significantly lower adipose tissue weight were observed in the combination-fed mice but not in the G-hesperidin-fed or caffeine-fed mice. DNA microarray analysis of mouse liver suggested that the feeding of G-hesperidin + caffeine was associated with lower lipogenesis. Therefore, we examined the anti-lipogenic effect of G-hesperidin + caffeine in fasted-refed KK mice. Hepatic triglyceride levels were significantly lower in the mice fed G-hesperidin + caffeine during the refeeding period but not in the mice fed each alone. In addition, hepatic expressions of genes related to lipogenesis, such as sterol regulatory element-binding protein-1c or fatty acid synthase, were significantly lower in the mice fed G-hesperidin + caffeine compared with that in the control mice. These results suggested that G-hesperidin + caffeine is effective for controlling obesity partly by the inhibition of hepatic lipogenesis.

A nonradioisotope, enzymatic microplate assay for in vivo evaluation of 2-deoxyglucose uptake in muscle tissue.

To determine 2-deoxy-D-glucose (2DG) and 2-deoxy-D-glucose 6-phosphate (DG6P) in mouse tissue after injection of 2DG, we have developed a novel assay. This assay is a simple procedure involving incubation of samples with four independent, single reaction mixtures followed by measurement of fluorescence. From differences between the values obtained with the four reactions, each of glucose, glucose 6-phosphate, 2DG and DG6P were able to be quantified in a sensitive manner. Using this assay system, glucose and 2DG in blood and DG6P-accumulation in muscle were easily determined. Therefore, this assay may be useful for measuring in vivo glucose uptake without the use of radioisotopes.

A combination of caffeine, arginine, soy isoflavones, and L-carnitine enhances both lipolysis and fatty acid oxidation in 3T3-L1 and HepG2 cells in vitro and in KK mice in vivo.

To develop an anti-obesity agent containing dietary components, we focused on the mechanisms that enhance both lipolysis and fatty acid oxidation. Caffeine and arginine (CA), a nonselective adenosine-receptor antagonist and an inducer of lipolytic hormone, respectively, were used to stimulate lipolysis. Soy isoflavones and L-carnitine (SL), stimulators of carnitine palmitoyl transferase 1A and a cofactor for beta-oxidation of fatty acids, respectively, were used to enhance fatty acid oxidation. Effects of a combination of CA and SL (CASL) on lipid metabolism were studied in vitro and in vivo. During 3T3-L1 differentiation, lipid accumulation was significantly lower in cells treated with CASL (50 micromol/L, 1 mmol/L, 1 micromol/L, and 1 mmol/L, respectively) compared with each alone. Lipolysis was also significantly greater in 3T3-L1 adipocytes treated with CASL (50 micromol/L, 1 mmol/L, 10 micromol/L and 0.5 mmol/L, respectively) compared with each alone. In addition, treatment with higher concentrations of CASL (2 mmol/L, 1 mmol/L, 10 micromol/L, and 1 mmol/L, respectively) significantly enhanced beta-oxidation in HepG2 cells. The effects of CASL-containing diets (250 mg, 6 g, 200 mg, and 1.5 g/kg diet, respectively) were studied in vivo. When KK mice were food deprived for 48 h and subsequently refed a fat-free diet for 72 h, hepatic triglyceride (TG) accumulation was significantly lower in mice fed CASL compared with the control mice. In addition, after obese KK mice were fed a low-fat diet for 2 wk, adipose tissue weights were significantly lower in those fed CASL, but not CA or SL alone, compared with the control mice. Plasma and liver TG levels were also lower in mice fed CASL than in the control mice. These results suggest that CASL is effective for controlling obesity.

Effects of intake of a mixture of thiamin, arginine, caffeine, and citric acid on adiposity in healthy subjects with high percent body fat.

We assessed the effects of intake of thiamin, arginine, caffeine, and citric acid (TACC) on lipid metabolism in healthy subjects. Thirty-one subjects with high percent body fat (> or = 25.0%) were randomly assigned to a 12-wk intervention with daily intake of TACC-supplemented tea (1.1, 1240, 52, and 540 mg, respectively; n=16) or control tea (n=15). The percent body fat decreased significantly during the intervention in both groups, especially in the TACC group. A percentage decrease in triceps skinfold was significantly greater in the TACC group than in the control group. The decrease in abdominal visceral fat in obese subjects was significantly greater in the TACC group than in the control group. Serum triglyceride was significantly lower during intervention than that during the non-intervention period in the TACC group. These results suggest that TACC may be effective in reducing body fat in obese subjects.

Anti-obesity effects of a mixture of thiamin, arginine, caffeine, and citric acid in non-insulin dependent diabetic KK mice.

Anti-obesity effects of a mixture of thiamin, arginine, caffeine, and citric acid (TACC) were investigated in non-insulin dependent diabetic KK mice. Feeding of either arginine or caffeine significantly suppressed an increase in hepatic lipid contents in fasted-refed KK mice. In addition, each component admixed with a low-calorie diet effectively reduced adipose tissue weight in KK mice previously fed a high-calorie diet. The decrease in adipose tissue weight was greater with a mixture of arginine and caffeine, and much greater with TACC than with arginine or caffeine alone. Moreover, plasma insulin concentration was significantly lower in mice fed TACC than in control mice. The anti-obesity effects of TACC were also shown when it was supplemented with a tea beverage. Adipose tissue weight, hepatic triglyceride contents, and plasma insulin concentration were significantly lower in mice given TACC-supplemented tea than in control mice. These results suggest that TACC is effective in reducing adipose tissue mass as well as improving disorders in lipid metabolism.

Nigerooligosaccharides augments natural killer activity of hepatic mononuclear cells in mice.

Nigerooligosaccharides (NOS), a mixture of nigerose and nigerosylmaltooligosaccharides, is immunopotentiating oligosaccharides found in foodstuffs. Augmentation of natural killer (NK) activity by NOS was studied in vitro and in vivo in mice. In vitro treatment of hepatic mononuclear cells (MNC) from normal mice with 1 micro/ml NOS 17 h or just prior to the cytotoxicity assay enhanced their cytotoxicity against YAC-1 cells. NK activity of hepatic MNC was also enhanced in mice injected intraperitoneally with 0.4 mg NOS or in mice given 1% NOS solution orally as a drinking fluid. In association with the augmentation of NK activity, increase in the liver weight induced by intravenous inoculation of EL-4 tumor cells was significantly suppressed by intraperitoneal pretreatment with 0.4 mg NOS. Moreover, drinking 1% NOS significantly improved the survival curve of mice intravenously inoculated with EL-4 cells. Our results suggest that the immunopotentiating activity of NOS is exerted partly through the augmentation of NK activity.