Assessing the risk of epidemic dropsy from black salve use

Epidemic dropsy is a potentially life‐threatening condition resulting from the ingestion of argemone oil derived from the seeds of Argemone mexicana Linn. Exposure to argemone oil is usually inadvertent, arising from mustard cooking oil adulteration. Sanguinarine, an alkaloid present in argemone oil, has been postulated as a causative agent with the severity of epidemic dropsy correlating with plasma sanguinarine levels. Cases of epidemic dropsy have also been reported following the topical application of argemone containing massage oil. Black salve, a topical skin cancer therapy also contains sanguinarine, but at significantly higher concentrations than that reported for contaminated massage oil. Although not reported to date, a theoretical risk therefore exists of black salve inducing epidemic dropsy. This literature review explores the presentation and pathophysiology of epidemic dropsy and assesses the risk of it being induced by black salve.     

In vivo DNA damaging potential of sanguinarine alkaloid, isolated from argemone oil, using alkaline Comet assay in mice.

Consumption of mustard oil contaminated with argemone oil is well known to cause clinical manifestation referred to as "Epidemic Dropsy". Our prior studies have shown that argemone oil produces genotoxic effects in mice [Ansari, K.M., et al., 2004. Int. J. Cancer 112, 890]. Since, sanguinarine alkaloid is the major component of argemone oil, the in vivo DNA damaging potential of the isolated alkaloid was investigated in blood and bone marrow cells of mice using alkaline Comet assay. Swiss albino male mice were given single intraperitoneal administration of 1.35, 2.70, 5.40, 10.80 and 21.60 mg sanguinarine alkaloid/kg b wt., while controls were treated with saline in the same manner. The results revealed a dose dependent increase in DNA damage in blood and bone marrow cells following 24 h treatment of sanguinarine alkaloid. All the three parameters of Comet assay including olive tail moment (OTM), tail length and tail DNA showed significant (p<0.05) increases in blood and bone marrow cells at respective doses of 10.80 and 5.40 mg alkaloid/kg b wt. However, some of the parameters were significantly increased even at lower doses of sanguinarine alkaloid (2.70 mg/kg b wt.). The frequency of cells exhibiting greater DNA damage were found to be increased by sanguinarine alkaloid in a concentration dependent manner. These results indicate that single exposure of sanguinarine alkaloid causes DNA damage in blood and bone marrow cells of mice, which could be responsible for the genotoxicity of argemone oil.     

Biochemical toxicology of argemone oil. I. Effect on hepatic cytochrome P-450 and xenobiotic metabolizing enzymes

The in vivo effect of argemone oil on hepatic xenobiotic metabolizing enzymes was investigated in albino rats following either a single (10 ml kg-1 body wt.) or multiple intraparenteral doses (5 ml kg-1 body wt.) for three days. Animals sacrificed 72 h after a single intraparenteral dose of argemone oil exhibited a significant loss of hepatic cytochrome P-450 (35%) and cytochrome b5 (34%) contents and inhibition of aminopyrine-N-demethylase (APD), aryl hydrocarbon hydroxylase (AHH) and ethoxycoumarin-O-deethylase (ECD) activities (21-39%). Three successive 24-hourly intraparenteral injections of argemone oil followed by sacrificing the animals after 24 h of the last injection, showed a greater degree of inhibition of the content of cytochrome P-450 (58%) and its dependent mixed-function oxidases (35-63%). Also, multiple treatment of argemone oil caused a depletion of endogenous hepatic glutathione (GSH) content (72%) with a concomitant increase in lipid peroxidation (177%) and decrease in glutathione-S-transferase (GST) activity (30%). A significant decrease in relative liver weight (39%) was observed in animals treated with multiple treatment of argemone oil. These results suggest that argemone oil can alter both membrane and cytosolic defences and destabilizes the hepatic cytochrome P-450 dependent mixed-function oxidase system, so that it tips in the direction of autooxidative peroxidation of lipids.     

Skin tumor promotion by argemone oil/alkaloid in mice: Evidence for enhanced cell proliferation,ornithine decarboxylase,cyclooxygenase-2 and activation of MAPK/NF-κB pathway

Consumption of argemone oil (AO) contaminated edible oil causes “Epidemic Dropsy”. Previously, we have shown that AO and isolated sanguinarine possess genotoxicity and skin tumor initiating activity. Here, we evaluate tumor-promoting potential of AO/sanguinarine alkaloid and investigate the molecular mechanisms involved therein. Single topical application of AO (50–400 μl/mouse) or sanguinarine alkaloid (1.5–12.0 μmol/mouse) afforded significant increase in (i) ornithine decarboxylase (ODC) activity, (ii) uptake of [³H]-thymidine in DNA, (iii) cyclooxygenase-2 (COX-2), proliferating cell nuclear antigen (PCNA) and ODC protein expressions, (iv) phosphorylation of extracellular signal-regulated kinase (ERK)1/2, c-jun-N-terminal kinase (JNK)1/2 and p38 mitogen-activated protein (MAP) kinases, (v) increased NF-κB activation and (vi) no significant increase in dark basal keratinocytes. Subsequently, when AO and sanguinarine alkaloid was tested either as complete or stage I or stage II tumor promoter in 7, 12-dimethyl benz(a)anthracene (DMBA)-initiated mice, there was enhanced tumor incidence, tumor body burden and higher % of mice with tumors, when AO (0.1 ml) or isolated sanguinarine (1.5 μmol) was tested as stage II tumor promoter. However, no tumors were found when AO or sanguinarine alkaloid was tested either as complete or stage I tumor promoter. These results indicate that AO/ sanguinarine alkaloid possesses tumor-promoting potential at stage II level involving MAPK/NF-κB pathway.     

Biochemical toxicology of argemone oil. IV. Short-term oral feeding response in rats

Consumption of edible oils contaminated with Argemone mexicana seed oil is known to cause various clinical manifestations. In the present study, the effect of dietary intake of argemone oil on histopathological changes, haematological indices and selected marker parameters of toxicity was investigated to observe the exact sites and mode of action of argemone oil in rats. Histopathological changes in the liver showed increased fibrosis, hyperplasia of bile ducts and congestion in a few portal tracts. Lungs of argemone oil-fed animals indicated congestion and thickening of interalveolar septa. Alveolar spaces were disorganised and irregular. Kidneys showed vascular and glomerular congestion and patchy tubular lesions. At 30 days only mild congestion was noted in the myocardium. Cardiac muscle fibres showed degenerative changes at 60 days which were more marked in the auricular wall. Haematological examination showed appearance of anaemia in experimental animals. Hepatic alkaline phosphatase, alanine transaminase and aspartate transaminase activities were inhibited by 30, 29 and 29% after 30 days of argemone intake along with concomitant enhancement in serum by 27, 29 and 66%, respectively. Liver showed decrease in glutathione (32-63%) content along with significant stimulation of lipid peroxidation (49-105%) in argemone-intoxicated animals. These results suggest that liver, lungs, heart and kidneys are the target tissues of argemone oil toxicity and that membrane destruction may be a possible mode of action.     

Metabolism of sanguinarine: the facts and the myths

Sanguinarine, a quaternary benzo[c]phenanthridine alkaloid, exhibits antimicrobial and anti-inflammatory activities and for this reason it is used in dental hygiene products and feed additives. Its metabolism and disposition is the subject of constant scientific discourse. In this paper we summarize current knowledge on sanguinarine metabolism. We show in particular that: (i) Sanguinarine is not transformed to 3,4-benzacridine and that the literature reporting this compound as a metabolite of sanguinarine is based on artifacts and misinterpretations that in course of time have created a dogma; (ii) Sanguinarine is converted to dihydrosanguinarine in vivo, the conversion being tentatively a detoxication pathway; (iii) Aryl hydrocarbon receptor metabolic signaling pathways modulate sanguinarine biological activity.     

Biochemical toxicology of argemone alkaloids. III. Effect on lipid peroxidation in different subcellular fractions of the liver

Consumption of edible oils contaminated with Argemone mexicana seed oil causes various toxic manifestations. In this investigation the in vivo effect of argemone oil on NADPH-dependent enzymatic and Fe2+-, Fe2+/ADP- or ascorbic acid-dependent non-enzymatic hepato-subcellular lipid peroxidation was studied. Parenteral administration of argemone oil (5 ml/kg body weight) daily for 3 days produced a significant increase in both non-enzymatic and NADPH-supported enzymatic lipid peroxidation in whole homogenate, mitochondria, and microsomes. Lipid peroxidation aided by various pro-oxidants, namely Fe2+, Fe2+/ADP and ascorbic acid also revealed a significant enhancement in the whole homogenate, mitochondria and microsomes of argemone oil-treated rats. Further, when compared with whole homogenate, the hepatic mitochondria and microsomes of either control or argemone oil-treated rats showed a 4- and 6-fold increase in non-enzymatic, and a 5- and 18-fold increase in NADPH-dependent enzymatic lipid peroxidation, respectively. Similarly, both mitochondrial and microsomal fractions showed a 5- and 7-fold increase in Fe2+-, and a 12- and 15-fold increase in either Fe2+/ADP- or ascorbic acid-aided lipid peroxidation, respectively. These results suggest that the hepatic microsomal as well as the mitochondrial membrane is vulnerable to the peroxidative attack of argemone oil and may be instrumental in leading to the hepatotoxicity symptoms noted in argemone poisoning victims.     

Involvement of heme oxygenase-1 in neuroprotection by sanguinarine against glutamate-triggered apoptosis in HT22 neuronal cells

Sanguinarine is a natural compound isolated from the roots of Macleaya cordata and M. microcarpa, has been reported to possess several biological activities such as anti-inflammatory and anti-oxidant effects. In the present study, we demonstrated that sanguinarine markedly induces the expression of HO-1 which leads to a neuroprotective response in mouse hippocampus-derived neuronal HT22 cells from apoptotic cell death induced by glutamate. Sanguinarine significantly attenuated the loss of mitochondrial function and membrane integrity associated with glutamate-induced neurotoxicity. Sanguinarine protected against glutamate-induced neurotoxicity through inhibition of HT22 cell apoptosis. JC-1 staining, which is a well-established measure of mitochondrial damage, was decreased after treatment with sanguinarine in glutamate-challenged HT22cells. In addition, sanguinarine diminished the intracellular accumulation of ROS and Ca²⁺. Sanguinarine also induced HO-1, NQO-1 expression via activation of Nrf2. Additionally, we found that si RNA mediated knock-down of Nrf2 or HO-1 significantly inhibited sanguinarine-induced neuroprotective response. These findings revealed the therapeutic potential of sanguinarine in preventing the neurodegenerative diseases.     

Inhibition of Na,K-ATPase activity and ouabain binding by sanguinarine

Sanguinarine, an isoquinoline alkaloid belonging to the benzophenanthridine class, has been found to be a time-dependent inhibitor of cardiac Na,K-ATPase activity. Enzyme inhibition depended on the concentration of enzyme in the assay. Dilution and centrifugation of the enzyme following incubation with sangunarine did not decrease inhibiton. Inhibiton of ouabain binding by sanguinarine was also time-dependent and resulted in a reduction of the equilibrium level of ouabian bound to the enzyme preparation. This suggests the possibility that the positive inotropic effect of sanguinarine on the heart may be due to interaction with the cardiac glycoside receptor site on Na,K-ATPase. Alternatively, sanguinarine may bind nonspecifically to phospholipids associated with the enzyme and exert its effects via conformational changes.     

Sanguinarine induces apoptosis in A549 human lung cancer cells primarily via cellular glutathione depletion

Sanguinarine is a plant-derived benzophenanthridine alkaloid and has been shown to possess anti-tumor activities against various cancer cells. In this study, we investigated whether sanguinarine induces apoptosis in A549 human lung cancer cells. Treatment of A549 cells with sanguinarine induced apoptosis in a dose- and time-dependent manner. Treatment with sanguinarine led to activation of caspases and MAPKs as well as increased MKP-1 expression. Importantly, pretreatment with z-VAD-fmk, a pan caspase inhibitor suppressed the sanguinarine-induced apoptosis in A549 cells. Moreover, pretreatment with NAC, a sulfhydryl group-containing reducing agent strongly suppressed the apoptotic response and caspase activation to sanguinarine. However, the sanguinarine-mediated cytotoxicity in A549 cells was not protected by pharmacological inhibition of MAPKs or MKP-1 siRNA-mediated knockdown of MKP-1. These results collectively suggest that sanguinarine induces apoptosis in A549 cells through cellular glutathione depletion and the subsequent caspase activation.     

The mechanism of action of sanguinarine against methicillin-resistant Staphylococcus aureus

Sanguinarine is a benzophenanthridine alkaloid derived from the root of Sanguinaria canadensis. It is known to perform a wide spectrum of biological activities. The aim of this study is to examine the antimicrobial actions of sanguinarine against methicillin-resistant Staphylococcus aureus (MRSA). Sanguinarine antimicrobial activity was assessed by broth dilution method; its mechanism of action was investigated by bacteriolysis, detergent or ATPase inhibitors and transmission electron microscopy were used to monitor the survival characteristics and the changes in bacteria morphology. The activity of sanguinarine against MRSA strains ranged from 3.12 to 6.25 μg/ml, while the minimum inhibitory concentrations of the two reference strains are 3.12 μg/ml and 1.56 μg/ml. The treatment of the cells with sanguinarine induced the release of membrane-bound cell wall autolytic enzymes, which eventually resulted in lysis of the cell. The OD(600s) of the suspensions treated with the combination of Tris-(hydroxymethyl) aminomethane and Triton X-100 with sanguinarine were reduced to 40% and 8%, respectively. Transmission electron microsco-py of MRSA treated with sanguinarine showed alterations in septa formation. The predisposition of lysis and the altered morphology seen by transmission electron microscopy suggest that sanguinarine compromises the cytoplasmic membrane.     

Safety evaluation studies on argemone oil through dietary exposure for 90days in rats.

Epidemic dropsy is a disease caused by the consumption of mustard oil contaminated with argemone oil (AO). During 1998 dropsy in New Delhi, which is so far the largest with more than 3000 victims and over 60 deaths, it was enquired at various scientific and regulatory meetings about the maximum tolerated dose of AO. Hence, the present study was aimed to investigate the safety levels of AO in rats. Animals were given AO in diet at a dose of 0.001%, 0.01%, 0.1%, 0.5% and 1% daily for 90 days and the two control groups received the standard diet with and without 1% mustard oil. A decrease in body weight gain (28-31%) was observed in 0.5% and 1% AO groups; while significant increases in relative lungs and liver weight was noticed in respective doses of 0.01% and 0.1% AO groups as well as in higher dosage animals. Reduction in RBC count and hemoglobin content (p<0.05) was noticed in 0.01% and 0.1% AO exposed animals. This effect was more pronounced in higher AO doses. Serum marker enzymes including alanine transaminase (ALT), aspartate transaminase (AST), lactate dehydrogenase (LDH) and alkaline phosphatase (ALP) were found to be significantly elevated in 0.01-1% AO groups. Further, a decrease in albumin/globulin ratio (42-78%) was observed in the serum of 0.01% to higher AO dose groups. The levels of serum triglycerides and VLDL cholesterol were found to be enhanced (p<0.05) in AO treated (0.01-1.0%) animals. Histopathological changes in lung were observed at 0.01% dose of AO while liver, kidney and heart produced changes at 0.1% AO and above doses. None of the parameters were found to be affected in 0.001% AO treated animals. These results suggest that the no observed adverse effect level (NOAEL) dose of AO is 0.001% in rats and considering a factor of 100 for humans for highly toxic compound, the safe limit of 0.00001% (100 ppb or 100 ng AO/g oil) AO can be implicated which shall contain only 0.55% of sanguinarine equivalent to 0.6 ng sanguinarine per gram oil. However, the minimum detectable limit of AO is 5 ppm (equivalent to 5 microg sanguinarine per gram oil) with the present existing HPLC method, thereby suggesting that mustard oil should be absolutely free from AO contamination.     

The anti-inflammatory effects of sanguinarine and its modulation of inflammatory mediators from peritoneal macrophages

The quaternary ammonium salt, sanguinarine (SANG), is of great practical and research interest because of its pronounced, widespread physiological effects, which promote anti-microbial and anti-inflammatory responses in experimental animals. Sanguinarine was originally shown to possess anti-inflammatory properties and it has been used to treat various inflammatory diseases. To gain insight into the anti-inflammatory effect of sanguinarine and its mechanisms of action, we used animal models of acute and chronic inflammation and lipopolysaccharide (LPS)-induced murine peritoneal macrophages to examine the anti-inflammatory function of sanguinarine. Sanguinarine displayed significant anti-inflammatory effects both in vitro and in vivo. Our findings further demonstrated that sanguinarine potently inhibited the expression of inflammatory mediators and inflammation in general. Additionally, our results demonstrated that sanguinarine inhibited the activation of mitogen-activated protein kinase (MAPK), which altered inflammatory mediator synthesis and release in vitro. This study extends our understanding of the anti-inflammatory activity of sanguinarine in acute and chronic inflammation. Furthermore, our findings provide clarification of the molecular mechanisms underlying the anti-inflammatory activity of sanguinarine, supporting the naturopathic use of sanguinarine for the treatment of various human inflammatory diseases.     

Induction of necrosis and apoptosis to KB cancer cells by sanguinarine is associated with reactive oxygen species production and mitochondrial membrane depolarization

Sanguinarine is a benzopheanthridine alkaloid present in the root of Sanguinaria canadensis L. and Chellidonium majus L. In this study, sanguinarine (2 and 3 microM) exhibited cytotoxicity to KB cancer cells by decreasing MTT reduction to 83% and 52% of control after 24-h of exposure. Sanguinarine also inhibited the colony forming capacity (>52-58%) and growth of KB cancer cells at concentrations higher than 0.5-1 microM. Short-term exposure to sanguinarine (>0.5 microM) effectively suppressed the adhesion of KB cells to collagen and fibronectin (FN). Sanguinarine (2 and 3 microM) induced evident apoptosis as indicated by an increase in sub-G0/G1 populations, which was detected after 6-h of exposure. Only a slight increase in cells arresting in S-phase and G2/M was noted. Induction of KB cell apoptosis and necrosis by sanguinarine (2 and 3 microM) was further confirmed by Annexin V-PI dual staining flow cytometry and the presence of DNA fragmentation. The cytotoxicity by sanguinarine was accompanied by an increase in production of reactive oxygen species (ROS) and depolarization of mitochondrial membrane potential as indicated by single cell flow cytometric analysis of DCF and rhodamine fluorescence. NAC (1 and 3 mM) and catalase (2000 U/ml) prevented the sanguinarine-induced ROS production and cytotoxicity, whereas dimethylthiourea (DMT) showed no marked preventive effect. These results suggest that sanguinarine has anticarcinogenic properties with induction of ROS production and mitochondrial membrane depolarization, which mediate cancer cell death.     

Degradation of cytochrome P-450 heme in ascorbic acid-deficient guinea pigs.

Degradation of hepatic cytochromc P-450 heme in ascorbic acid-deficient and ascorbic acid-dosed (p.o., 25 mg/100 g/body wt/day) guinea pigs was investigated by determining the turnover of radioactive cytochrome P-450 and the formation of in vivo expired carbon monoxide (¹⁴CO). ¹⁴CO is a specific degradation product of the labeled heme. Hepatic levels of cytochrome P-450 and cytochrome b₅ were decreased significantly (61 per cent, P < 0.001. and 29 per cent. P < 0.01, respectively) in scorbutic animals compared to ascorbic acid-adequate animals. After the administration of δ-amino-levulinic acid [3, 5-³H], the disappearance of radioactivity from the fast-phase and slow-phase components of cytochrome P-450 heme (CO-binding pigments) exhibited half-lives of 4.2 and 30.8 hr, respectively, in the ascorbic acid-depleted guinea pigs. The turnover of cytochrome P-450 heme was similar for both phases in animals fed the ascorbic acid-deficient diet and dosed with ascorbic acid or animals fed a normal stock diet. In guinea pigs deficient in ascorbic acid and injected with [5-¹⁴C]δ-aminolevulinic acid, the cumulative expired ¹⁴CO was similar to guinea pigs adequate in ascorbic acid. These studies demonstrate that the decrease in hepatic cytochrome P-450 content in ascorbic acid deficiency is not due to a defect in cytochrome heme catabolism. The role of ascorbic acid in stabilizing cytochrome P-450 remains to be determined.     

Disposition of sanguinarine in the rat

Sanguinarine is an alkaloid with known antibiotic and anti-inflammatory activity and its pharmacokinetics have been studied in the rat after a single oral dose (10?mg?kg^?1 body weight). Alkaloid determination in the plasma and liver was carried out by high-performance liquid chromatography–electrospray ionization mass spectrometry (HPLC/ESI-MS). The pharmacokinetic parameters (t_max, c_max, AUC_0→t and AUC_0→∞) were determined for sanguinarine and dihydrosanguinarine, the major components detected in plasma. The first step in sanguinarine metabolism in the rat was the reduction of the iminium bond resulting in formation of the less toxic dihydrosanguinarine. Both compounds were completely eliminated from the plasma and liver after 24?h and not detected in urine. After a single oral dose of ³H-sanguinarine, more than 42% of the ingested radioactivity was present in gastrointestinal tract. Benz[c]acridine, up to date the only sanguinarine metabolite referred to in the literature, was not detected in the plasma, liver or urine.     

Effect of pyrrolizidine alkaloids from tansy ragwort (Senecio jacobaea) on hepatic drug-metabolizing enzymes in male rats

Ad lib. consumption of diets containing 5% tansy ragwort (Senecio jacobaea) for 1–4 weeks produced a 5- to 6-fold increase in hepatic microsomal epoxide hydrase and significant increases in cytosolic glutathione S-transferase activities in male Long-Evans rats. An enhancement of these enzyme activities was also observed when a diet containing 1% tansy ragwort was fed for a period of 3 weeks. Feeding a diet containing 0.5% pyrrolizidine alkaloid (PA) mixture extracted from tansy ragwort for 1 week produced a 5-fold increase in hepatic epoxide hydrase and a 73 per cent increase in glutathione S-transferase activities. In contrast, hepatic microsomal aryl hydrocarbon hydroxylase activity (AHH) was reduced significantly by feeding diets containing 5% tansy ragwort or a 0.5% alkaloid mixture. Hepatic microsomal cytochrome P-450 content was lowered following consumption of the 0.5% alkaloid mixture but not by feeding a 5% tansy ragwort diet, the difference presumably being a result of the lowered PA intake by the latter animals. Exposure to the pyrrolizidine alkaloids, therefore, may influence significantly the capacity of animals to metabolize endogenous or foreign compounds and possibly also affect the subsequent biotransformation and toxicity of these plant constituents.     

Effects of 5-Bromo-2'Deoxyuridine (BrdU) Implants on Hepatic Cytochrome P-450 Content and Beta-Oxidation Activity in Rats and Mice

ABSTRACT

Standard regulatory toxicity tests are frequently supplemented with additional compound specific analysis. Analysis of hepatic cytochrome P-450 content, hepatic β-oxidation activity (biochemical analysis), and cell proliferation rates are examples of these analyses that are included when past experience or similarity to other compounds, suggest that a presently tested compound may have an effect. Until now, separate subsets of animals have been designated for cell proliferation analysis and biochemical analysis, because it was unknown if implantation of 5-bromo-2'deoxyuridine (BrdU) filled osmotic pumps (BrdU implants) would effect the rate of hepatic-β or hepatic cytochrome P-450 content.

The purpose of the current study was to determine if BrdU implants had an effect on hepatic cytochrome P-450 content, β-oxidation activity, or the measurement of these enzymes in rats and mice. The BrdU was administered through subcutaneous osmotic pump implants.

The rate of hepatic peroxisomal p-oxidation was not altered in male or female rats or mice with the BrdU implants when compared to those of the control groups. The total hepatic cytochrome P-450 content was also not altered in male or female rats or mice with the BrdU implants when compared to those of the control groups.

BrdU implants do not appear to have an effect on the rate of hepatic peroxisomal β-oxidation or the total hepatic cytochrome P-450 content in male or female rats and mice. It can be concluded that in future studies, rats or mice which are designated for cell proliferation analysis using BrdU implants are also suitable for use in evaluating chemically induced effects on hepatic peroxisomal β-oxidation activity and/or total hepatic cytochrome P-450 content.     

Sanguinarine induces apoptosis of HT-29 human colon cancer cells via the regulation of Bax/Bcl-2 ratio and caspase-9-dependent pathway

Sanguinarine is an alkaloid obtained from the bloodroot plant Sanguinaria canadensis and has beneficial effects on oxidative stress and inflammatory disorders. Previous reports have demonstrated that sanguinarine also exhibit anticancer properties. In the current study, we investigated the effects of sanguinarine on HT-29 human colon cancer cells. It was observed that sanguinarine treatment induces a dose-dependent increase in apoptosis of human colon cancer cells. We also investigated the effects of sanguinarine on the expression of apoptosis-associated proteins, and the results revealed that there was an increase in Bax and a decrease in B-cell lymphoma 2 (Bcl-2) protein levels. Moreover, sanguinarine treatment significantly increases the activation of caspases 3 and 9 that are the key executioners in apoptosis. Our results suggest that sanguinarine induces apoptosis of HT-29 human colon cancer cells and may have a potential therapeutic use in the treatment of human colon cancer.