Pulicaria incisa (Lam.) DC. as a Potential Source of Antioxidant, Antibacterial, and Anti-Enzymatic Bioactive Molecules: Phytochemical Constituents, In Vitro and In Silico Pharmacological Analysis.

Plants with medicinal benefits are a crucial source of compounds for developing drugs. This study was designed to determine the chemical composition, antibacterial, antibiofilm, antioxidant, and anti-enzymatic activities of Pulicaria incisa (Lam.) DC. We also reported the molecular interaction between identified molecules and several receptors associated with antimicrobial and antibiofilm activities. A total of seventeen and thirteen compounds were identified in aqueous and methanolic extracts of P. incisa, respectively. The methanolic extract yielded a higher total content of polyphenols and flavonoids of about 84.80 ± 2.8 mg GAE/g and 28.30 ± 1.2 mg QE/g, respectively. Significant antibacterial activity was recorded for both extracts, with minimum inhibitory concentration (MIC) values ranging from 30 to 36 µg/mL, and the result was comparable to the reference antibiotic control. Antibiofilm assays revealed that both extracts were able to reduce the attachment of bacterial cells to 96-well plates, but the highest antibiofilm activity was recorded against Staphylococcus aureus. The methanolic extract also showed anti-enzymatic potency and high antioxidant activity, as demonstrated by all assays used, including DPPH, FRAP, and ABTS. These results were further validated by in silico approaches, particularly the molecular interaction of the identified compounds with the targeted receptors. These findings present P. incisa as a significant source of antibacterial, antibiofilm, antioxidant, and anti-enzymatic molecules.

Investigation of Antimicrobial, Anti-Quorum Sensing, and Cytotoxic Activities of Flavonoids Isolated from Pulicaria armena Boiss. & Kotschy ex Boiss. (Asteraceae).

This is the first report on the separation and biological assessment of all metabolites derived from Pulicaria armena (Asteraceae) which is an endemic species narrowly distributed in the eastern part of Turkey. The phytochemical analysis of P. armena resulted in the identification of one simple phenolic glucoside together with eight flavon and flavonol derivatives whose chemical structures were elucidated by NMR experiments and by the comparison of the spectral data with the relevant literature. The screening of all molecules for their antimicrobial, anti-quorum sensing, and cytotoxic activities revealed the biological potential of some of the isolated compounds. Additionally, quorum sensing inhibitory activity of quercetagetin 5,7,3' trimethyl ether was supported by molecular docking studies in the active site of LasR which is the primary regulator of this cell-to-cell communication system in bacteria. Lastly, the critical molecular properties indicating drug-likeness of the compounds isolated from P. armena were predicted. As microbial infections can be a serious problem for cancer patients with compromised immune systems, this comprehensive phytochemical research on P. armena with its anti-quorum sensing and cytotoxic compounds can provide a new approach to the treatment.

Constituents of Pulicaria inuloides and Cytotoxic Activities of Two Methoxylated Flavonols.

Plants of the genus Pulicaria are known for providing traditional medicines, spices, herbal teas, and insect deterrents. Pulicaria inuloides (Poir.). DC. is one of the less chemically studied species within the genus. Hydroalcoholic extracts from roots and aerial parts of P. inuloides were analyzed using the UHPLC-PAD-MSn technique and revealed the presence of six caffeoylquinic and eleven caffeoylhexaric conjugates together with hydroxykaempferol dimethyl ether and quercetagetin trimethyl ether. Moreover, constituents of chloroform extract from the whole P. inuloides plants were isolated and identified by spectroscopic methods. One new and four known caryophyllene derivatives, three thymol derivatives, and four polymethoxylated flavonols were found in the analyzed extract. The structure of the new compound was established by spectroscopic methods (HRESIMS, 1H NMR, 13C NMR, COSY, HSQC, HMBC, NOESY). The cytotoxicity of 6-Hydroxykaempferol 3,7-dimethyl ether and quercetagetin 3,7,3'-trimethyl ether (chrysosplenol C), which are major flavonols isolated from the plant, were tested on prostate epithelial cells (PNT2), prostate cancer cells (DU145 and PC3), human keratinocytes (HaCaT), and melanoma cells (HTB140 and A375). Both flavonols demonstrated moderate cytotoxic activity against PC3 cells (IC50 = 59.5 µM and 46.6 µM, respectively). The remaining cell lines were less affected (IC50 > 150 µM).

Effect of Pulicaria mauritanica on Glucose Metabolism and Glycogen Content in Streptozotocin-Induced Diabetic Rats.

AIMS: The study aimed to assess the antihyperglycemic activity of Pulicaria mauritanica. BACKGROUND: Pulicaria mauritanica is a medicinal and aromatic plant used for the treatment of many diseases such as inflammation, diabetes, and intestinal disorders. OBJECTIVE: The main goals of this present paper were to confirm the antihyperglycemic capacity of aqueous extract from Pulicaria mauritanica in normoglycemic and diabetic rats over a period of time (7 days of treatment). METHODS: The effect of the aqueous extract of Pulicaria mauritanica from aerial parts (AEPM) on glucose and lipid metabolism was tested using an acute test (single dose during 6 hours) and subchronic assay (repeated oral administration for seven days) at a dose of 60 mg/kg and the serum glucose levels were measured in normoglycemic and streptozotocin(STZ)-induced diabetic rats. In addition, the glycogen content in the liver, extensor digitorum longus (EDL), and soleus was evaluated. The antioxidant activity, phytochemical screening, and quantification of some secondary metabolites of this extract were also performed. RESULTS: AEPM at a dose of 60 mg/kg reduced the plasma glucose concentrations significantly in STZ-induced diabetic rats after a single oral administration (p<0.05). This lowering effect became more significant during the repeated oral administration in hyperglycemic rats (p<0.0001). Also, the findings showed that this plant exhibited a significant increase in liver and skeletal soleus muscle glycogen content in diabetic rats. AEPM revealed a remarkable antioxidant activity in addition to the presence of polyphenol compounds such as flavonoids, tannins, saponins, sterols, glucides, terpenoids, quinones, anthraquinones, and mucilage. CONCLUSION: The study shows that AEPM exhibits antihyperglycemic activity in diabetic rats, and it increases liver and muscle glycogen content.

Analgesic, antipyretic, anti-inflammatory, and hepatoprotective activities of Pulicaria crispa (Forssk. ) Oliv. (Asteraceae)

Abstract Some plants of the genus Pulicaria have been used in traditional medicines for treating back pain and inflammation. They possess various bioactivities such as antipyretic, analgesic, and hepatoprotective. This study aimed to investigate the potential analgesic, antipyretic, anti- inflammatory, and hepatoprotective activities of Pulicaria crispa (P. crispa) extract (PCE). Analgesic activity was evaluated using the hot plate and acetic acid-induced writhing tests. Antipyretic and anti-inflammatory activities were evaluated using rectal temperature and carrageenan-induced hind paw edema methods, respectively. CCl4-intoxication was used for hepatoprotective activity. Also, liver histopathology was assessed. PCE, at 500 mg/kg, exhibited significant analgesic, antipyretic, and anti-inflammatory effects. The increased serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), gamma-glutamyl transferase (GGT), and bilirubin of CCl4-exposed rats reflects their liver injury. PCE significantly decreased the elevated liver markers. The hepatoprotective effect of PCE was confirmed, as it successfully reversed the altered levels of total protein, malondialdehyde (MDA), and non-protein sulfhydryls (NP-SH) in the liver tissues of CCl4-exposed rats. Histopathological studies confirmed the hepatoprotective nature of PCE. Pretreatment of rats with PCE reduced the severity of CCl4-induced liver damage. These findings concluded that PCE possesses analgesic, antipyretic, anti-inflammatory, and hepatoprotective activities.

Downregulation of fibrosis and inflammatory signalling pathways in rats liver via Pulicaria crispa aerial parts ethanol extract.

CONTEXT: Liver is a vital organ for the detoxification of toxic substances in the body, where fibrosis is the major cause of liver damage. Pulicaria crispa processes many therapeutic applications such as antioxidant, antimicrobial, anticancer and anti-ulcerative agent. OBJECTIVE: This study aimed to modulate the fibrosis and inflammatory signalling pathways in carbon tetrachloride (CCl4)-induced liver damage in rats via treatment with Pulicaria crispa aerial parts ethanol extract (PCEE). MATERIALS AND METHODS: CCl4 was intraperitoneally injected at a dose of 0.5 mL/kg b.wt./twice a week/six consecutive weeks, PCEE was orally allocated at a dose of 250 mg/kg b.wt./day/six weeks and silymarin was orally administrated at a dose of 100 mg/kg, b.wt/day/six weeks. The plant extract evaluation was done through measuring aspartate and alanine aminotransferases (AST& ALT), alkaline phosphatase (ALP), total lipids (TP), total cholesterol (TC), triglycerides (TG), high-density lipoprotein cholesterol (HDL-C), low level glycoprotein-cholesterol (LDL-C), alpha fetoprotein (AFP), reduced glutathione (GSH), superoxide dismutase (SOD), malondialdehyde (MDA), nitric oxide (NO), tumour necrosis factor-α (TNF-α) and interleukin 6 (IL-6). The liver architectures were also estimated. RESULT: The phytochemical analysis of the extract showed the presence of sterols and/or triterpenoids. Treatments with plant extract suppressed significantly (p < 0.0001) the levels of AST, ALT, ALP, TP, TG, TC, LDH-C, MDA, NO, AFP, TNF-α and IL-6, while increased (p < 0.0001) the levels of HDL-C, GSH and SOD. The histopathological features confirmed the therapeutic role of the plant extract. CONCLUSION: PCEE succeeded to exert anti-fibrotic, anti-inflammatory and anti- oxidants effects in CCl4-induced liver fibrosis.

Essential Oil Analysis and Antimicrobial Evaluation of Three Aromatic Plant Species Growing in Saudi Arabia.

Arabian flora is a rich source of bioactive compounds. In this study, we investigated three aromatic plant species with the aim of finding valuable sources of antimicrobial agents against common pathogenic microorganisms. We focused especially on microorganisms, which cause outbreaks of infectious disease during mass gatherings and pilgrimages season in Saudi Arabia. The essential oils of three aromatic plant species were hydrodistilled from flowering aerial parts of Lavandula pubescens Decne. and Pulicaria incisa subsp. candolleana E.Gamal-Eldin, and from leaves, stems, ripe and unripe fruits of Juniperus procera Hochst. Ex Endl. They were subsequently analyzed by gas chromatography-mass spectrometry (GC-MS). The main constituents of L. pubescens were found to be carvacrol (55.7%), methyl carvacrol (13.4%), and ß-bisabolene (9.1%). P. incisa subsp. Candolleana essential oil was rich in linalool (33.0%), chrysanthenone (10.3%), eugenol (8.9%), and cis-chrysanthenol (8.0%); the major components of J. procera essential oil were α-pinene (31.3-62.5%) and δ-3-carene (7.3-30.3%). These essential oils were tested against thirteen American Type Culture Collection (ATCC) strains of Gram-positive and Gram-negative bacteria using the agar diffusion assay. The only effective essential oil was that of L. pubescens and the most sensitive strains were Acinetobacter baumannii, Salmonella typhimurium, Shigella sonnei, Enterococcus faecalis and Staphylococcus epidermidis. Carvacrol, the major constituent of L. pubescens, was tested on these strains and was compared with vancomycin, amikacin, and ciprofloxacin. The Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC) assays of L. pubescens essential oil and carvacrol revealed that Gram-negative strains were more susceptible than the Gram-positive ones.

Stabilization of water-in-oil emulsion of Pulicaria jaubertii extract by ultrasonication: Fabrication, characterization, and storage stability.

This study investigated the effect of ultrasonic treatments on the properties and stability of the water-in-oil (W/O) emulsion of Pulicaria jaubertii (PJ) extract. The study used different ultrasound powers (0, 100, 200, 400, and 600 W) at two storage degrees (4 and 25 °C) for 28 days. The findings showed that the emulsifying properties were improved to different extents after ultrasonic treatments. The treatment at 600 W showed optimum particle size, polydispersity index, emulsifying property, viscosity properties, and release of total phenolic content than the other powers. However, the ultrasonic power of 400 W gave positive effects on creaming index and antioxidant release compared to 600 W. The emulsion stored at 4 °C presented higher stability than that stored at 25 °C during the 28 days of storage. Microscopically, the increase in sonication power up to 600 W reduced particle size and decreased flocculation, thus resulted in stable emulsions, which is desirable for its applications in food systems.

Phytochemical Characterization, In Vitro Anti-Inflammatory, Anti-Diabetic, and Cytotoxic Activities of the Edible Aromatic Plant; Pulicaria jaubertii.

Pulicaria jaubertii is a medicinal herb that alleviates inflammations and fever. Chromatographic separation, phytochemical characterization, and in vitro biological activities of the plant n-hexane extract were conducted for the first time in this study. Six compounds were isolated for the first time from the n-hexane fraction of Pulicaria jaubertii aerial parts and were identified on the bases of NMR and MS analyses as pseudo-taraxaterol (1), pseudo-taraxasterol acetate (2), 3ß-acetoxytaraxaster-20-en-30-aldehyde (3), calenduladiol-3-O-palmitate (4), stigmasterol (5), and α-tocospiro B (6). Compound (6) was a rare tocopherol-related compound and was isolated for the first time from family Asteraceae, while compound (3) was isolated for the first time from genus Pulicaria. The total alcoholic extract and n-hexane fraction were tested for their anti-inflammatory, antidiabetic, and cytotoxic activities. The n-hexane fraction has dose dependent red blood cells (RBCs) membrane stabilization and inhibition of histamine release activities with IC50: 60.8 and 72.9 µg/mL, respectively. As antidiabetic activity, the alcoholic extract exerted the most inhibition on the activity of yeast α-glucosidase, with an IC50: 76.8 µg/mL. The n-hexane fraction showed cytotoxic activity against hepatocarcinoma (HepG-2), breast carcinoma (MCF-7), and prostate carcinoma (PC-3) cell lines with IC50: 51.8, 90.8 and 62.2 µg/mL, respectively. In conclusion, the anti-inflammatory effect of Pulicaria jaubertii might be attributed to the triterpenoid constituents of the n-hexane extract of the plant.

2α-Hydroxyalantolactone from Pulicaria undulata: activity against multidrug-resistant tumor cells and modes of action.

BACKGROUND: Sesquiterpene lactones having α-methylene-γ-lactone moiety are promising natural metabolites showing various biological activity. One of the major metabolites isolated from Pulicaria undulata, 2α-hydroxyalantolactone (PU-1), has not been investigated in detail yet. Multidrug resistance (MDR) represents a major obstacle for cancer chemotherapy and the capability of novel natural products to overcoming MDR is of great interest. PURPOSE: Exploring the molecular modes of action for potent natural product metabolites. METHODS: The resazurin reduction assay was employed to evaluate the cytotoxicity of PU-1 on sensitive and their corresponding drug-resistant cell lines (overexpressing P-glycoprotein, BCRP, ABCB5, ΔEGFR, or TP53 knockout). Gene expression profiling was performed by transcriptome-wide mRNA microarray in the human CCRF-CEM leukemic cells after treatment with PU-1. The top significantly up- or down-regulated genes were identified by Chipster program and analyzed using Ingenuity Pathway Analysis (IPA) software. Finally, flow cytometry and Western blotting were performed for cell cycle analyses and apoptosis detection. RESULTS: The sesquiterpene lactone, PU-1, showed potent cytotoxicity towards the drug-sensitive and -resistant cell lines. Transcriptome-wide mRNA expression profiling and pathway analysis pointed to genes involved in DNA damage response and G2/M cell cycle arrest. G2/M arrest was verified by flow cytometry and further confirmed by the upregulation of p21 and downregulation of p-CDC25C expression in Western blotting. Moreover, the suggested DNA damage checkpoint regulation was confirmed by immunofluorescence and Western blotting by upregulation of pS345 Chk1, p-H3 and γ-H2AX. Furthermore, PU-1 inhibited PI3K/AKT pathway, which is involved in signaling DNA damage and G2/M arrest. Cells ultimately induced apoptosis upon PU-1 treatment. CONCLUSIONS: PU-1 is a potent natural product inhibiting otherwise drug-resistant human tumor cell growth through DNA damage, G2/M cell cycle arrest and apoptosis.

In Vitro Studies on the Immunomodulatory Effects of Pulicaria crispa Extract on Human THP-1 Monocytes.

BACKGROUND: Pulicaria crispa (P. crispa) is a plant from the Compositae family that exhibits antioxidant, anti-inflammatory, antibacterial, and cytotoxic activities. OBJECTIVE: The current study aimed at investigating the immunomodulatory effects of P. crispa extract in lipopolysaccharide- (LPS-) stimulated human monocytic THP-1 cells. METHODS: To induce macrophage differentiation, THP-1 cell lines were treated with phorbol-12-myristate 13-acetate, followed by exposure to LPS with or without 50 or 100 µg/ml of P. crispa extract. The following tests were employed to test the immunomodulatory effects of the extract: MTT assay, ELISA, Western blotting analysis, cell migration and phagocytosis assays, and Annexin V staining method. RESULTS: Exposure to 100 µg/ml P. crispa extract significantly reduced THP-1 cell proliferation, migration, and phagocytosis (in LPS-stimulated cells, but not in unstimulated cells). Moreover, the extract alone significantly reduced the rate of THP-1 cell apoptosis, while it increased the rate of late apoptosis. Molecular investigations showed that treatment with P. crispa extract significantly upregulated the expression of ERK1, p-MAPK, P-P38, and Bcl2, while it significantly reduced the expression of ERK5, Bax, NF-κB, P-NF-κB, CCL1, CCL2, CCL5, CCL22, CXCL1, and CXCL10. CONCLUSION: Pulicaria crispa extract exhibited anti-inflammatory, antiproliferative, antimigratory, and antiphagocytic effects in LPS-stimulated THP-1 cells. Future studies should investigate these mechanisms in animal models with chronic inflammatory diseases.

Chemical composition and biological activities of the essential oil from Pulicaria undulata (L.) C. A. Mey. growing wild in Egypt.

Pulicaria undulata is used as a traditional herbal remedy in Egypt. We used gas chromatography-mass spectrometry for analysis of essential oil of this plant growing wild in Egypt and 64 compounds were identified. The oil was rich in oxygenated monoterpenes (64.0%) and aromatic derivatives (18.8%). The major components were carvacrol (46.5%), xanthoxylin (18.1%) and carvotanacetone (8.7%). The oil of the Egyptian plant showed significant differences from the oil results reported on this species derived from different accessions. Antioxidant activity was performed by FRAP, DPPH and ABTS assays, and the oil demonstrated a powerful antioxidant properties. Furthermore, cytotoxicity was assessed using MTT assay against three cell lines (A375, T98G, HCT116) and the oil showed moderate results with IC50 of 18.53, 40.64 and 22.23 µg/ml; respectively. The oil showed a good anti-acetylcholinesterase activity (IC50 = 139.2 µg/ml) using Ellman method. In conclusion, the studied oil exhibited a peculiar fingerprint and promising biological activities.

Suppression of LPS-Induced Hepato- and Cardiotoxic Effects by Pulicaria petiolaris via NF-κB Dependent Mechanism.

Recently, there is an increasing interest in searching for harmless natural products isolated from plant materials that can be used as beneficial dietary supplements and/or therapeutic drug candidates. The present study aimed to test the potential protective role of Pulicaria petiolaris (PP, Asteraceae) against hepatic and cardiotoxic effects associated with lipopolysaccharide (LPS) injection. PP was given orally for 5 days at two different doses before LPS injection. Results have shown that LPS induced remarkable hepatic and cardiac injurious effects in mice. Hepatic damage was evident through increased serum transaminases, lactate dehydrogenase (LDH), alkaline phosphatase (ALP), and activity. Estimation of high levels of serum creatine kinase-MB (CK-MB) and cardiac troponin I indicated cardiac damage. Histopathological examination of liver and heart confirmed the biochemical results. Increase in oxidative stress along with a depressed antioxidant status of liver and heart were observed in LPS-intoxicated animals. Furthermore, LPS induced activation of nuclear factor-κB (NF-κB) and subsequent elevation of inflammatory cytokines (TNF-α, IL-6). On the other hand, PP treatment successfully safeguards both organs against LPS-induced injury as indicated by the improvement of the biochemical and histopathological parameters. These results suggest that PP ameliorates LPS-induced hepatic and cardiac oxidative injurious effects via antioxidant and anti-inflammatory effects.

Assessment of anticancer activity of Pulicaria undulata on hepatocellular carcinoma HepG2 cell line.

Searching for new sources of safe nutraceuticals antitumor drugs is an important issue. Consequentially, this study designed to assess the antitumor activity of Pulicaria undulata extract in vitro in the treatment of hepatocellular carcinoma HepG2 cell line. Aerial parts of P. undulata plants were collected, used for phytochemical analysis, and assessed for anticancer activity. The antitumor activity was evaluated through studying the cell viability and apoptotic pathway. The gas chromatography-mass spectrometry phytochemical analysis revealed that P. undulata is a promising new source of several known antioxidant and antitumor compounds which could participate in drug development and exploration of alternative strategies to the harmful synthetic antitumor drugs. P. undulata stifled HepG2 cell viability in a concentration-dependent manner. Meanwhile, P. undulata tempted substantial apoptosis in HepG2 cells and enhanced the expression of miR-34a. However, the mRNA expression level of antiapoptotic B-cell lymphoma-2 was markedly decreased by P. undulata treatment. Moreover, P. undulata increased the protein expression of proapoptotic p53 and caspase 3/9 with reducing B-cell lymphoma-2 protein expression level. Thus, P. undulata induced apoptosis in the HepG2 cells by overexpression of miR-34a which regulates p53/B-cell lymphoma-2/caspases signaling pathway. These findings were well appreciated with morphological studies of cells treated with P. undulata. In conclusion, P. undulata could be a probable candidate agent for the initiation of cell apoptosis in HepG2 and thereby can serve as promising therapeutic agent for treatment of hepatocellular carcinoma which should attract further studies.

Cytotoxicity of 40 Egyptian plant extracts targeting mechanisms of drug-resistant cancer cells.

BACKGROUND: The multidrug resistance (MDR) phenotype encounters a major challenge to the success of established chemotherapy in cancer patients. We hypothesized that cytotoxic medicinal plants with novel phytochemicals can overcome MDR and kill MDR-cells with similar efficacy as drug sensitive cells. PURPOSE: We evaluated plant extracts from an unexplored ecosystem in Egypt with unusual climate and nutrient conditions for their activity against sensitive and multidrug-resistant cancer cell lines. MATERIAL AND METHODS/STUDY DESIGN: Methylene chloride: methanol (1:1) and methanol: H2O (7:3) extracts of 40 plants were prepared resulting in a sum of 76 fraction containing compounds with varying polarity. The resazurin reduction assay was employed to evaluate the cytotoxicity of these extracts on five matched pairs of drug-sensitive and their drug-resistant cell lines. Flow cytometry and Western blotting was used to determine cell cycle analyses, apoptosis, and autophagy. Reactive oxygen species (ROS) were measured spectrophotometrically. RESULTS: Extracts derived from Withania obtusifolia (WO), Jasonia candicans (JC), Centaurea lippii (CL), and Pulicaria undulata (PU) were the most active ones among 76 extracts from 40 Egyptian medicinal plants. They showed a significant reduction of cell viability on drug-sensitive CCRF-CEM leukemia cell line with IC50 values less than 7 µg/ml. Low cross-resistance degrees were observed in multidrug-resistant CEM/ADR5000 cells towards CL (1.82-fold) and JC (6.09-fold). All other drug-resistant cell lines did not reveal cross-resistance to the four extracts. Further mechanistic assessment have been studied for these four extracts. CONCLUSION: The methylene chloride: methanol (1:1) fractions of WO, JC, CL, and PU are promising cytotoxic extracts that could be used to combat MDR cancer cells through different cell death pathways.

Pulicaria gnaphalodes powder in broiler diets: consequences for performance, gut health, antioxidant enzyme activity, and fatty acid profile.

The search constantly continues to identify potential alternatives to the use of antimicrobial growth promoters (AGP) in broiler production. This trial was conducted with broiler chicks to investigate the effect of different levels of Pulicaria gnaphalodes powder (PGP) in comparison with AGP, and probiotic (PRO) on growth performance, gut microflora, intestinal morphology, antioxidant enzyme activity, and fatty acid profile of meat. Ross 308 male broiler chicks (n = 576) were randomly assigned into 6 dietary treatments with 8 replicate pens per treatment and 12 birds per pen. The dietary treatments consisted of a basal diet as control (CON, with no additive), CON + 0.1% PGP, CON + 0.2% PGP, CON + 0.3% PGP, CON + 0.1% probiotic mixture (PRO), and CON + 0.05% bacitracin methylene disalicylate (AGP). Higher body weight gain and lower feed conversion ratio were obtained in birds fed AGP and 0.3% PGP compared with those fed CON and 0.1% PGP during grower, finisher, and the entire study (P < 0.05). On day 42, birds on PRO, 0.2 and 0.3% PGP treatments had lower counts of Escherichia coli and higher lactobacillus spp. in ileum and cecal contents compared to the CON and 0.1% PGP (P < 0.05). Villus height and villus height to crypt depth ratio of the duodenum were increased (P < 0.05) in response to dietary AGP, PRO, and 0.3% PGP. The diets containing PRO and different levels of PGP increased superoxide dismutase and glutathione peroxidase activities and decreased malondialdehyde level in serum, liver, and thigh muscle (P < 0.05). Total polyunsaturated fatty acid and n-3 fatty acid of birds fed PRO and PGP diets were higher than birds in CON and AGP groups (P < 0.05). In summary, supplementation of PGP could be a potential alternative to AGP in broiler diets due to its combined positive impacts on performance, serum cholesterol, intestinal health, antioxidant activity, and fatty acid profile in meat. Such effects, however, need to be further verified under compromised health or a disease challenge condition.

Pulicaria crispa mitigates gastric ulcer induced by ethanol in rats: role of treatment and auto healing.

Context: Stomach ulcers are the common gastrointestinal disorders worldwide. Objective: This study aimed to investigate the therapeutic impact of Pulicaria crispa aerial parts ethanol extract against gastric ulcer in rats. Materials and methods: Ulcer was induced by one oral dose of ethanol (0.5 ml/100g body weight) on 24 hours empty stomach, then the plant extract (500 mg/kg b.wt.) was orally administered daily for one week. Ranitidine (100 mg/kg b.wt.); as a reference drug was evaluated. Stomach acidity and volume, as well as lesion counts were measured. Levels of malondialdehyde (MDA), glutathione (GSH) and superoxide dismutase (SOD) were estimated. Assay of different marker enzymes; succinate dehydrogenase (SDH), lactate dehydrogenase (LDH), glucose-6-phosphatase (G-6-Pase), acid phosphatase (AP) and 5'-nucleotidase (5'NT) were determined. Interlukin-10 (IL-10), intracellular adhesion molecule-1 (ICAM-1) and tumor necrosis factor alpha (TNF-α) were also determined. Stomach histopathological assessment was detected. Results: Gastric ulcer showed drastic changes in oxidative stress, cell organelles and inflammatory markers. These biomarkers served as good tools to identify the presence of gastric ulcer. Treatment with P. crispa recorded amelioration in most parameters exceeding the auto healing effect. Conclusion: Healing potency of P. crispa is possibly related to its content of glycosides, coumarins, flavonoids, tannins, sterols and triterpenes.

Pulicaria vulgaris Gaertn. essential oil: an alternative or complementary treatment for Leishmaniasis.

Leishmaniasis is a neglected parasitic protozoal disease that affects approximately 12 million people and represents a public health problem in Iran. The objectives of this study were to obtain the essential oil (EO) from Pulicaria vulgaris Gaertn. growing in Iran and to carry out in-vitro antileishmanial screening of the EO against promastigotes of Leishmania major and Leishmania infantum. The EO from the aerial parts of P. vulgaris was extracted by hydrodistillation. Serial dilutions of the EO were screened for in-vitro antileishmanial activity using 96-well microtiter plates. The P. vulgaris EO was active against the promastigote forms of L. major and L. infantum, with IC50 values of 244.70 and 233.65 µg/mL, respectively. Pulicaria vulgaris EO may serve as an alternative or complementary treatment for leishmaniasis.

Letter to the Editor.

I read with great interest the paper by Dehvari and Ghahghaei (Dehvari and Ghahghaei, 2018 [1]). Their paper aimed to prove that biosynthesized AgNPs mediated by Pulicaria undulata L. has the capability in inhibiting amyloid fibril formation and thus could be considered as a therapeutic agent in the treatment of amyloidosis disorders. According to the literature (Jangholi et al., 2018 [2]), Thioflavin T (ThT) is a commonly used probe to monitor in vitro amyloid fibril formation. Also, since ThT fluorescence originates only from the bound population of the dye molecules, the several orders of magnitude increase in the fluorescence intensity makes ThT an unusually sensitive and efficient reporter. The authors used ThT to monitor in vitro amyloid fibril formation of α-lactalbumin and fluorescence emission spectra were recorded upon titration of the indicated concentrations of nanoparticles or α-casein. They excited the assay solutions at 450nm and the emissions were measured over the range 460-600nm. Interestingly, upon binding to α-lactalbumin's amyloid fibrils, ThT displayed a (slight) relative increase of fluorescence signal at approximately ~530-540nm, when excited at 450nm with no background ThT fluorescence subtraction/reportage. Upon binding to amyloid fibrils, ThT generally exhibits a dramatic shift of the excitation maximum (from 385nm to 450nm) and the emission maximum from 445nm to the higher wavelengths, along with a strong fluorescence signal at ~482nm (Biancalana, 1804 [3]). Regarding the wavelength of ThT fluorescence maxima, the obtained results by Dehvari and Ghahghaei are inconsistent with what has yet been reported in the literature. Moreover, the respected authors, in the present work, have not tried to describe these unusual ThT results. However, I could not find evidence/fingerprint of "cotton effect" of ThT and/or absorption flattening within the results.

The effect of green synthesis silver nanoparticles (AgNPs) from Pulicaria undulata on the amyloid formation in α-lactalbumin and the chaperon action of α-casein.

The formation and deposition of protein fibrillar aggregates in the tissues is associated with several neurodegenerative diseases such as Alzheimer's and Parkinson's disease. Molecular chaperones are a family of proteins that are believed to have the ability to inhibit protein aggregation. The present study examines the effect of different concentrations of green synthesis silver nanoparticles (AgNPs) from Pulicaria undulata L. on the aggregation of α-lactalbumin (α-LA) and the chaperone action of αs-casein. The effects of the AgNPs were determined by measuring light scattering absorption, fluorescence (ThT assay, intrinsic fluorescence assay and ANS binding assay), TEM, CD spectroscopy and SDS-PAGE. The results showed that AgNPs have the ability to prevent the aggregation of α-LA in a concentration-dependent manner. In fact, by increasing the concentration of AgNPs within a specified range, the adsorption and interaction between AgNPs and protein have increased and protein conformational changes and self-association decreased, thus amyloid aggregation is prevented. Our results also showed that α-casein effectively prevented the aggregation of the α-lactalbumin which increased in the presence of the AgNPs. Standard experimental results, however, proved that nanoparticles had no effect on the structure and hence the chaperone ability of α-casein. Our findings showed that AgNPs can prevent protein aggregation and have no effect on the chaperone ability of αs-casein. In the main, results of this study show that biosynthesized AgNPs mediated by Pulicaria undulata L. has the capability in inhibiting amyloid fibril formation and thus could be consider as a therapeutic agent in the treatment of amyloidosis disorders.