Potent Radical Scavenging Activity Research Articles

Phytochemical Constituents and Biological Activities of Salvia suffruticosa

Background and objectives: Salvia suffruticosa is a perennial plant from Lamiaceae family. Many Salvia species have been employed as medicinal plants; despite the medicinal potentials of S. suffruticosa, there is limited studies regarding its phytochemical profile or biological properties. The aim of the present study was to investigate the chemical constituents of the essential oil and extract of the plant and evaluate its biological activities. Methods: Essential oil from the aerial parts of the plant was extracted by hydrodistillation and analyzed using gas chromatography/mass spectroscopy. Isolation of compounds from methanol and petroleum ether fractions was achieved by using column chromatography with different stationary phases. The structures of the isolated compounds were elucidated by NMR techniques. Cytotoxicity potentials were evaluated using MTT assay and acridine orange/ethidium bromide staining method. Antioxidant activity was assessed by DPPH method. Results: Hydrocarbon sesquiterpenes were identified as the predominant components of the oil, with β-caryophyllene (27.35%), bicyclogermacrene (22.15%), germacrene-D (9.49%) and β-farnesene (9.08%) as the major constituents. Phytochemical analysis of the extract resulted in isolation of lupeol (1), β-sitosterol (2), stigmasterol (3), caffeic acid (4) and 1-feruloyl-β-D-glucopyranose (5). Among the tested samples, lupeol demonstrated the most potent inhibitory activity toward breast cancer cell lines including MCF-7, T-47D and MDA-MB-231 with IC50 values equal to 33.38±2.6, 36.70±3.1 and 23.66±1.4 μg/mL, respectively; caffeic acid with IC50 value of 12.1±1.2 μg/mL showed the most potent radical scavenging activity. Conclusion: The results of this study suggested S. suffruticosa as a promising source of bioactive compounds useful in prevention and treatment of cancer.

Phytochemical Evaluation, Antioxidant Activity and Toxicity of Paeonia daurica ssp. macrophylla Root

Background and objectives: Paeonia daurica ssp. macrophylla, is an herbaceous and perennial plant which belongs to Paeoniaceae family. Two species of this plant grow in northern parts of Iran. The roots in powder form have been used in Persian traditional medicine for treatment of epilepsy, nightmares and gynecological diseases. Several biological activities such as antioxidant and anti-tumor effects of Paeonia species have been reported. Methods: methanol-water (80-20) extract (total extract) was fractionated using n-hexane, chloroform and ethyl acetate. Antioxidant activity of the total extract and fractions were evaluated using DPPH and FRAP assays. Total phenolics content of the extracts was determined by Folin-Ciocalteu method. In addition, cytotoxic activity of the fractions was determined against brine shrimp larvae. Column chromatography with normal phase silica gel and preparative TLC were also used for the isolation and purification of compounds. Results: Evaluation of the results indicated that the ethyl acetate and chloroform fractions with IC50 values of 16.55, 23.9 µg/mL, respectively showed potent radical scavenging activity. As well, the ethyl acetate and chloroform fractions indicated the highest antioxidant power by FRAP assay. Due to the potent antioxidant activity, the chloroform fraction was chosen for further investigations. Three compounds were identified as benzoic acid, veratric acid and oleanolic acid by different spectroscopic methods. Conclusion: According to our findings in this study, the root of Paeonia daurica ssp. macrophyla has beneficial antioxidant activity without toxicity and the therapeutic use of this plant in traditional medicine can be somewhat justifiable.

PP.29.16] NITROSONIFEDIPINE, A NOVEL ANTIOXIDANT, AMELIORATES NEUROLOGICAL SYMPTOMS AND PROLONGS THE SURVIVAL IN A MALIGNANT STROKE-PRONE SPONTANEOUSLY HYPERTENSIVE RATS

Objective: Nitrosonifedipine (NO-NIF) is photolytic metabolite of nifedipine, an anti-hypertensive drug. We have revealed that NO-NIF possesses potent radical scavenging activity and protective effects against several pathological conditions involving oxidative stress. A malignant stroke-prone spontaneously hypertensive rat (M-SHRSP) is a model animal showing high rate of cerebral stroke. The onset and progressions of stroke are known to be involved in oxidative stress. Therefore, we investigated the effects of NO-NIF on the onset and progressions of stroke in M-SHRSP. Design and method: M-SHRSPs were treated with NO-NIF (30 mg/kg/day, i.p.) or vehicle from the age of five weeks. Neurological symptoms were observed every day and scored by severity of general status and disordered motility of anterior/posterior limbs. The day of the stroke onset was determined by neurological symptoms and the change in body weight. Brain was harvested after death and histologically analyzed. For in vitro study, nerve growth factor (NGF)-induced neurite elongation and intracellular signaling pathway were investigated using PC12 cells. Results: There was no significant difference in the incidence and the timing of stroke onset between control and NO-NIF treated group. However, the periods until 50% rats died from stroke were extended for 29 days in NO-NIF group compared to control. Although the neurological score showed linear worsening after stroke onset, NO-NIF significantly suppressed its exacerbation. Consistently, rats showed less pathological lesion, such as hemorrhage, thrombus, and liquefaction degeneration in brains. In PC12 cells, NGF-induced neurite elongation was enhanced by NO-NIF existence. Moreover, NO-NIF prolonged the co-localization of NGF receptor and flotillin, a lipid raft marker, and the activations of Akt and ERK1/2. In our previous study, we have already showed that NO-NIF affects the fluidity of cellular membrane. Therefore, it was considered that NO-NIF enhanced NGF signaling by changing the cellular membrane fluidity and localization of NGF receptor. Conclusions: In the present study, we demonstrated that NO-NIF could prolong the lifespan after stroke induced by malignant hypertension. NO-NIF were suggested to elongate neurite by enhancing NGF-stimulated intracellular signaling activities and finally improved the stroke-related neurological symptoms in M-SHRSP.

Comparison of Volatile Compounds Profile and Antioxydant Activity of Allium sativum Essential Oils Extracted using Hydrodistillation, Ultrasound- Assisted and Sono-Hydrodistillation Processes

Abstract: Objective: This work aims to evaluate the impact of the use of ultrasound during essential oils extraction from garlic bulbs. Material and methods: The essential oils were extracted by Conventional Hydrodistillation method (HD), Ultrasound-Assisted hydro-distillation (US-HD) and Sono-hydrodistillation (SHD) and compared using yield profile, chemical composition and antioxidant activity. Results: Higher performances were obtained when the direct application of ultrasound were used. Indeed, SHD allowed a higher yield after only 30 min of extraction time against 70 min and 90 min for the US-HD and the HD respectively. Among the total identified compounds by GC/MS, Sulphur compounds were found to be the most abundant with qualitative similarities for the three processes. Essential oils showed a potential DPPH radical scavenging activity with IC50 values of 0.96, 1.17 and 1.23 respectively for the three HD, US-HD and SHD processes. Conclusion: The use of ultrasound for the extraction of essential oil from garlic reduced extraction time without causing significant changes in volatile composition and antioxidant activity. Keywords: Garlic essential oil, GC-MS, Antioxidant activity, Hydrodistillation, Ultrasound.

Phytochemical screening and antioxidant activity of Terminalia muelleri benth. leaf extract

In the present investigation, leaves of Terminalia muelleri were assayed for their phytochemical constituents, as well as its free radical scavenging activity. The antioxidant effect was measured using 2,2-diphenyl-1-picrylhydrazyl (DPPH•) radical scavenging assay, the phytochemical constituents were screened to assess their corresponding effect on antioxidant activity. The results showed that the ethanol soluble fraction of T. muelleri possesses a potent radical-scavenging activity using the DPPH• (2,2-diphenyl-1-picrylhydrazyl) radical-scavenging assay (IC50 value = 2.7 μg/mL, while the standard ascorbic acid has an IC50 value = 10.5 μg/mL).

Antioxidant, anticancer and electrochemical redox properties of new bis(2,3-dihydroquinazolin-4(1H)-one) derivatives.

In this paper, a series of bis(2,3-dihydroquinazolin-4(1H)-one) derivatives (4a-i, 10a-k) were synthesized by the one-pot pseudo-five-component reaction of isatoic anhydride with aromatic aldehydes and aromatic amines under reflux in glacial acetic acid. The synthesized compounds were screened for their antioxidant properties using the DPPH radical scavenging method. Compounds 4i and 10h showed potent radical scavenging activities at 20 [Formula: see text] compared to BHA and ascorbic acid. The anticancer activity of compound 4f was evaluated against human breast cancer cell line (MCF 7), and the observed [Formula: see text] was found to be 11.4 [Formula: see text]. The redox behaviour of some analogues was evaluated by cyclic voltammetric methods, and it is found that compound 7d possesses the maximum redox potential.

Proliferation, Accumulation of Polyphenols, and Antioxidant Activities of Callus from the ‘Anji Baicha’ Cultivar of Tea [Camellia sinensis (L.) O. Ktze.

Tea is one of the most consumed beverages worldwide and the relatively high levels polyphenols is benefit for health. In this study, we developed an efficient system for proliferation of callus from ‘Anji Baicha’, a cultivar of tea (Camellia sinensis ). Callus tissue was initially induced by culturing leaf explants on medium containing different plant growth regulators. For callus induction, thidiazuron (TDZ) was more effective than 2,4-dichlorophenoxyacetic acid (2,4-D), α-naphthalene acetic acid (NAA), and N6-benzyladenine (BA). The frequency of callus induction from leaf explants reached 90.21% on 1.0 mg·L -1 TDZ and the developed callus was reddish and friable. We also tested the effect of different concentrations of NAA, 2,4-D, indole 3-acetic acid (IAA), BA, and TDZ, alone and in combinations, on callus proliferation. Medium supplemented with TDZ in combination with IAA was suitable for callus proliferation and accumulation of tea polyphenols. The growth index value and tea polyphenol content of callus cultured on MS medium containing 0.5 mg·L -1 TDZ and 1.0 mg·L -1 IAA was maximally 1,351% and 23.24%, respectively, and the relative abundance of epicatechin was as high as 17.449%. We also measured the antioxidant activity of all samples and the callus with the highest tea polyphenol content also exhibited high potential radical scavenging activity.

Characterization of antioxidants from Detarium microcarpum Guill. et Perr. leaves using HPLC-DAD coupled with pre-column DPPH assay

Detarium microcarpum is an African tree which is widely used as a folk medicine and food. A methanol extract from the leaves as well as its hexane, dichloromethane, ethyl acetate and n-butanol fractions were evaluated for their total phenolic content and radical scavenging activities. Additionally, a pre-column DPPH-HPLC assay was performed to screen and identify the compounds responsible for the antioxidant properties of the extract. Results indicated that the methanol extract contained a substantial amount of phenolic compounds with a value of 51.7 ± 0.2 mg gallic acid equivalent/g. It also exerted potent radical scavenging activities in both DPPH and ORAC assays, with respective values of 937 ± 16 and 2247 ± 63 μmol trolox equivalent/g. Its ethyl acetate and n-butanol fractions were shown to possess the highest TPC and radical scavenging activities. Pre-column DPPH-HPLC experiments led to the direct detection of five potential radical scavenging compounds which were identified as gallic acid, quercetin 3,7-O-dirhamnoside, myricetin 3-O-rhamnoside, quercetin 3-O-glucoside and quercetin 3-O-rhamnoside. All of them were confirmed to exert potent radical scavenging activities using common DPPH spectrophotometric assay. These findings revealed that D. microcarpum leaves possess potent antioxidant activity and could be recommended as a nutraceutical resource. Additionally, the identified bioactive compounds can be considered as appropriate markers for quality control.

Antioxidant and anti-osteoporotic effects of anthraquinones and related constituents from the aqueous dissolved Aloe exudates

In this study, 25 known anthraquinones and related compounds were isolated from aqueous dissolved Aloe exudates. The antioxidant and anti-osteoporotic activities of the isolated compounds were then investigated. Compounds 8, 11, 20, and 23 showed good antioxidant activity based on peroxyl radical-scavenging and reducing capacity assays at a concentration of 10.0 μM. Additionally, compounds 7, 9, 15–16, 18, 21–22 and 24–25 showed potent peroxyl radical-scavenging activities with values ranging from 5.28 to 14.60 at 10.0 μM. Moreover, compounds 8, 11, 15, 20 and 22 exhibited significantly suppressed tartrate-resistant acid phosphatase (TRAP) activity in nuclear factor-κB ligand-activated osteoclastic RAW 264.7 cells, with values of 125.67, 118.54, 127.64, 125.82 and 124.98%, respectively. These results indicate that Aloe is an excellent source of antioxidant and anti-osteoporotic phytochemicals.

IN VITRO ANTI-DIABETIC AND ANTI-OXIDANT ACTIVITIES OF ETHANOL EXTRACT OF TINOSPORA SINENSIS

Objective: The aim of the present study was to evaluate the alpha (α)-amylase and alpha (α)-glucosidase inhibitory activities and in vitro antioxidant activities of the 80 % aqueous ethanol extracts of Tinosporasinensis Lour (Merr.).Methods: The 80% aq. ethanol extract of the plant was prepared. The plant extract was examined for its antioxidant activity by using free radical 1,1-diphenyl-2-picryl hydrazyl (DPPH) scavenging method, ABTS radical scavenging ability, reducing power capacity, estimation of total phenolic content, flavonoid content and flavonol content. Different concentrations (2, 4, 8,10and 15 μg/ml) of the extract was subjected to α-amylase inhibitory and α-glucosidase inhibitory activities and IC50were calculated.Results: The study revealed that the different concentrations of the plant extract possessed a very good amount of total phenolics, flavonoid and flavonol and exhibited potent radical scavenging activity using DPPH and ABTS as a substrate. The ethanol extracts exhibited significant α-amylase and α-glucosidase inhibitory activities with an IC50 value1.093µg and 1.04µg dry extract respectively and well compared with standard acarbose drug.Conclusion: Thus, it could be concluded that due to the presence of antioxidant components the plant extracts could be used for the treatment of hyperglycemia, diabetes and the related condition of oxidative stress. This knowledge will be useful in finding more potent components from the natural resources for the clinical development of antidiabetic therapeutics.

In vitro Evaluation of Antioxidant and Antiproliferative Activities of Artemisia nilagirica Extracts

Medicinal plants serve as unlimited source for phytoconstituents that possess potent antioxidant and antiproliferative properties. Artemisia nilagirica L. is a potent medicinal plant widely found in India and has been used to treat human diseases for centuries. The present investigation was conducted to investigate the antioxidant and antiproliferative abilities of different crude extracts of A. nilagirica. A detailed study was performed on the antioxidant activity and antiproliferative activity of the methanol extract of A. nilagirica by in vitro chemical analysis. The chemical composition of extracts, studied in terms of phenolics, total flavonoids, triterpenoids, tannins and alkaloids were also determined. Results suggested that the amounts of different phytoconstituents in extracts vary based on polarity of solvents. The extracts possessed different antioxidant and radical-scavenging activities in different assays. Among the extracts, ethyl acetate and alcohol extracts showed the most potent radical-scavenging activities. Ethyl acetate and alcohol extracts showed cytotoxicity towards eight different human cancer cell lines in vitro with IC50 values ranging between 30.43±0.86 and 982.46±14.40 μg/ml and they were less toxic to normal cell lines. The findings of this study provide evidence that A. nilagirica extracts can be used potentially as ready accessible and valuable bioactive source for isolation of antioxidant and anticancer agents.

Antioxidative and Anti-Inflammatory Activities of Galloyl Derivatives and Antidiabetic Activities of Acer ginnala.

Chromatographic isolation of the 80% MeOH extract of Acer ginnala (AG) yielded seven galloyl derivatives: gallic acid (1), ginnalin B (2), acertannin (3), maplexin D (4), maplexin E (5), quercetin-3-O-(2′′-galloyl)-α-L-rhamnopyranoside (6), and kaempferol-3-O-(2′′-galloyl)-α-L-rhamnopyranoside (7). This is the first study to report the isolation of compounds 4 and 5 from AG. Galloyl derivatives 3–7 exhibited potent radical scavenging activities, with 5 and 7 showing particularly strong inhibitory activities against nitric oxide production in lipopolysaccharides- (LPS-) stimulated RAW264.7 cells. In addition, oral administration of AG extract (500 mg/kg b.w.) improved symptoms of hyperglycemia and blunted the increases in serum GOT/GPT levels in a rat model of streptozotocin-induced diabetes. These results suggest that galloyl derivatives (1–7) are antioxidant and anti-inflammatory agents and that AG extract has potential as a functional material or novel herbal medicine for treating diabetes mellitus.

A Primary Study of the Antioxidant, Hypoglycemic, Hypolipidemic, and Antitumor Activities of Ethanol Extract of Brown Slimecap Mushroom, Chroogomphus rutilus (Agaricomycetes).

In vivo and in vitro treatments were carried out to investigate the effects of a 95% ethanol extract of Chroogomphus rutilus (CRE) on antioxidant, hypoglycemic, hypolipidemic, and antitumor properties. CRE showed potent radical scavenging activity against DPPH in vitro. It could increase antioxidant enzymatic activities (superoxide dismutase and glutathione peroxidase) and could reduce malondialdehyde content in vivo in mice in which aging was induced by D-galactose. CRE had hypoglycemic activity and could significantly inhibit α-glucosidase activity in vitro and decrease blood glucose concentration in vivo. CRE could decrease the serum total cholesterol, triglyceride, and low-density lipoprotein cholesterol levels and increase the high-density lipoprotein cholesterol level in diabetic mice. The MTT assay showed that CRE also had a certain inhibitory effect on the tumor cells. These results suggest that CRE may be beneficial for human health and could be useful for applications in medicine, the food industry, and agriculture.

가는갯능쟁이 (Atriplex gmelinii)의 항산화 효과

In the present study, antioxidant activity of crude extract and its solvent-partitioned subfractions (n-hexane, 85% aqueous methanol, n-butanol, and water) obtained from Atriplex gmelinii was investigated using several different antioxidant assays. The tested samples possessed different antioxidant and radical-scavenging activities in different assays. n-butanol fraction showed the most potent radical-scavenging activity on reducing power while 85% aqueous methanol fraction exhibited the highest radical-scavenging activity on DPPH radicals and intracellular reactive oxygen species (ROS). On the otherhand, n-BuOH and 85% aqueous methanol revealed the similar inhibitory effect on peroxynitrite-scavenging and genomic DNA oxidation. These results suggest that the Atriplex gmelinii can be used as the valuable source for developing a natural antioxidant.

In vitro antioxidant potential and free radical scavenging activity of various extracts of pollen of Nelumbo nucifera Gaertn.

In vitro antioxidant potential and free radical scavenging activity of various extracts of pollen of Nelumbo nucifera Gaertn.

Prophylactic and curative anti-ulcerogenic activity and the possible mechanisms of action of some desert plants

The present study aimed to evaluate the anti-ulcerogenic activities and the possible mechanisms of action of seven desert plants from different families. Conyza dioscoridis (L.) Desf. (Asteraceae), Euphorbia hirta L. (Euphorpiaceae), Origanum syriacum L., Salvia lanigera L. (Lamiaceae), Sisymbrium irio L., Solanum nigrum Linn. (Solanaceae) and Solenostemma arghel (Del.) Hayne. (Asclepiadaceae), were tested using prophylactic and curative models of absolute ethanol-induced ulcer, at three doses (125, 250 & 500mg/kg) of each extract.The investigated extracts possessed dose dependent anti-ulcerogenic activities in both models, with LD50 higher than 5g/kg. The most effective extracts were C. dioscoridis and S. irio with percent protection of control ulcer; 91.1% and 85.4% respectively. The antisecretory activity of both C. dioscoridis and S. irio appears to be mainly related to the suppression of gastrin release. The in vitro potential radical (DPPH) scavenging activities of the investigated extracts were well supported with the reduction in gastric MDA (50.6% and 43.3%) and enhancing the level of reduced GSH (2.84, 2.59mg/g tissue) for C. dioscoridis and S. irio respectively. In addition, suppression of the inflammatory mediator TNF-α may be one of the possible mechanisms of action. The alcohol extracts of C. dioscoridis and S. irio showed no alteration on liver and kidney functions. Phytochemical screening of the investigated extracts revealed the presence of flavonoids, tannins and sterols which could be related to the activities.

Design, Synthesis, and Biological Evaluation of Novel Tetramethylpyrazine Derivatives as Potential Neuroprotective Agents.

Oxidative stress plays a crucial role in neurological diseases, resulting in excessive production of reactive oxygen species, mitochondrial dysfunction and cell death. In this work, we designed and synthesized a series of tetramethylpyrazine (TMP) derivatives and investigated their abilities for scavenging free radicals and preventing against oxidative stress-induced neuronal damage in vitro. Among them, compound 22a, consisted of TMP, caffeic acid and a nitrone group, showed potent radical-scavenging activity. Compound 22a had broad neuroprotective effects, including rescuing iodoacetic acid-induced neuronal loss, preventing from tert-butylhydroperoxide (t-BHP)-induced neuronal injury. Compound 22a exerted its neuroprotective effect against t-BHP injury via activation of the phosphatidyl inositol 3-kinase (PI3K)/Akt signaling pathway. Furthermore, in a rat model of permanent middle cerebral artery occlusion, compound 22a significantly improved neurological deficits, and alleviated the infarct area and brain edema. In conclusion, our results suggest that compound 22a could be a potential neuroprotective agent for the treatment of neurological disease, particularly ischemic stroke.

LC/MS characterization of antioxidant flavonoids from Tragia involucrata L.

Chromatographic and spectroscopic methods were developed for the identification of flavonoids with potent radical scavenging activity from Tragia involucrata, an important medicinal plant. The separation consisted of column chromatography and high pressure preparative liquid chromatography. In vitro radical scavenging activity of different fractions was screened by DPPH radical scavenging assay. The ESI MS/MS analysis was employed for the characterization of active fractions. Flavonoids such as iridin, dihexosyl quercetin, quercetin-3-O-rutinoside, rhamnosyl hexosyl methyl quercetin, gentenstein 7-glucoside, orientin, C-(O-caffeoyl-hexosyl)-O-hexoside and tricin 7-O-hexosyl-O-hexoside were identified, for the first time, from T. involucrata.

Comparative Analysis of Fatty Acids and Antioxidant Activity of Betula utilis Bark Collected from Different Geographical Region of India

Objective: The present study investigated the comparative analysis of fatty acid and antioxidant activity from the bark of Betula utilis, collected from Kashmir and Sikkim coded as BUK and BUS respectively. Methods: Fatty acid constituents were analyzed by GCMS (gas chromatography mass spectroscopy) in BUK and BUS bark. B. utilis bark extracts of both the locations were phytochemically investigated and radical scavenging activity was evaluated by DPPH in all solvent fractions. Results: Phenolic content were found to be higher in methanolic extracts of BUK bark, 5.8 ± 0.1 mg/gm and flavonoid content were higher in the water extract of BUK bark, 6.16 ± 0.2 mg/gm. The radical scavenging activity was found to be higher methanolic, alcoholic and water extracts of BUK bark. The lowest IC50 value for radical scavenging activity of methanolic and water extracts of B. utilis bark collected from Kashmir were found 18.7 ± 1.1 and 18.2 ± 0.3 μg/mL, respectively. Thirteen fatty acids were identified in the sample BUK as well as BUS in which the major fatty acids were found-Palmitic acid, linoleic acid and oleic acid. BUK possess the highest amount of Palmitic acid (18.07%) in oily portion of the bark. Conclusion: The present study concluded that BUK and BUS extracts have shown significant antioxidant activity in comparison to standard but BUK possess potent radical scavenging activity over BUS, and higher amounts palmitic acid, linoleic acid and oleic acid was present in the extract, and may a play an important role in nutritional and pharmaceutical applications.

Abstract: Topical Application of Nitrosonifedipine, a Novel Free Radical Scavenger, Ameliorate the Ischemic Skin Flap Necrosis

INTRODUCTION: Ischemic flap necrosis is often occurred by insufficient blood supply, which prolongs the treatment period and occasionally requires an additional surgery. Ischemic flaps generate an excess amount of free radicals which is regarded as a major factor of ischemic skin necrosis. Thus free-radical scavengers would be an effective drugs in improvement of flap survival. In our previous studies, we have reported nitrosonifedipine (NO-NIF), which is a photolytic compound of nifedipine, possesses a potent radical scavenging activity, and shows the favorable effects against vascular endothelial dysfunction and type 2 diabetic nephropathy. In this study, we evaluated the ameliorating effect of NO-NIF on the ischemic flap model mice. MATERIAL AND METHODS: 9–10 weeks old Male C57BL/6 mice were divided into 2 groups, NO-NIF or control (n=6 in each group), respectively. A 1.0 × 3.0 cm cranially based random pattern flap was elevated on the dorsum of mice. NO-NIF 30 mg/kg or vehicle was injected subcutaneously immediate after the operation and once a day until evaluation. Seven days after surgery, the survival area was calculated as a percentage of the total flap area. To detect the oxidative stress, malondialdehyde (MDA) in the distal part of the flap at post-operative day 1 and 3 was measured by thiobarbituric acid reactive substances assay. Protein expression of p22phox, an essential component of NADPH-oxidase, in the flap was measured by western blotting. RESULTS: At post-operative day 7, the flap survival area was significantly larger in the NO-NIF-treated mice than controls (78.29 ± 7.04% vs. 51.81 ± 6.85%, p=0.021). The amount of MDA significantly decreased in the NO-NIF-treated mice at post-operative day 3 (2.31 ± 0.28µmol/g protein vs. 4.21 ± 0.32µmol/g protein, p=0.001), whereas MDA was same level in the both groups at the post-operative day 1 (2.77 ± 0.61µmol/g protein vs. 2.96 ± 0.51µmol/g protein). In a manner consistent with MDA levels, protein expression level of p22phox was decreased in the NO-NIF-treated mice at post-operative day 3 (p=0.002). CONCLUSIONS: We present the ameliorating effect of NO-NIF on ischemic flap survival. MDA and p22phox protein was decreased by NO-NIF treatment, which suggests the ameliorating effect was exerted via free radical scavenging. This investigation indicates that free-radical scavengers including NO-NIF are effective drugs in improvement of flap survival. DISCLOSURE/FINANCIAL SUPPORT:None.