Diclofenac exhibits cytotoxic activity associated with metabolic alterations and p53 induction in ESCC cell lines and decreases ESCC tumor burden in vivo.

Esophageal squamous cell carcinoma (ESCC) is one of the most aggressive forms of human malignancy, often displaying limited therapeutic response. Here, we examine the non-steroidal anti-inflammatory drug diclofenac (DCF) as a novel therapeutic agent in ESCC using complementary in vitro and in vivo models. DCF selectively reduced viability of human ESCC cell lines TE11, KYSE150, and KYSE410 as compared with normal primary or immortalized esophageal keratinocytes. Apoptosis and altered cell cycle profiles were documented in DCF-treated TE11 and KYSE 150. In DCF-treated TE11, RNA-Sequencing identified differentially expressed genes and Ingenuity Pathway Analysis predicted alterations in pathways associated with cellular metabolism and p53 signaling. Downregulation of proteins associated with glycolysis was documented in DCF-treated TE11 and KYSE150. In response to DCF, TE11 cells further displayed reduced levels of ATP, pyruvate, and lactate. Evidence of mitochondrial depolarization and superoxide production was induced by DCF in TE11 and KYSE150. In DCF-treated TE11, the superoxide scavenger MitoTempo improved viability, supporting a role for mitochondrial reactive oxygen species in DCF-mediated toxicity. DCF treatment resulted in increased expression of p53 in TE11 and KYSE150. p53 was further identified as a mediator of DCF-mediated toxicity in TE11 as genetic depletion of p53 partially limited apoptosis in response to DCF. Consistent with the anticancer activity of DCF in vitro, the drug significantly decreased tumor burdene in syngeneic ESCC xenograft tumors and 4-nitroquinoline 1-oxide-mediated ESCC lesions in vivo. These preclinical findings identify DCF as an experimental therapeutic that should be explored further in ESCC.

Redox and biometal status in Wistar rats after subacute exposure to fluoride and selenium counter-effects.

This study aimed to investigate the effect of 150 mg/L sodium fluoride (NaF) on redox status parameters and essential metals [copper (Cu), iron (Fe), and zinc (Zn)] in the blood, liver, kidney, brain, and spleen of Wistar rats and to determine the protective potential of selenium (Se) against fluoride (F-) toxicity. Male Wistar rats were randomly distributed in groups of five (n=5) receiving tap water (control) or water with NaF 150 mg/L, NaF 150 mg/L + Se 1.5 mg/L, and Se 1.5 mg/L solutions ad libitum for 28 days. Fluorides caused an imbalance in the redox and biometal (Cu, Fe, and Zn) status, leading to high superoxide anion (O2 .-) and malondialdehyde (MDA) levels in the blood and brain and a drop in superoxide dismutase (SOD1) activity in the liver and its increase in the brain and kidneys. Se given with NaF improved MDA, SOD1, and O2 .- in the blood, brain, and kidneys, while alone it decreased SH group levels in the liver and kidney. Biometals both reduced and increased F- toxicity. Further research is needed before Se should be considered as a promising strategy for mitigating F- toxicity.

Neuroprotective effect of Biochanin a against Bisphenol A-induced prenatal neurotoxicity in zebrafish by modulating oxidative stress and locomotory defects.

Exogenous toxicants cause oxidative stress and damage to brain cells, resulting in inflammation. Neuroinflammation is important in the pathobiology of various neurological illnesses, including Alzheimer's disease (AD). In this context, Bisphenol A (BPA), a common toxin, causes oxidative damage and has been linked to neurological problems. An O-methylated isoflavone known as Biochanin A (5,7-dihydroxy-4'-methoxy-isoflavone, BCA) is considered to be a phytoestrogen, which is abundant in some legume plants and soy which have preventive effects against cancer, osteoporosis, menopausal symptoms and oxidative stress. However, the mechanism by which BCA protected the prenatal neurological stress are not known. So that, in this study we investigated the BCA neuroprotective effect against BPA-induced neuroinflammation in zebrafish embryo models. For this study, fertilized zebrafish embryos are exposed to BPA (1 µM) with or without BCA. Our finding suggested that BCA co-exposure prevented the depletion of antioxidant defense enzymes by BPA and reduced the production of intracellular ROS production, superoxide anion (O2-), lipid peroxidation (LPO), lactate dehydrogenase (LDH) and nitric oxide (NO) levels in the head that aided in safeguarding neuronal development. Baseline locomotion was rendered and a total distance was calculated to assess the motor function. Exposure to BCA increased acetylcholinestrase (AChE) and improved motor neuron functions. It also reduced the pro-inflammatory response expression and prevented neuroinflammation. Our study suggests that BCA has a positive role in the attenuation or amelioration of neuronal oxidative damage and locomotory behaviour induced by BPA.

Superoxide dismutase: an updated review on its health benefits and industrial applications.

Many short-lived and highly reactive oxygen species, such as superoxide anion (O2-) and hydrogen peroxide (H2O2), are toxic or can create oxidative stress in cells, a response involved in the pathogenesis of numerous diseases depending on their concentration, location, and cellular conditions. Superoxide dismutase (SOD) activities as an endogenous and exogenous cell defense mechanism include the potential use in treating various diseases, improving the potential use in treating various diseases, and improving food-stuffs preparation dietary supplements human nutrition. Published work indicates that SOD regulates oxidative stress, lipid metabolism, inflammation, and oxidation in cells. It can prevent lipid peroxidation, the oxidation of low-density lipoprotein in macrophages, lipid droplets' formation, and the adhesion of inflammatory cells into endothelial monolayers. It also expresses antioxidant effects in numerous cancer-related processes. Additionally, different forms of SOD may also augment food processing and pharmaceutical applications, exhibit anticancer, antioxidant, and anti-inflammatory effects, and prevent arterial problems by protecting the proliferation of vascular smooth muscle cells. Many investigations in this review have reported the therapeutic ability and physiological importance of SOD. Because of their antioxidative effects, SODs are of great potential in the medicinal, cosmetic, food, farming and chemical industries. This review discusses the findings of human and animal studies that support the advantages of SOD enzyme regulations to reduce the formation of oxidative stress in various ways.

Anticancer Effects and Mechanisms of Action of Plumbagin: Review of Research Advances.

Plumbagin (PLB), a natural naphthoquinone constituent isolated from the roots of the medicinal plant Plumbago zeylanica L., exhibited anticancer activity against a variety of cancer cell lines including breast cancer, hepatoma, leukemia, melanoma, prostate cancer, brain tumor, tongue squamous cell carcinoma, esophageal cancer, oral squamous cell carcinoma, lung cancer, kidney adenocarcinoma, cholangiocarcinoma, gastric cancer, lymphocyte carcinoma, osteosarcoma, and canine cancer. PLB played anticancer activity via many molecular mechanisms, such as targeting apoptosis, autophagy pathway, cell cycle arrest, antiangiogenesis pathway, anti-invasion, and antimetastasis pathway. Among these signaling pathways, the key regulatory genes regulated by PLB were NF-kß, STAT3, and AKT. PLB also acted as a potent inducer of reactive oxygen species (ROS), suppressor of cellular glutathione, and novel proteasome inhibitor, causing DNA double-strand break by oxidative DNA base damage. This review comprehensively summarizes the anticancer activity and mechanism of PLB.

HPLC-DAD-ESI/MSn phenolic profile and in vitro biological potential of Centaurium erythraea Rafn aqueous extract.

Centaurium erythraea Rafn is a food flavouring used in both food and beverage industries. Despite the existence of some works on this species, only few focus their attention on its common form of consumption (infusion). Thus, we intended to explore the chemical composition and the biological properties of infusions from C. erythraea aerial parts. Phenolic compounds were identified by HPLC-DAD-ESI/MSn, twenty-two flavonoid derivatives being reported for the first time. Concerning to the biological potential, the best scavenging activity was observed for the superoxide anion radical. Moreover, this extract revealed to have some effect on the inhibition of 5-lipoxygenase, no cytotoxicity against RAW 264.7 macrophage cells line, and a weak potential to decrease the nitric oxide levels in this cell system. The results suggest that C. erythraea aerial parts could be valuable sources of health benefits compounds, contributing to its valorisation and its further application in functional foods.

Intracellular hydrogen peroxide and superoxide poison 3-deoxy-D-arabinoheptulosonate 7-phosphate synthase, the first committed enzyme in the aromatic biosynthetic pathway of Escherichia coli.

In Escherichia coli, aromatic compound biosynthesis is the process that has shown the greatest sensitivity to hydrogen peroxide stress. This pathway has long been recognized to be sensitive to superoxide as well, but the molecular target was unknown. Feeding experiments indicated that the bottleneck lies early in the pathway, and the suppressive effects of fur mutations and manganese supplementation suggested the involvement of a metalloprotein. The 3-deoxy-D-arabinoheptulosonate 7-phosphate synthase (DAHP synthase) activity catalyzes the first step in the pathway, and it is provided by three isozymes known to rely upon a divalent metal. This activity progressively declined when cells were stressed with either oxidant. The purified enzyme was activated more strongly by ferrous iron than by other metals, and only this metalloform could be inactivated by hydrogen peroxide or superoxide. We infer that iron is the prosthetic metal in vivo. Both oxidants displace the iron atom from the enzyme. In peroxide-stressed cells, the enzyme accumulated as an apoprotein, potentially with an oxidized cysteine residue. In superoxide-stressed cells, the enzyme acquired a nonactivating zinc ion in its active site, an apparent consequence of the repeated ejection of iron. Manganese supplementation protected the activity in both cases, which matches the ability of manganese to metallate the enzyme and to provide substantial oxidant-resistant activity. DAHP synthase thus belongs to a family of mononuclear iron-containing enzymes that are disabled by oxidative stress. To date, all the intracellular injuries caused by physiological doses of these reactive oxygen species have arisen from the oxidation of reduced iron centers.

Further in vitro evaluation of antiradical and antimicrobial activities of phytol.

The antiradical activity of phytol was evaluated by electron paramagnetic resonance towards hydroxyl radical (·OH), superoxide anion radical (·O2(-)), methoxy radical (·CH2OH), carbon-dioxide anion radical (·CO2(-)), as well as towards nitric-oxide radical (·NO) and 2,2-diphenyl-1-picrylhydrazyl (·DPPH) radical. It reduced the production of all tested radicals showing more promising activity against ·CO2(-), ·CH2OH and ·DPPH radicals (56%, 50% and 48%, respectively) in comparison with ·NO, ·O2(-) and ·OH radicals (38%, 23% and 15%, respectively). The antimicrobial activity of phytol was evaluated by the microdilution method against eight bacterial and eight fungal strains. To varying degrees, it was proven to be active against all tested bacteria and fungi (MIC 0.003-0.038 mg/mL and MBC 0.013-0.052 mg/mL, MIC 0.008-0.016 mg/mL and MFC 0.090-0.520 mg/mL, respectively). According to the obtained results, medical foods containing phytol may support development of new therapies for heart disease.

Antiradical, antimicrobial and cytotoxic activities of commercial beetroot pomace.

The by-product of food processing is often utilized as feed, and for the preparation of dietary fiber and biofuel. However, these products are also promising sources of bioactive antioxidants and color giving compounds, which could be used as additives in the food, pharmaceutical and cosmetic industry. The aim of this study was to investigate the phytochemical profile, and the antiradical, antimicrobial and cytotoxic activities of industrial beetroot pomace extract (BPE). The content of phenolics (45.68 mg gallic acid equivalents g(-1)), flavonoids (25.89 mg rutin equivalents g(-1)) and betalains (4.09 mg betanin g(-1); 7.32 mg vulgaxanthin I g(-1)) were determined spectrophotometrically. The antiradical activity on DPPH (EC(50)(DPPH·) = 0.0797 mg ml(-1)), hydroxyl (EC(50)(·OH) = 0.0655 mg ml(-1)) and superoxide anion (EC(50)(O2·-) = 1.0625 mg ml(-1)) radicals were measured by electron spin resonance (ESR) spectroscopy. The antimicrobial activity was determined using the agar-well diffusion method. Gram(-) bacteria (Salmonella typhimurium, Citrobacter freundii) and Gram(+) bacteria, (Staphylococcus aureus, Staphylococcus sciuri, Bacillus cereus) showed high susceptibility, while yeasts and moulds were resistant. BPE exhibits cytotoxic properties against Ehrlich carcinoma (EAC) cells in vivo due to induction of oxidative stress. The largest decreases in EAC cell numbers were observed in the pre-treated male (approximately 53%) and female (approximately 47%) mice, and also the EAC cell viability was decreased after administration of BPE. The activities of the antioxidant enzymes, xanthine oxidase (XOD) and peroxidase (Px), were significantly different between the untreated EAC control group and all other groups that were treated with BPE. The XOD and Px activities were very low in untreated malignant cells, but increased significantly after administration of BPE. Our results show that BPE holds promise in the food industry as a source of bioactive compounds.

Purification and characterization of a novel ~18 kDa antioxidant protein from Ginkgo biloba seeds.

Ginkgo biloba seeds are widely used as a food and traditional medicine in China. In the present study, a novel antioxidant protein named GBSP was purified from Ginkgo biloba seeds. The protein (GBSP) was purified by homogenization of Ginkgo biloba seed powder in saline solution, 70% ammonium sulphate precipitation, filtration on a DEAE-Cellulose52 anion exchange column, gel filtration on a Sephadex G-50 column, and preparative chromatography on a C(18) column using RP-HPLC. GBSP showed an apparent molecular weight of 18 kDa by SDS-PAGE and MALDI-TOF/MS analyses. The amino acid sequence obtained by MALDI-TOF/TOF MS analysis showed GBSP was a novel protein, as no matching protein in was found the database. The protein exhibited significant antioxidant activities against free radicals such as DPPH, ABTS and superoxide anion and showed higher activity than α-tocopherol in a linoleic acid emulsion assay system. Furthermore, GBSP exhibited notable reducing power and a strong chelating effect on Cu(2+) and Fe(2+). Therefore, the present study demonstrates, for the first time, that this novel protein from Ginkgo biloba seeds is an excellent antioxidant.

Tetrahydrobiopterin protects the kidney from ischemia-reperfusion injury.

Tetrahydrobiopterin (BH4) is an essential cofactor for the nitric oxide (NO) synthases and represents a critical determinant of NO production. BH4 depletion during ischemia leads to the uncoupling of the synthases, thus contributing to reperfusion injury due to increased superoxide formation. To examine whether BH4 supplementation attenuates ischemia-reperfusion injury, we clamped the left renal arteries of male Lewis rats immediately following right-side nephrectomy. BH4 tissue levels significantly decreased after 45 min of warm ischemia compared with levels in non-ischemic controls. Histopathology demonstrated significant tubular damage and increased peroxynitrite formation. Intravital fluorescent microscopy found perfusion deficits in the microvasculature and leakage of the capillary mesh. Supplemental BH4 treatment before ischemia significantly reduced ischemia-induced renal dysfunction, and decreased tubular histologic injury scores and peroxynitrite generation. BH4 also significantly improved microcirculatory parameters such as functional capillary density and diameter. These protective effects of BH4 on microvasculature were significantly correlated with its ability to abolish peroxynitrite formation. We suggest that BH4 significantly protects against acute renal failure following ischemia reperfusion. Whether BH4 has a therapeutic potential will require more direct testing in humans.

Silibinin induces protective superoxide generation in human breast cancer MCF-7 cells.

The pharmacological activity of polyphenolic silibinin from milk thistle (Silybum marianum) is primarily due to its antioxidant property. However, this study found that silibinin promoted sustained superoxide (O(2)(.-)) production that was specifically scavenged by exogenous superoxide dismutase (SOD) in MCF-7 cells, while the activity of endogenous SOD was not changed by silibinin. Previous work proved that silibinin induced MCF-7 cell apoptosis through mitochondrial pathway and this study further proved that O(2)(.-) generation induced by silibinin was also related to mitochondria. It was found that respiratory chain complexes I, II and III were all involved in silibinin-induced O(2)(.-) generation. Moreover, it was found that silibinin-induced O(2)(.-) had protective effect, as exogenous SOD markedly enhanced silibinin-induced apoptosis.

The in vitro effects of superoxide, some commercially available antioxidants and red palm oil on sperm motility.

In this study, two commercially available superoxide scavengers, tetrakis (1-methyl-4-pyridyl) porphyrin (Mn[III]TMPyP) and superoxide dismutase (SOD), as well as red palm oil (RPO), a natural vegetable oil, had been used to investigate their possible in vitro effects against the toxic effects of superoxide (O(2).) on human sperm motility. Semen samples were obtained from 12 normozoospermic healthy volunteer donors aged between 19 and 23 years. The O(2). donor 2,3-dimetoxyl-1,4-naphthoquinone (DMNQ) (2.5 micromol L(-1)-100 micromol L(-1)) was added to normozoospermic post-swim-up sperm in the presence or absence of Mn(III)TMPyP (50 micromol L(-1)), SOD (50 IU) or RPO (0.1% or 0.5%). Computer-assisted semen analysis was used to analyze various motility parameters. The parameters of interest were percentage of motile cells, progressive motility, rapid cells and static cells. Concentrations of higher than 25 micromol L(-1) DMNQ were detrimental to sperm motility. Mn(III)TMPyP was able to attenuate the effect of O(2). on the motility parameters. In vitro addition of SOD and RPO showed harmful effects on sperm motility.

C-geranylated chalcones from the stems of Angelica keiskei with superoxide-scavenging activity.

An ethyl acetate-soluble fraction of a methanol extract from the stems of Angelica keiskei was subjected to chromatographic separation to give three new chalcones, designated as xanthokeismins A-C (1-3), in addition to a known chalcone, xanthoangelol B (4). The structures of 1-3 were determined on the basis of the analysis of their spectroscopic data. Compounds 1-4 exhibited potent superoxide-scavenging activity.

Antioxidant properties of Viburnum opulus and Viburnum lantana growing in Turkey.

In the present study, antioxidant properties of the water extracts of different parts of Viburnum opulus and Viburnum lantana (Caprifoliaceae) were investigated using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging and superoxide anion scavenging methods. The extracts were prepared from the fruits, branches, and leaves of V lantana and V opulus species. The branch extracts of V lantana and V opulus inhibited superoxide anion in a concentration-dependent manner. Compared with t-tocopherol, the fruit extract of V lantana did not show any scavenging effect on superoxide anion formation. V lantana leaf extracts, however, showed a moderate scavenging effect on superoxide anion formation, whereas V lantana branch extracts showed a strong scavenging effect (IC50 = 3.1 mg/ml) on superoxide anion in higher concentration. On the other hand, all extracts exhibited a scavenging effect on the DPPH radical with various potencies. When compared with butylated hydroxytoluene, V opulus branch and V lantana leaf extracts, as well as V lantana branch, V opulus fruit and V lantana fruit extracts, showed strong DPPH radical scavenging activity with IC50 values of 0.014, 0.035, 0.052, 0.057 and 0.085 mg/ml, respectively.

Stictic acid derivatives from the lichen Usnea articulata and their antioxidant activities.

Two new beta-orcinol depsidones, 1 and 2, together with 13 known compounds were isolated from the lichen Usnea articulata. The structures of 1 and 2 were elucidated by spectroscopic analyses and those of known compounds by comparison of their spectroscopic data with literature values or by direct comparison with authentic standards. Compounds 1, 2, and 5 exhibited moderate antiradical activity in the 1,1-diphenyl-2-picrylhydrazyl (DPPH) assay. The depsidones 4 and 5 showed better superoxide anion scavenging activity (IC50 = 566 and 580 microM, respectively) than quercetin (IC50 = 754 microM).

Resistance of Lactobacillus plantarum KCTC 3099 from Kimchi to oxidative stress.

The antioxidative capacity of two lactic acid bacteria isolated from Kimchi, a Korean fermented food, was evaluated by measuring the resistance to reactive oxygen species (ROS) and compared with that of Lactobacillus rhamnosus GG as a positive control. Both intact cells and cell-free extracts of Lactobacillus plantarum KCTC 3099 exhibited higher antioxidative activity in inhibiting lipid peroxidation among the strains evaluated with an inhibitory level of 38.6% and 48.5%, respectively. To evaluate the resistance of the two lactic acid bacteria to ROS, we tested their survival in the presence of 1 mM hydrogen peroxide, 0.4 mM hydroxyl radicals, and superoxide anions induced by 10 mM paraquat. L. plantarum KCTC 3099 was viable even after 8 hours in the presence of both 1 mM hydrogen peroxide and 0.4 mM hydroxyl radicals. Moreover, the survival of L. plantarum KCTC 3099 was not affected by superoxide anions generated by using paraquat, indicating that it has resistance to superoxide anions. To define the antioxidative mechanism, superoxide dismutase (SOD) and metal ion chelating activities were determined. L. plantarum KCTC 3099 presented little SOD activity, but had the higher level of chelating activity for both Fe2+ and Cu2+ metal ions at 13.6 ppm and 23.9 ppm, respectively. These results suggested that the antioxidative capacity of L. plantarum KCTC 3099 is apparently caused by chelating metal ions rather than by SOD activation.

Effect of endocrine disruptor para-nonylphenol on the cell growth and oxygen radical generation in Escherichia coli mutant cells deficient in catalase and superoxide dismutase.

para-Nonylphenol (NP) had previously been found to have strong suppressive effects of growth of bacterial and yeast cells, and these effects were associated with NP-induced generation of radical oxygen species (ROS). In the present study, we determined that wild-type strains of Escherichia coli (CSH 7, SY-11, and IFO-3545) were resistant to NP compared with other sensitive microorganisms reported previously. To elucidate the relationship between NP-induced ROS generation and cell growth inhibition in more detail, we analyzed the effect of NP on cell growth and survival of wild-type and mutant E. coli strains deficient in ROS-scavenging enzymes such as catalase and superoxide dismutase (SOD). The SOD-deficient strain QC 774 (sod A- and sod B-) was much more sensitive to NP than wild-type (CSH 7) and catalase-deficient (UM 1 kat E- and kat G-) strains. As a comparative experiment, when hydrogen peroxide was applied to the same growth and survival assays, UM 1 cells were more sensitive to hydrogen peroxide than QC 774 and CSH 7. A chemiluminescence (CHL) experiment using MCLA (2-methyl-6-Lf-methylphenyl]-3,7-dihydroimidazc [1,2-alpha] pyrazin-3-one) reflecting predominantly superoxide generation showed that NP caused marked CHL generation in QC 774 cells, but not in CSH 7 and UM 1 cells. However, the CHL experiment using L-012 reflecting predominantly hydroxyl radical and hypochlorite did not exhibit significant CHL generation in QC 774 cells at the same concentrations of NP. Furthermore, supplementation with SOD prevented NP-induced ROS generation and cell survival inhibition of QC 774 cells, but the catalase and metal-chelating agent deferoxamine did not have significant effects. These results suggest that one of the primary actions of NP in cells is the generation of superoxide which may be responsible for NP-induced cell growth inhibition.

Reactive oxygen species promote tyrosine phosphorylation and capacitation in equine spermatozoa.

The objective of this study was to examine the influence of reactive oxygen species (ROS) on equine sperm capacitation. Motile equine spermatozoa were separated on a discontinuous Percoll gradient, resuspended at 10 x 10(6)ml in Tyrode's medium supplemented with BSA (0.5%) and polyvinyl alcohol (0.5%) and incubated at 39 degrees C for 2h with or without the xanthine (X; 0.1mM)-xanthine oxidase (XO; 0.01 U/ml) system or NADPH (0.25 mM). The importance of hydrogen peroxide or superoxide for capacitation was determined by the addition of catalase (CAT; 150 U/ml) or superoxide dismutase (SOD; 150 U/ml), respectively. Following incubation, acrosomal exocytosis was induced by a 5 min incubation at 39 degrees C with progesterone (3.18 microM), and sperm viability and acrosomal integrity were then determined by staining with Hoechst 33258 and fluoroisothiocyanate-conjugated Pisum sativum agglutin. To examine tyrosine phosphorylation, treatments were subjected to sodium dodecyl sulfate-polyacrylaminde gel electrophoresis (SDS-PAGE) followed by Western blot analysis with the anti-phosphotyrosine antibody (alpha-PY; clone 4G10). Capacitation with the X-XO system or NADPH led to a significant (P<0.0001) increase in live acrosome-reacted spermatozoa compared to controls. The addition of CAT or SOD prevented the increase in live acrosome-reacted spermatozoa associated with X-XO treatment. Incubation with the X-XO system was also associated with a significant (P<0.005) increase in tyrosine phosphorylation when compared to controls, which could be prevented by the addition of CAT but not SOD. This study indicates that ROS can promote equine sperm capacitation and tyrosine phosphorylation, suggesting a physiological role for ROS generation by equine spermatozoa.

A neutrophil multitarget functional bioassay to detect anti-inflammatory natural products.

A multitarget functional bioassay was optimized as a method for detecting substances interacting with the inflammatory process of activated neutrophil granulocytes, mainly to release elastase detected by p-nitroanilide (pNA) formation. Using this bioassay, 100 fractionated extracts of 96 plants were screened, with results presented in a manner that links recorded biological activity to phylogenetic information. The plants were selected to represent a major part of the angiosperms, with emphasis on medicinal plants, Swedish anti-inflammatory plants, and plants known to contain peptides. Of the tested extracts, 41% inhibited pNA formation more than 60%, and 3% stimulated formation. The extract of Digitalis purpurea enhanced pNA formation, and digitoxin, the active compound, was isolated and identified. Plant extracts that exhibited potent nonselective inhibition (>80% inhibition) were evaluated further for direct inhibition of isolated elastase and trypsin enzyme. The inhibitory effect of most tested extracts on the isolated enzyme elastase was similar to that of PAF- and fMLP-induced pNA formation. Compared to trypsin, inhibition of elastase by extracts of Rubus idaeus and Tabernaemontana dichotoma was significantly higher (80% and 99%, respectively). Inhibition of trypsin by the extract of Reseda luteola was high (97%). Orders such as Lamiales and Brassicales were shown to include a comparably high proportion of plants with inhibitory extracts.