Anti-Inflammatory Flavonoids from Agrimonia pilosa Ledeb: Focusing on Activity-Guided Isolation.

To elucidate the anti-inflammatory properties and constituents of Agrimonia pilosa Ledeb. (A. pilosa), a comprehensive investigation was conducted employing activity-guided isolation. The anti-inflammatory effects were evaluated through an in vitro nitric oxide (NO) assay on lipopolysaccharide (LPS)-treated RAW 264.7 macrophage cells. Seven bio-active compounds with anti-inflammatory properties were successfully isolated from the butanol fraction and identified as follows: quercetin-7-O-ß-d-rhamnoside (1), apigenin-7-O-ß-d-glucopyranoside (2), kaempferol-7-O-ß-d-glucopyranoside (3), quercetin (4), kaempferol (5), apigenin (6), and apigenin-7-O-ß-d-glucuronide-6″-butylester (7). All isolated compounds showed strong NO inhibitory activity with IC50 values ranging from 1.4 to 31 µM. Compound 6 demonstrated the most potent NO inhibition. Compound 7, a rare flavonoid, was discerned as a novel anti-inflammatory agent, ascertained through its inaugural demonstration of nitric oxide inhibition. Subsequently, a comprehensive structure-activity relationship (SAR) analysis was conducted employing eight flavonoids derived from A. pilosa. The outcomes elucidated that flavones exhibit superior NO inhibitory effects compared to flavonols, and the aglycone form manifests greater potency in NO inhibition than the glycone counterpart. These results highlight A. pilosa as a promising source of effective anti-inflammatory agents and indicate its potential as a health-beneficial dietary supplement and therapeutic material.

Effect of preoperative nutritional support in malnourished patients with pancreatobiliary cancer: a quasi-experimental study.

BACKGROUND: In malnourished patients, postoperative morbidity, hospitalization period, and medical expenses are reportedly to be high. We evaluated the clinical impact of a preoperative nutritional support program (PNSP) among malnourished cancer patients. METHODS: For this quasi-experimental study, we enrolled 90 patients who underwent major pancreatobiliary cancer surgery. Malnutrition was defined as at least one of the following: (1) Patient-Generated Subjective Global Assessment (PG-SGA) grade B or C; (2) > 10% weight loss within 6 months; (3) body mass index <18.5 kg/m2; and (4) serum albumin level < 3.0 g/dL. Forty-five malnourished patients allocated to the PNSP group received in-hospital PNSP for a median of 6 (4-35) days. In the PNSP group, the nutrition support team calculated the patients' daily nutritional requirements based on their nutritional status and previous day's intake. The supplementation targets were as follows: total calorie intake, 30-35 kcal/kg/day; protein intake, 1.2-1.5 g/kg/day; and lipid intake, 1-1.5 g/kg/day. Patients who did not meet the diagnostic criteria for malnutrition were allocated to the well-nourished group and underwent surgery without receiving the PNSP (n = 45). We compared the perioperative nutritional indices (as measured using PG-SGA), postoperative outcome, and quality of life (QOL) according to the European Organization for Research and Treatment of Cancer Quality of Life Questionnaire version 3.0. RESULTS: In the PNSP group, the proportion of patients with serum prealbumin <16 mg/dL decreased significantly after PNSP (29.5% vs. 8.9%, p = 0.013). Moreover, patients with PG-SGA grade A had a statistically significant increase (2.2% vs. 50%, p < 0.001). The overall and major complication rates were higher in the PNSP group than in the well-nourished group without significance (51.1%, 33.3%; 42.2%, 26.7%, respectively). However, the overall and major complication rates were similar between the subgroup with PG-SGA improvement after PNSP and the well-nourished group (40.9% vs. 42.2%, p = 0.958; 27.3% vs. 26.7%, p = 0.525, respectively). QOL indicators in the PNSP group were comparable with those in the well-nourished group after PNSP. CONCLUSION: PNSP may improve perioperative nutritional status and clinical outcomes among malnourished patients with pancreatobiliary cancer.

Prevalence of Psychological Symptoms in Patients Undergoing Pancreatoduodenectomy and Results of a Distress Management System: A Clinic-Based Study.

PURPOSE: Patients undergoing pancreatoduodenectomy are a high-risk group that requires psychosocial support. This study retrospectively reviewed the prevalence of psychological symptoms in patients undergoing pancreatoduodenectomy for periampullary neoplasm and the psychosocial referral rate after implementing full screening and triage algorithm for administering a distress management protocol based on the integrated supportive care system established in 2010. MATERIALS AND METHODS: From September 2010 to December 2018, insomnia, anxiety, and depression were screened on the first day of admission (T1) and on the 10th postoperative day (T2). Patients with clinical levels of distress were referred to a mental health clinic for appropriate aftercare. RESULTS: The adherence rate to routine screening was 82.7% (364/440). Among the 364 patients, the prevalence of insomnia, anxiety, and depression increased from 22.0% (T1) to 32.6% (T2, p=0.001), 29.1% to 33.6% (p=0.256), and 18.4% to 27.6% (p=0.001), respectively. Less than 45% of those with psychological symptoms expressed their needs for psychological supportive care. Among those with psychological symptoms at T2, clinical insomnia, anxiety, and depression were detected via in-depth evaluations among 77.2%, 38.1%, and 82.5% of patients, respectively. Patients who had two or more symptoms at T2 had a longer postoperative hospital stay, as compared to those with one or no symptoms (a median of 20.5 days vs. 18.0 days, p=0.006). Psychiatric consultation rate was 72.8% among patients with clinical psychological symptoms, and 74% of the consulted patients completed psychiatric intervention before discharge. CONCLUSION: Over one-third of the patients had psychological symptoms before and after pancreatoduodenectomy. Implementing a routine psychological symptoms screening with a systematic psychiatric referral protocol enhanced surgeons' responsiveness to patients' psychological symptoms.

Isolation of Isocoumarins and Flavonoids as α-Glucosidase Inhibitors from Agrimonia pilosa L.

Agrimonia pilosa L. (AP) showed potent α-glucosidase inhibitory (AGI) activity, but it is uncertain what phytochemicals play a key factor. The phytochemical study of AP based on AGI activity led to the isolation of four isocoumarins; agrimonolide (1), agrimonolide-6-O-ß-d-glucopyranoside (2), desmethylagrimonolide (3), desmethylagrimonolide-6-O-ß-d-glucopyranoside (4), and four flavonoids; luteolin (5), quercetin (6), vitexin (7), and isovitexin (8). The four isocoumarins were isolated as α-glucosidase inhibitors for the first time. Isocoumarins, compound 1 (agrimonolide) and 3 (desmethylagrimonolide) showed strong α-glucosidase inhibitory activities with IC50 values of 24.2 and 37.4 µM, respectively. Meanwhile, isocoumarin and flavonoid glycosides showed weak AGI activity. In the kinetic analysis, isocoumarins, compounds 1 and 3 showed non-competitive inhibition, whereas flavonoid, compound 6 showed competitive inhibition.

Favorable glycemic response after pancreatoduodenectomy in both patients with pancreatic cancer and patients with non-pancreatic cancer.

Diabetes mellitus (DM) is prevalent in patients with pancreatic cancer and tends to improve after tumor resection. However, the glycemic response of non-pancreatic cancer patients after surgery has not been examined in detail. We aimed to investigate the changes in glucose metabolism in patients with pancreatic cancer or non-pancreatic cancer after pancreatoduodenectomy (PD).We prospectively enrolled 48 patients with pancreatic cancer and 56 patients with non-pancreatic cancer, who underwent PD. Glucose metabolism was assessed with fasting glucose, glycated hemoglobin (HbA1c), plasma C-peptide and insulin, quantitative insulin check index (QUICKI), and a homeostatic model assessment of insulin resistance (HOMA-IR) and ß cell (HOMA-ß) before surgery and 6 months after surgery. Patients were divided into 2 groups: "improved" and "worsened" postoperative glycemic response, according to the changes in HbA1c and anti-diabetic medication. New-onset DM was defined as diagnosis of DM ≤ 2 years before PD, and cases with DM diagnosis >2 years preceding PD were described as long-standing DM.After PD, insulin resistance (IR), as measured by insulin, HOMA-IR and QUICKI, improved significantly, although C-peptide and HOMA-ß decreased. At 6 months after PD, new-onset DM patients showed improved glycemic control in both pancreatic cancer patients (75%) and non-pancreatic cancer patients (63%). Multivariate analysis showed that long-standing DM was a significant predictor for worsening glucose control (odds ratio = 4.01, P = .017).Favorable glycemic control was frequently observed in both pancreatic cancer and non-pancreatic cancer after PD. PD seems to contribute improved glucose control through the decreased IR. New-onset DM showed better glycemic control than long-standing DM.

Profiles of Glucosinolates, Their Hydrolysis Products, and Quinone Reductase Inducing Activity from 39 Arugula (Eruca sativa Mill.) Accessions.

Glucosinolates, their hydrolysis product concentrations, and the quinone reductase (QR) inducing activity of extracts of leaf tissue were assayed from 39 arugula (Eruca sativa Mill.) accessions. Arugula accessions from Mediterranean countries (n = 16; Egypt, Greece, Italy, Libya, Spain, and Turkey) and Northern Europe (n = 2; Poland and United Kingdom) were higher in glucosinolates and their hydrolysis products, especially glucoraphanin and sulforaphane, compared to those from Asia (n = 13; China, India, and Pakistan) and Middle East Asia (n = 8; Afghanistan, Iran, and Israel). The QR inducing activity was also the highest in Mediterranean and Northern European arugula accessions, possibly due to a significant positive correlation between sulforaphane and QR inducing activity (r = 0.54). No nitrile hydrolysis products were found, suggesting very low or no epithiospecifier protein activity from these arugula accessions. Broad sense heritability (H(2)) was estimated to be 0.91-0.98 for glucoinolates, 0.55-0.83 for their hydrolysis products, and 0.90 for QR inducing activity.

Caffeic acid: potential applications in nanotechnology as a green reducing agent for sustainable synthesis of gold nanoparticles.

The sustainable synthesis of gold nanoparticles from gold ions was conducted with caffeic acid as a green reducing agent. The formation of gold nanoparticles was confirmed by spectroscopic and microscopic methods. Spherical nanoparticles with an average diameter of 29.99 ± 7.43 nm were observed in high- resolution transmission electron microscopy and atomic force microscopy images. The newly prepared gold nanoparticles exhibited catalytic activity toward the reduction of 4-nitrophenol to 4-aminophenol in the presence of sodium borohydride. This system enables the preparation of green catalysts using plant natural products as reducing agents, which fulfills the growing need for sustainability initiatives.

Identification and evaluation of flavone-glucosides isolated from barley sprouts and their inhibitory activity against bacterial neuraminidase.

Neuraminidase (NA) is one of the key enzymes responsible for bacterial infection and pathogenesis. This study aimed to gain deeper insights into the inhibitory effects of flavone-glucosides (1-9) isolated from barley sprouts (BS) on neuraminidase activity. The isolated compounds were identified as, lutonarin (1), saponarin (2), isoorientin (3), orientin (4), isovitexin (5), isoscoparin-7-O-[6-sinapoyl]-glucoside (6), isoscoparin-7-O-[6-feruloyl]-glucoside (7), isovitexin-7-O-[6-sinapoyl]-glucoside (8), and isovitexin-7-O-[6-feruloyl]-glucoside (9). Among them, compounds 1-5 exhibited neuraminidase-inhibitory activities in a dose-dependent manner, with IC50 values ranging from 20.1 to 32.7 µM, in a non-competitive inhibition mode according to kinetic studies. Moreover, the individual flavone-glucoside levels differed notably, in particular, lutonarin (1) and saponarin (2) were shown to be present in the greatest amounts, according to UPLC analysis. Consequently, our results suggest that BS may be utilized as an effective NA inhibitor in human health food, additives, and feed.

Antioxidant and enzyme inhibitory activities of Plebeian herba (Salvia plebeia R. Br.) under different cultivation conditions.

An adaptation of cultural management to the specific cultural system, as well as crop demand, can further result in the improvement of the quality of horticultural products. Therefore, this study focused on the antioxidant and enzyme inhibitory activities of Plebeian herba (Salvia plebeia R. Br.) grown in hydroponics in comparison with those of the plant grown in soil. The antioxidant activities of Plebeian herba extract were measured as 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radical scavenging abilities as well as the reducing power by decreasing nitric oxide (NO) and superoxide dismutase activity (SOD) in vitro. Interestingly, by comparison with hydroponics and traditional cultivation, Plebeian herba cultivated in nutrition-based soil improved inhibitory effect on free radicals of DPPH, ABTS, and NO and increased the contents of phenolics such as caffeic acid (1), luteolin-7-glucoside (2), homoplantaginin (3), hispidulin (4), and eupatorin. Free radical scavenging and SOD activity, as well as α-glucosidase inhibitory effect, were higher in Plebeian herba grown in nutrition-based soil than in plants grown in hydroponics and traditional condition.

Biogenic silver nanoparticles with chlorogenic acid as a bioreducing agent.

We report the synthesis of biogenic silver nanoparticles using chlorogenic acid as a bioreducing agent. Chlorogenic acid is a polyphenol compound abundant in coffee. UV-Vis spectra showed the characteristic surface plasmon resonance band at 415 nm, indicating the successful synthesis of biogenic silver nanoparticles. Spherical and irregular shaped nanoparticles were observed with an average diameter of 19.29 +/- 8.23 nm. The reaction yield from silver ion to silver nanoparticles was observed as 95.43% by using inductively coupled plasma-mass spectrometry. Fourier transform infrared spectra revealed that the -C = O groups of chlorogenic acid may coordinate or complex into silver nanoparticles. Biogenic silver nanoparticles exerted higher antibacterial activity against Gram-negative bacteria than against Gram-positive bacteria. Interestingly, a comparable antibacterial activity to a standard antibiotic was observed against two strains of Pseudomonas aeruginosa (minimum inhibitory concentration of 0.66 microg/mL). The synergistic effect of a combination of silver nanoparticles and chlorogenic acid on antibacterial activity is obvious, leading to approximately 8-fold enhancement in the case of Pseudomonas aeruginosa when compared with chlorogenic acid alone. The present report suggests that a pure compound with a plant origin is capable of being a bioreducing agent for the synthesis of biogenic silver nanoparticles with superior antibacterial activity, opening up many applications in nanomedicine and nanobiotechnology.

Changes in aurantio-obtusin and glucoaurantio-obtusin content in Cassiae Semen via treatment with a crude enzyme extract from Aspergillus usamii.

Cassiae Semen (seeds of Cassia tora) showed a remarkably different HPLC chromatogram after being treated with a crude enzyme extract from Aspergillus usamii. Increased and decreased compounds were identified as aurantio-obtusin and glucoaurantio-obtusin, respectively. The aurantio-obtusin content reached its maximum level (133.58 +/- 0.39 microg/mg extract) after being incubated for 50 min at 37 degrees C, whereas the inactivated crude enzyme-treated control remained unchanged (54.13 +/- 1.33 microg/mg). On the other hand, the glucoaurantio-obtusin content decreased by less than one-third (51.09 +/- 1.63 microg/ mg) of the untreated control (143.19 +/- 2.12 microg/mg), suggesting that an increase in aurantio-obtusin content originated from the enzymatic cleavage of its glucoside glucoaurantio-obtusin.

Platelet-activating factor (PAF) receptor binding antagonists from Alpinia officinarum.

The bioassay-guided purification of ether extracts of Alpinia officinarum led to the isolation of two new compounds 6-hydroxy-1,7-diphenyl-4-en-3-heptanone (1) and 6-(2-hydroxy-phenyl)-4-methoxy-2-pyrone (4) as well as three known compounds 1,7-diphenyl-4-en-3-heptanone (2), 1,7-diphenyl-5-methoxy-3-heptanone (3), and apigenin (5). Their structures were established on the basis of spectral methods. All three diarylheptanoids 1, 2, and 3 exhibited potent PAF receptor binding inhibitory activities with an IC(50) of 1.3, 5.0, and 1.6 microM, respectively. These studies have identified diarylheptanoids as a novel class of potent PAF antagonists.

Production of triterpene acids by cell suspension cultures of Olea europaea.

Olive (Olea europaea) contains large quantity of triterpene acids including oleanolic acid (6) as a major one. Varieties of biological activities exhibited by triterpene acids attracted our attentions, especially from pharmaceutical viewpoints. Cell culture of olive plant was induced and its triterpene constituents were studied. From the cell suspension cultures, six ursane type triterpene acids; ursolic acid (9), pomolic acid (10), rotundic acid (11), tormentic acid (12), 2alpha-hydroxyursolic acid (13) and 19alpha-hydroxyasiatic acid (14), and two oleanane type acids; oleanolic acid and maslinic acid (7), have been isolated. Quantity of ursane type triterpene acids produced by cell cultures was larger than that of oleanane type. Further, a multifunctional oxidosqualene cyclase (OSC) named OEA was cloned by homology based PCRs from the same cultured cells. Major product of OEA is alpha-amyrin (ursane skeleton), showing good accordance to higher content of ursane-type triterpene acids in the cultured cells, and strongly suggesting OEA to be a major contributor OSC for their production.

Beccaridiol, an unusual 28-nortriterpenoid from the leaves of Diplectria beccariana.

A C29-triterpene, beccaridiol (1), a dihydrochalcone natural product, 2',4'-dihydroxy-3-(4-methoxyphenyl)-propiophenone (2), as well as three known compounds, 4'-hydroxy-1',2'-dihydro-beta-ionone, 4'-O-methyldavidigenin (3), and ursolic acid, have been isolated from an EtOAc-soluble extract of the leaves of Diplectria beccariana. Beccaridiol (1) was characterized as an ursane-type 28-nortriterpene possessing an unusual aromatic E-ring by spectroscopic data interpretation. The relative configuration of this unusual isolate was established by analyzing the observed NOESY NMR correlations, and the absolute stereochemistry of 1 was then determined based on the circular dichroism (CD) spectrum of its 2,3-di-p-bromobenzoate (1b) derivative. All isolates were evaluated for their potential cancer chemopreventive properties utilizing a cell culture assay to determine quinone reductase induction.

A comparative evaluation of the anticancer properties of European and American elderberry fruits.

European elderberry (Sambucus nigra), recognized in Europe for its health-promoting properties for many generations, is known to contain a range of anthocyanins, flavonoids, and other polyphenolics that contribute to the high antioxidant capacity of its berries. American elderberry (Sambucus canadensis), on the other hand, has not been cultivated, bred, and promoted as a medicinal plant like its better-characterized European counterpart. In this study, aqueous acetone extracts of the berries from these two species were fractionated and tested in a range of assays that gauge anticarcinogenic potential. Both cultivated S. nigra and wild S. canadensis fruits demonstrated significant chemopreventive potential through strong induction of quinone reductase and inhibition of cyclooxygenase-2, which is indicative of anti-initiation and antipromotion properties, respectively. In addition, fractions of S. canadensis extract showed inhibition of ornithine decarboxylase, an enzyme marker related to the promotion stage of carcinogenesis. Analysis of active fractions using mass spectrometry and liquid chromatography-mass spectrometry revealed, in addition to flavonoids, the presence of more lipophilic compounds such as sesquiterpenes, iridoid monoterpene glycosides, and phytosterols.

Beta-secretase (BACE1) inhibitors from Sanguisorbae Radix.

In the course of screening anti-dementia agents from natural products, two beta-secretase (BACE1) inhibitors were isolated from the ethyl acetate soluble fraction of Sanguisorbae Radix by the activity-guided purification using silica gel, Sephadex LH-20, and RP-HPLC. They were identified as 1,2,3-trigalloyl-4,6-hexahydroxydiphenoyl-beta-D-glucopyranoside (Tellimagrandin II, 1) and 1,2,3,4,6-pentagalloyl-beta-D-glucopyranoside (2) and were shown to non-competitively inhibit beta-secretase (BACE1) with the IC50 values of 3.10x10(-6) M and 3.76x10(-6) M, respectively. The Ki values of 1 and 2 were 6.84x10(-6) M and 5.13x10(-6) M. They were less inhibitory to alphasecretase (TACE) and other serine proteases such as chymotrypsin, trypsin, and elastase, suggesting that they were relatively specific inhibitors of BACE1.

Xanthohumol isolated from Humulus lupulus Inhibits menadione-induced DNA damage through induction of quinone reductase.

The female parts of hops (Humulus lupulus L.) show estrogenic effects as well as cancer chemopreventive potential. We analyzed the chemopreventive mechanism of hops by studying its antioxidative activities and its effect on the detoxification of a potentially toxic quinone (menadione). The detoxification enzyme quinone reductase [(NAD(P)H:quinone oxidoreductase, QR] protects against quinone-induced toxicity and has been used as a marker in cancer chemoprevention studies. Although the hop extract was only a weak quencher of free radicals formed from 1,1-diphenyl-2-picrylhydrazyl, it demonstrated strong QR induction in Hepa 1c1c7 cells. In addition, compounds isolated from hops including xanthohumol (XH) and 8-prenylnaringenin were tested for QR induction. Among these, XH was the most effective at inducing QR with a concentration required to double the specific activity of QR (CD value) of 1.7 +/- 0.7 microM. In addition, pretreatment of Hepa1c1c7 cells with XH significantly inhibited menadione-induced DNA single-strand breaks. The QR inhibitor dicumarol reversed the protective effect of XH against menadione-induced DNA damage. Because the expression of QR and other detoxifying enzymes is known to be upregulated by binding of the transcription factor Nrf2 to the antioxidant response element (ARE), the reporter activity mediated by ARE in HepG2-ARE-C8 cells was investigated after incubation with XH for 24 h. Under these conditions, XH increased ARE reporter activity in a dose-dependent manner. One mechanism by which XH might induce QR could be through interaction with Keap1, which sequesters Nrf2 in the cytoplasm, so that it cannot activate the ARE. Using LC-MS-MS, we demonstrated that XH alkylates human Keap1 protein, most likely on a subset of the 27 cysteines of Keap1. This suggests that XH induces QR by covalently modifying the Keap1 protein. Therefore, XH and hops dietary supplements might function as chemopreventive agents, through induction of detoxification enzymes such as QR.

beta-Secretase (BACE1) inhibitors from pomegranate (Punica granatum) husk.

In the course of screening for anti-dementia agents from natural products, two beta-secretase (BACE1) inhibitors were isolated from the husk of pomegranate (Punica granatum) by activity-guided purification. They were identified as ellagic acid and punicalagin with IC50 values of 3.9 x10(-6) and 4.1x10(-7) M and Ki values of 2.4x10(-5) and 5.9x10(-7) M, respectively. The compounds were non-competitive inhibitors with a substrate in the Dixon plot. Ellagic acid and punicalagin were less inhibitory to alpha-secretase (TACE) and other serine proteases such as chymotrypsin, trypsin, and elastase, thus indicating that they were relatively specific inhibitors of BACE1.

Potentiation by alpha-tocopheryl succinate of the etoposide response in multidrug resistance protein 1-expressing glioblastoma cells.

Multidrug resistance protein 1 (MRP1) is one of the representative members of the ATP-binding cassette superfamily of transporters that is involved in resistance to chemotherapeutic agents in cancer patients. MRP1 functions as an efflux pump of drugs, primarily those conjugated to glutathione (GSH). Decreases in the intracellular concentration of GSH have been shown to enhance the response of MRP1-overexpressing cells to MRP1-substrate drugs by limiting the available drug-GSH conjugates. We report here that alpha-tocopheryl succinate (TOS), a vitamin E analogue, decreased intracellular GSH concentration and blocked MRP1 function in glioblastoma cells. Functional blockade by TOS of MRP1 was confirmed by the enhanced accumulation of etoposide (VP-16), an MRP1-substrate drug. As a result, co-treatment of TOS with VP-16 or treatment with liposomes containing both TOS and VP-16 greatly enhanced the response of MRP1-expressing glioblastoma cells to VP-16. TOS may be a promising adjuvant for enhancing the therapeutic efficacy of VP-16 in patients with MRP1-expressing glioblastomas.

Role of reactive oxygen species in the induction of apoptosis by alpha-tocopheryl succinate.

Alpha-tocopheryl succinate (TOS), a vitamin E analog, is a promising anticancer agent due to its abilities to inhibit proliferation and to induce apoptosis in a variety of human malignant cell lines, while being relatively less active toward normal cells. However, the molecular mechanisms underlying the apoptotic effects of TOS are not precisely understood. Reports that TOS can generate reactive oxygen species (ROS) prompted us to investigate the role of ROS in TOS-induced apoptosis in cancer cells. We found that the human lung cancer A549 and H460 cell lines were much more sensitive to TOS-induced apoptosis than the human glioblastoma T98G and U87MG cell lines. Our data suggested that the differential TOS sensitivity was not caused by differences in the uptake and retention of TOS between TOS-sensitive and -resistant cancer cells. The differential ability of cancer cells to generate ROS in response to TOS appears to be an important factor in determining the susceptibility of cells to TOS-induced apoptosis. Our results further suggest that TOS-induced generation of ROS is involved in caspase-independent apoptosis. Taken together, our findings suggest an important role of ROS generation in TOS-induced, caspase-independent apoptosis of cancer cells.