Anti-elastase Activity Research Articles

Screening of the Antioxidant and Anti-elastase Activities for the Extracts of Jeju Endemic Plants

Screening of the Antioxidant and Anti-elastase Activities for the Extracts of Jeju Endemic Plants

Synthesis, antibacterial, antielastase, antiurease and antioxidant activities of new methoxy substitued bis-1,2,4-triazole derivatives

The methoxy substitued two novel bis triazole-schiff bases (6 a–b) were synthesized with 4-amino-3,5-diethyl-4H-1,2,4-triazole and various bis-aldehydes. Their amine derivatives prepared by reduced with NaBH4 (5 a–b). The obtained products 6 a–b and 7 a–b were identified by FT-IR, 1H-NMR, 13C-NMR. The bis triazole-schiff bases and amine derivatives were tested for antimicrobial activity using the agar diffusion technique against 11 bacteria. The synthesized compounds (6 a–b and 7 a–b) were screened for their antielastase, antiurease and antioxidant activities. The resuts showed that the synthesized compounds (6 a–b and 7 a–b) had effective antielastase and antiurease activities.

Antioxidant and potential anti-inflammatory activity of extracts and formulations of white tea, rose, and witch hazel on primary human dermal fibroblast cells

BackgroundNumerous reports have identified therapeutic roles for plants and their extracts and constituents. The aim of this study was to assess the efficacies of three plant extracts for their potential antioxidant and anti-inflammatory activity in primary human skin fibroblasts.MethodsAqueous extracts and formulations of white tea, witch hazel and rose were subjected to assays to measure anti-collagenase, anti-elastase, trolox equivalent and catalase activities. Skin fibroblast cells were employed to determine the effect of each extract/formulation on IL-8 release induced by the addition of hydrogen peroxide. Microscopic examination along with Neutral Red viability testing was employed to ascertain the effects of hydrogen peroxide directly on cell viability.ResultsConsiderable anti-collagenase, anti-elastase, and antioxidant activities were measured for all extracts apart from the witch hazel distillate which showed no activity in the collagenase assay or in the trolox equivalence assay. All of the extracts and products tested elicited a significant decrease in the amount of IL-8 produced by fibroblast cells compared to the control (p < 0.05). None of the test samples exhibited catalase activity or had a significant effect on the spontaneous secretion of IL-8 in the control cells which was further corroborated with the microscopy results and the Neutral Red viability test.ConclusionsThese data show that the extracts and products tested have a protective effect on fibroblast cells against hydrogen peroxide induced damage. This approach provides a potential method to evaluate the claims made for plant extracts and the products in which these extracts are found.

Elastase Inhibitory Activity of Airway α1-Antitrypsin Is Protected by Treatment With a Catalytic Antioxidant in a Baboon Model of Severe Bronchopulmonary Dysplasia

Recent studies in animal models of bronchopulmonary dysplasia (BPD) suggest that antioxidant treatments may be beneficial for the disease. However, the mechanisms by which these drugs improve the course of BPD are not completely known. Alpha1-antitrypsin (α1-AT) is one of the major serine protease inhibitors in human plasma that has antielastase and antiapoptotic activities. Both activities of α1-AT are dependent on its reactive site loop (RSL), which is highly susceptible to oxidative inactivation. In this study, we investigated the elastase inhibitory activity of α1-AT in two different baboon models of BPD, the "new BPD" and the "severe BPD" models, and determined the effect of treatment with a catalytic antioxidant, Mn(III) meso-tetrakis(N-ethylpyridinium-2-yl)porphyrin (MnTE-2-PyP), on the elastase inhibitory activity of α1-AT in the severe BPD model. Our results demonstrate the presence of sufficient elastase inhibitory activity of the airway α1-AT in the new but not in the severe BPD model. Treatment of severe BPD group baboons with the catalytic antioxidant MnTE-2-PyP resulted in augmentation of the elastase inhibitory activity of α1-AT. These findings suggest that prevention of the oxidative inactivation of α1-AT may be one of the mechanisms by which antioxidant therapy improves the pulmonary outcomes in animal models of severe BPD.

In vitro antifungal, anti-elastase and anti-keratinase activity of essential oils of Cinnamomum-, Syzygium- and Cymbopogon-species against Aspergillus fumigatus and Trichophyton rubrum

This study was aimed to evaluate effects of certain essential oils namely Cinnamomum verum, Syzygium aromaticum, Cymbopogon citratus, Cymbopogon martini and their major components cinnamaldehyde, eugenol, citral and geraniol respectively, on growth, hyphal ultrastructure and virulence factors of Aspergillus fumigatus and Trichophyton rubrum. The antifungal activity of essential oils and their major constituents was in the order of cinnamaldehyde > eugenol > geraniol = C. verum > citral > S. aromaticum > C. citratus > C. martini, both in liquid and solid media against T. rubrum and A. fumigatus. Based on promising antifungal activity of eugenol and cinnamaldehyde, these oils were further tested for their inhibitory activity against ungerminated and germinated conidia in test fungi. Cinnamaldehyde was found to be more active than eugenol. To assess the possible mode of action of cinnamaldehyde, electron microscopic studies were conducted. The observations revealed multiple sites of action of cinnamaldehyde mainly on cell membranes and endomembranous structures of the fungal cell. Further, test oils were also tested for their anti-virulence activity. More than 70% reduction in elastase activity was recorded in A. fumigatus by the oils of C. verum, C. martini, eugenol, cinnamaldehyde and geraniol. Similar reduction in keratinase activity in A. niger was recorded for the oils of C. martini and geraniol. Maximum reduction (96.56%) in elastase activity was produced by cinnamaldehyde whereas; geraniol caused maximum inhibition (97.31%) of keratinase activity. Our findings highlight anti-elastase and anti-keratinase activity of above mentioned essential oils as a novel property to be exploited in controlling invasive and superficial mycoses.

Anti-hyaluronidase and anti-elastase activity screening of tannin-rich plant materials used in traditional Polish medicine for external treatment of diseases with inflammatory background

The aim of the study was to examine extracts from twelve tannin-rich plant materials used in traditional Polish medicine for external treatment of skin and mucosa diseases considering their ability to inhibit hyaluronidase activity and elastase release from stimulated neutrophils in vitro. In vitro anti-hyaluronidase and anti-elastase assays together with phytochemical qualitative and quantitative screening were performed. The strongest inhibition of hyaluronidase was observed for extract from Lythri herba, with IC(50) value 8.1 ± 0.8 μg/mL. The most active extract towards elastase release was from Hippocastani cortex which at concentration 10 μg/mL showed 62.0 ± 6.9% inhibition. Anti-hyaluronidase and anti-elastase activity of chosen tannin-rich plant materials can support their traditional use in folk medicine. Strong inhibition of both enzymes by extract from Lythri herba makes this pharmacopeial plant material an interesting topic for further biological and phytochemical examination.

The influence of hydrosalpinx on endometrial elafin expression

The endometrium of women with hydrosalpinx has an increased number of neutrophils and lower expression of elafin, an elastase inhibitor and natural antimicrobial molecule. These findings suggest that women with hydrosalpinx have a reduced antimicrobial and antielastase activity.

A Bowman–Birk inhibitor with anti-elastase activity from Lathyrus sativus L. seeds

Four Bowman-Birk inhibitors, named LSI-1/4, were isolated and purified from Lathyrus sativus L. seeds. The purification procedure consisted of two cation-exchange chromatography steps, followed by gel-filtration and RP-HPLC. Mass spectrometry analysis of LSI-1/4 inhibitors yielded relative molecular masses of 7914.41 for LSI-1, 6867.67 for LSI-2, 7341.24 for LSI-3 and 7460.01 for LSI-4. N-terminal sequences (up to 30 residues) of LSI-1/4 inhibitors were identical with the exception of sequence positions 21, 27 and 28 and highly similar to those of other Bowman-Birk inhibitors isolated from Leguminosae plants. Inhibitors LSI-1/4 were active towards trypsin and α-chymotrypsin, with IC(50) values for 12.6 nM of trypsin ranging from 4.9 to 24.3 nM. A lower activity was observed against bovine α-chymotrypsin (IC(50) values ranging from 0.5 to 3.4 μM for 15.0 nM of α-chymotrypsin). Peptide mapping of the LSI-1 sequence showed the presence of an Ala residue in the second reactive site, thus explaining the low anti-chymotrypsin activity of this inhibitor. In addition, LSI-1 was endowed with anti-elastase activity, being able to inhibit human leukocyte elastase.

Cucumis sativus fruit-potential antioxidant, anti-hyaluronidase, and anti-elastase agent

The aim of this study was to screen the antioxidant, anti-hyaluronidase, and anti-elastase activity of the lyophilized juice of Cucumis sativus fruit (CSLJ). The CSLJ was subjected to DPPH and superoxide radical scavenging assay in reference to butylated hydroxytoluene. The hyaluronidase and elastase inhibitory assay was performed in reference to oleanolic acid. Furthermore, the activities have been rationalized with HPLC analysis of the CSLJ with standard reference compound of ascorbic acid. The CSLJ exhibited DPPH-free radical and superoxide radical scavenging activity, IC(50) at a concentration of 14.73 ± 1.42 and 35.29 ± 1.30 μg/mL, respectively. The CSLJ also showed strong anti-hyaluronidase ((c) P < 0.001) and anti-elastase ((c) P < 0.001) activity, IC(50) at a concentration of 20.98 ± 1.78 and 6.14 ± 1.74 μg/mL, respectively. The HPLC 'chromatogram' of standard and CSLJ showed specific peak at retention time 2.905 and 3.066 min, respectively. Content of ascorbic acid was calculated with respect to the standard compound and it was found to be 3.5 ± 0.23% w/w. CSLJ is the rich source of ascorbic acid and this study thereby rationalizes the use of C. sativus as potential anti-wrinkle agent in cosmetic products.

Effects of sulphurous water on human neutrophil elastase release

Molecules bearing a sulphide (HS) group, such as glutathione, play a fundamental role in the defensive system of human airways, as shown by the fact that the lining fluid covering the epithelia of the respiratory tract contains very high concentrations of glutathione: the lungs and nose, respectively, contain about 140 and 40 times the concentrations found in plasma. Consequently, various low-weight soluble molecules bearing an HS group (including N-acetylcysteine, mesna and thiopronine, and prodrugs such as stepronine and erdosteine) have been used for therapeutic purposes. HS groups can also be therapeutically administered by means of sulphurous thermal water containing HS groups. The aim of this study was to investigate the direct activity of such water on the release of elastase by activated human neutrophils. After the neutrophils were incubated with increasing amounts of sulphurous water or the HS/hydrogen sulphide donor sodium hydrosulphide (NaHS), elastase release was initiated by N-formyl-methionyl-leucyl-phenylalanine and measured by means of spectrofluorimetry using methylsuccinylalanylprolylvalyl-methylcoumarin amide as the fluorogenic substrate. To verify the presence of direct action on elastase we determined the diameter of the area of elastinolysis on elastine-agarose gel plates. The sulphurous water significantly inhibited elastase release at HS concentrations ranging from 4.5 to 18 μg/ml, as assayed using the iodometric method; in the case of NaHS, the inhibition was significant at HS concentrations ranging from 2.2 to 18 μg/ml. The concentration-effect regression lines of both were parallel and neither showed any direct elastolytic activity. Previous claims concerning the activity of sulphurous water have been based on the patients' subjective sense of wellbeing and on symptomatic (or general) clinical improvements that are not easy to define or quantify exactly. Our findings indicate that, in addition to its known mucolytic and antioxidant activity, sulphurous water also has an anti-elastase activity that may help to control the inflammatory processes of upper and lower airway diseases.

Anti-hyaluronidase, Anti-elastase Activity of Garcinia indica

Anti-hyaluronidase, Anti-elastase Activity of Garcinia indica

Expression and purification of recombinant human α 1-proteinase inhibitor and its single amino acid substituted variants in Escherichia coli for enhanced stability and biological activity

Human α 1-proteinase inhibitor (α 1-PI) is the most abundant protease inhibitor found in the blood and expression of biologically active recombinant α 1-PI has great potential in therapeutic applications. We report here the expression of a synthetic α 1-PI gene and its variants in Escherichia coli. Modified α 1-PI gene and its single amino acid variants were cloned in pMAL-c2X vector, which allowed expression of recombinant protein(s) as a fusion of maltose-binding protein (MBP) with factor Xa protease recognition site between the fusion partners. The synthetic gene(s) were expressed in different E. coli strains and maximum expression of recombinant α 1-PI and variants up to 24% of total soluble protein (TSP) was achieved with engineered strain carrying extra copies of tRNAs for rare codons. Recombinant α 1-PI protein(s) were purified by amylose affinity chromatography with high homogeneity and overall yield of about 7–9 mg l −1 of bacterial culture (∼5.2 g wet cell mass). E. coli expressed recombinant α 1-PI showed specific anti-elastase activity and appeared as a single band of ∼45.0 kDa on SDS-PAGE. Primary structure of purified protein and integrity of N-terminus has been verified by mass spectrometric analysis. Recombinant α 1-PI expressed in E. coli was fully intact having molecular mass similar to native unglycosylated protein purified from human plasma. Increased thermostability and specific activities of purified α 1-PI variant proteins confirmed the stabilizing effect of incorporated mutations. Our results demonstrate efficient expression and purification of stable and biologically active α 1-PI and its variants in E. coli for further therapeutic applications.

Probucol inhibits tobacco smoke-induced decrease in plasma anti-elastase activity and ferroxidase activity in rats

Elastolytic enzymes and active oxygen species derived from leukocytes and alveolar macrophages during exposure to tobacco smoke, together with active oxygen species directly derived from tobacco smoke, are thought to play a crucial role in the pathogenesis of pulmonary emphysema by inactivating alpha 1 protease inhibitor (alpha 1 PI), a novel anti-elastase. We studied the inhibitory effect of probucol, an oral hypocholesterolemic agent, on tobacco smoke-induced decrease in plasma anti-elastase activity (EIA) and ferroxidase activity (FA) in conscious venous catheter instrumented rats. Rats exposed to the smoke of 5 cigarettes (nicotine 11 mg, tar 115 mg) in a plastic chamber showed a prompt increase in plasma COHb to 17.9 +/- 2.7%, and a prompt decrease in plasma EIA by -17.9% (p less than 0.05) and FA by -14.8% (p less than 0.01), which lasted for 6 hours after exposure. Rats administered probucol (1% probucol in food) for 3 days showed normal cholesterol plasma levels, and rats administered probucol for 4 weeks showed hypocholesterolemic plasma levels. EIA and FA were not depressed after smoking, and lipid peroxide product (TBA reactive substance) in lung tissue (p less than 0.05) and serum (p less than 0.1) showed a smaller increase in association with a smaller decrease in the ratio of lung tissue GSH/GSSG (p less than 0.01) compared with control rats. These results indicate that probucol, via its antioxidant action rather than its cholesterol lowering effect, has a protective effect on lung exposed to tobacco smoke in terms of protease-antiprotease balance and oxidant-antioxidant balance.

Optimized and Standardized Isolation and Structural Characterization of Anti-inflammatory Sulfated Polysaccharides from the Red Alga Delesseria sanguinea (Hudson) Lamouroux (Ceramiales, Delesseriaceae)

The red seaweed Delesseria sanguinea dominantly populates a large artificial reef in the southwestern Baltic Sea. It contains sulfated polysaccharides (SPs), which exhibit a pharmacological profile indicating anti-inflammatory and anti-skin aging potencies. To establish and optimize an extraction procedure for these SPs and to evaluate the influence of several parameters on their quality, 23 algae batches were harvested over the period of four years and extracted by different methods, resulting in 56 SP batches. Extraction with water at 85 degrees C proved to be superior and led to highly reproducible products with average yields of 11.6 +/- 3.9%, reaching 18% in spring. Their quality was independent of generation form and vitality of the algae. The SPs were identified as sulfated branched xylogalactans (degree of sulfation 0.50 +/- 0.08, mean M(r) 142000). The coextraction of floridean starch turned out to be the only parameter causing any seasonal variability. However, by using water, this concerns solely the yields of the isolated products. Compared to NaOH extracts, the antielastase activity of H(2)O extracted SPs was about twice as strong (IC(50) 0.204 +/- 0.024 microg/mL) and the batch to batch variability was much lower (CV 11.8 vs 28.6%). In conclusion, SPs from Delesseria sanguinea can be isolated in reproducibly high quality by using a specific extraction procedure. Thus, an important prerequisite for a potential economical utilization is fulfilled.

Effects of Alchornea cordifolia on elastase and superoxide anion produced by human neutrophils

The ability of Alchornea cordifolia (Schum. and Thonn.) Müll. Arg. (Euphorbiaceae) leaves to inhibit human neutrophil elastase (HNE) and superoxide anion (O2•–) activities was evaluated on aqueous and ethyl acetate extracts as they allow for a targeted extraction of polyphenols. The direct effect of A. cordifolia extracts on HNE and O2•– was assessed in an acellular system. Results showed that extracts scavenge HNE and O2•– in a dose-dependent manner. Better activity was exhibited by the ethyl acetate extract with lower IC50 (2.2 and 4. 1 mg/L for HNE and O2•–, respectively) than for the aqueous extract. Cellular systems including isolated human polymorphonuclear neutrophils (PMN) were investigated to assess the effect of extracts on PMN metabolism. PMN were stimulated with 4β-phorbol-12-myristate-13-acetate (PMA), calcium ionophore (CaI), or N-formyl-methionyl-leucine-phenylalanine (fMLP), each stimulant having its own stimulation pathway. From the IC50 obtained, it can be concluded that A. cordifolia reduces HNE and O2•– liberation. Furthermore it was demonstrated that A. cordifolia extracts have no cytotoxic activity on PMN by measuring release of the cytosolic enzyme lactate dehydrogenase. As the ethyl acetate extract offers a higher rate of total phenols than the aqueous extract as well as better scavenging activity, it can be supposed that polyphenols, which are well known for their potent antioxidant and antielastase activity, are implicated in the activity of the plant. Phenolic substances such as quercetin, myricetin-3-glucopyranoside, myricetin-3-rhamnopyranoside, and proanthocyanidin A2 were identified in the ethyl acetate extract. In conclusion, the study provides proof of ethnomedical claims and partly explains the mechanisms of the anti-inflammatory action of A. cordifolia leaves.

Anti-collagenase, anti-elastase and anti-oxidant activities of extracts from 21 plants.

BackgroundOwing to their roles in tissue remodelling in health and disease, several studies have reported investigations on plant extracts as inhibitors of proteinases and as anti-oxidants.MethodsThe anti-ageing and anti-oxidant properties of 23 plant extracts (from 21 plant species) were assessed as anti-elastase and anti-collagenase activities and in selected anti-oxidant assays along with phenolic content.ResultsAnti-elastase activities were observed for nine of the extracts with inhibitory activity in the following order: white tea (~89%), cleavers (~58%), burdock root (~51%), bladderwrack (~50%), anise and angelica (~32%). Anti-collagenase activities were exhibited by sixteen plants of which the highest activity was seen in white tea (~87%), green tea (~47%), rose tincture (~41%), and lavender (~31%). Nine plant extracts had activities against both elastase (E) and collagenase (C) and were ranked in the order of white tea (E:89%, C:87%) > bladderwrack (E:50%, C:25%) > cleavers (E:58%, C:7%) > rose tincture (E:22%, C:41%) > green tea (E:10%: C:47%) > rose aqueous (E: 24%, C:26%) > angelica (E:32%, C:17%) > anise (E:32%, C:6%) > pomegranate (E:15%, C:11%).Total phenolic content varied between 0.05 and 0.26 mg gallic acid equivalents (GAE)/mL with the exception of white tea (0.77 mg GAE/mL). For anti-oxidant assessment, the Trolox equivalent anti-oxidant capacity (TEAC) assay revealed activity for all extracts. White tea had the highest activity equivalent to ~21 μM Trolox for a 6.25 μg aliquot. In addition, seven extracts exhibited activities = 10 μM Trolox with witch hazel (6.25 μg = 13 μM Trolox) and rose aqueous (6.25 μg = 10 μM Trolox) showing very high activities at low concentrations. A high activity for white tea was also found in the superoxide dismutase (SOD) assay in which it exhibited ~88% inhibition of reduction of nitroblue tetrazolium. High activities were also observed for green tea (86.41%), rose tincture (82.77%), witch hazel (82.05%) and rose aqueous (73.86%).ConclusionFrom a panel of twenty three plant extracts, some one dozen exhibit high or satisfactory anti-collagenase or anti-elastase activities, with nine having inhibitory activity against both enzymes. These included white tea which was found to have very high phenolic content, along with high TEAC and SOD activities.

HDL antielastase activity prevents smooth muscle cell anoikis, a potential new antiatherogenic property

Various studies using proteomic approaches have shown that HDL can carry many proteins other than its constitutive apolipoprotein A-I (apoA-I). Using mass spectrometry and Western blotting, we showed the presence of alpha(1)-antitrypsin (AAT) (SERPINA1, serpin peptidase inhibitor, clade A, an elastase inhibitor) in HDL, isolated either by ultracentrifugation or by selected-affinity immunosorption using an anti-apoA-I column. Furthermore, we report that HDL possesses potent antielastase activity. We further showed that only HDL but not LDL is able to bind AAT. HDL-associated AAT was able to inhibit extracellular matrix degradation, cell detachment, and apoptosis induced by elastase in human vascular smooth muscle cells (VSMCs) and in mammary artery cultured ex vivo. Degradation of fibronectin by elastase used as a marker of pericellular proteolysis was prevented by addition of HDL. Elastase present in aortic abdominal aneurysm (AAA) thrombus samples was also able to induce apoptosis of VSMCs in culture. This phenomenon was prevented by addition of HDL but not of LDL. Finally, we report that the proportion of AAT in HDL isolated from patients with an AAA is decreased relative to that from matched control subjects, suggesting a reduced capacity of HDL to inhibit elastase in these patients. In conclusion, our data provide evidence of a new potential antiatherogenic property of HDL attributable to AAT and its antielastase activity.

Potent Antielastase and Antityrosinase Activities of Astilbe chinensis

Problem statement: In the current scenario, photo-aging is a major problem causing skin wrinkling and hyperpigmentation. Therefore, elastase and tyrosinase inhibitors play an important role in the treatment of skin aging and thus, gaining a special attention in cosmetic industries. Approach: In the screening of Korean medicinal plants to search the potent elastase and tyrosinase inhibitors, the extract of the rhizomes of Astilbe chinensis (A. chinensis) exhibited the strongest potential. Further the crude 100% methanolic extract of A. chinensis along with its n-hexane, methylene chloride, ethyl acetate and aqueous fractions were investigated for elastase and tyrosinase enzyme inhibition activities. Results: The A. chinensis extracts showed remarkable elastase inhibitory activities (at a concentration of 20 μg 155 μL-1 reaction) ranging from 38-92% and tyrosinase inhibition activities (at a concentration of 20 μg 150 μL-1 reaction) ranging from 2-90%. Among all the fractions, water fraction demonstrated the highest elastase and tyrosinase inhibitory activities (92 and 90%, respectively). Conclusion: Based on the noteworthy antielastase and antityrosinase activities by the rhizome of A. chinensis, it might prove a strong candidate as an active ingredient in cosmaceutical formulations and further in vitro and in vivo investigations is needed.

In Vivo Introduction of an Extrachromosomal Plasmid Expressing Long-term Human Alpha-1-antitrypsin protects Islet Allografts (141.48)

Abstract Alpha-1-antitrypsin (AAT) exerts anti-inflammatory and tolerogenic activities during islet allograft transplantation in diabetes mouse models. Although a serine protease inhibitor, evidence suggests that AAT possesses activities that are independent of protease inhibition. The previously reported plasmid, pEF-hAAT, contains the genomic sequence of human AAT (hAAT) and sustains circulating levels after single hydrodynamic tail-vein injection (HDI) in mice. To asses whether expression of hAAT by pEF-hAAT protects islets from acute rejection, pEF-hAAT (100 μg, n = 6) or PBS (n = 4, control) were hydrodynamically introduced to mice. Liver expression and circulating hAAT levels were determined. Supernatant of transfected Hepa1c cells exhibited anti-elastase activity. Sixteen days post HDI mice were injected streptozotocin (225 mg/kg) and grafted with 450 allogenic islets. Circulating hAAT levels (1-350 μg/ml) persisted for over 30 days. Normoglycemia was achieved and islets were accepted in 6 out of 6 recipients that expressed hAAT. All 4 control animals exhibited acute islet allograft rejection by day 12. We conclude that plasmid-derived long-term expression of as low as 1 μg/ml hAAT exhibits islet-allograft protection. Using site-directed mutagenesis of pEF-hAAT we will examine whether activities that are independent of protease inhibition partake in the protection of islet allografts.

Free radical scavenging and antielastase activities of flavonoids from the fruits of Thuja orientalis

Bioassay-guided fractionation of the MeOH extract of Thuja orientalis fruits using a DPPH (2,2-diphenyl-1-picrylhydrazyl) assay led to the isolation of 9 flavonoids: cupressuflavone (1), amentoflavone (2), robustaflavone (3), afzelin (4), (+)-catechin (5), quercitrin (6), hypolaetin 7-O-beta-xylopyranoside (7), isoquercitrin (8) and myricitrin (9). Their chemical structures were determined by spectroscopic analyses. The free radical scavenging and human neutrophil elastase (HNE) inhibitory activities were evaluated for the isolated compounds. By DPPH scavenging assay, compounds 5, 6, 7, 8 and 9 showed anti-oxidant activities with IC(50) values of 28.66, 31.19, 18.30, 26.63 and 15.10 microM, respectively. By ABTS [2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt] scavenging assay, these compounds also exhibited potent anti-oxidant activities with IC(50) values of 6.77, 13.96, 6.97, 22.79 and 9.96 microM, respectively. Of note, compounds 1, 2 and 3 showed significant HNE inhibitory activities with IC(50) values of 8.09, 1.27 and 1.33 microM, respectively.