Indian Journal of Experimental Biology Vol. 55, February 2017, pp. 122-126 Note Effect of ethanolic extract of esophagus resulting in mucosal damage and oxidative stress. About 10-20% of western population is Quisqualis indica L. flower on experimental 1 esophagitis in albino Wistar rats reported to suffer from this disease . Investigation suggests that GERD condition may lead to damage of Sarita Singh1, Amit Rai1, Siddhartha Maity2, mucosal and sub mucosal cells, which releases 2 Srimanta Sarkar3, Sutanu Maji4 & Sudipta Saha1* various anti-inflammatory mediators . This biochemical process finally enumerates for various 1Department of Pharmaceutical Sciences; 4Department of Applied pathophysiological conditions such as irritable bowel Plant Sciences, Babasaheb Bhimrao Ambedkar University, 3 Lucknow-226 025, Uttar Pradesh, India syndrome and functional dyspepsia . 2Department of Pharmaceutical Technology, Jadavpur University, Quisqualis indica L. Syn. Combretum indicum (L.) Kolkata-700 032, West Bengal, India DeFilipps., (fam. Combretaceae), commonly known 3Department of Pharmacy, National University of Singapore, 18, Science Drive 4, Singapore, 116543 as Rangoon creeper, Chinese Honeysuckle or Madhumati4,5, native to Southeast Asia, and now Received 11 September 2015; revised 31 March 2016 broadly grown in India too as an ornamental plant6. This plant mostly used against pyrexia, Quisqualis indica L., Syn. Combretum indicum (L.) DeFilipps., staphylococcal and helminth infection is also known known as Rangoon creeper or Chinese Honeysuckle, is an abundant 7 source of phenols and flavonoids thathave crucial role in free radical to possess antidiarrhoeal , anti-inflammatory, 8 scavenging. Therefore, here we investigated whether extract of antiseptic and immunomulatory activities . Sahu et al. Q. indica flower has any role against esophagitis through scavenging suggested that the flowers of Q. indica are rich in of free radical oxygen species. In this study, we elucidated the effect flavonoids and phenols9 and they have important role of ethanolic flower extract of Q. indica on experimental esophagitis in free radical scavenging activity. As the flower in albino Wister rats. The fasted animals divided into six groups and received carboxymethyl cellulose (CMC) (0.25%, 3 mL/kg, Sham contains these polar compounds in higher amounts, control) or toxic control or pantoprazole (30 mg/kg) or flower we hypothesized that the flower might be active extract of different doses (100, 200 and 300 mg/kg) were subjected against GERD via scavenging of free radical oxygen to pylorus and fore stomach ligation. All the animals were sacrificed species generated from esophageal site. In this after 8 h and evaluated for various parameters such as total acidity, context, we explored if ethanolic extract of Q. indica free acidity, gastric pH, volume of gastric juices and esophagitis index. Esophageal tissues were subjected to estimation of various flower had any significant effect on GERD. oxidative stress parameters like malonaldehyde (MDA), glutathione Materials and Methods (GSH), catalase (CAT), superoxide dismutase (SOD) and protein carbonyl (PC). In a separate experiment, in vitro antioxidant assays Plant material and preparation of extract such as DPPH and H2O2 assays, total phenolic and flavonoid The fresh flowers of Q. indica were collected contents were also conducted. The results revealed that treatments during the month of July from Lucknow, Uttar with pantoprazole and flower extracts significantly inhibited the gastric secretion, total acidity and esophagitis index. Various Pradesh, India and authenticated by Department oxidative stress parameters also restored to normal level in the of Horticulture, Babasaheb Bhimrao Ambedkar treated groups. This action could be due to the presence of higher University (A Central University), Lucknow, India. phenolic and flavonoid contents. All these findings collectively A voucher specimen has been deposited for suggest that the flower extract of Q. indica possibly possess anti- future reference (45/SM/DAPS/BBAU/14). The plant esophagitis potential. materials (200 g) were air dried under shade, Keywords: Burma Creeper, Chinese Honeysuckle, Flavonoids, powdered and extracted with ethanol (1.0 L) using Gastro esophageal reflux disease, Oxidative stress, soxhlet apparatus by successive solvent extraction Phenols, Rangoon Creeper, Ulcer method for three consecutive days. Finally, the extracted samples were evaporated to dryness using Gastro esophageal reflux disease (GERD) is rotary vacuum evaporator (IKA, Germany). The final nothing but reflux of stomach content back to yield was 13.5%. —————— Experimental animals *Correspondence: Phone: +91 8090747008 Albino Wistar rats (90-140 g) were purchased E-mail: [email protected] from Animal House, CDRI, Lucknow and were kept SINGH et al.: EFFECT OF QUISQUALIS INDICA FLOWER EXTRACT ON ESOPHAGITIS 123 in polypropylene cages under standard conditions of SEM analysis of the tissue was performed using the temperature (25±1°C) with 12 h light/dark cycle, free method as described previously17. access to commercial pellet diet and water ad libitum. Free radical scavenging assay and total phenolic and All the rats were transferred to the laboratory one flavonoids contents week before the experiment in order to acclimatize Free radical scavenging assays like 1,1-diphenyl- them in the laboratory conditions. Animals were 2-picrylhydrazine (DPPH) and hydrogen peroxide randomized and divided into six groups with six (H2O2) assays were performed by the method as animals in each group. The experimental protocol was described previously18,19. Total phenolic and flavonoid approved by Institutional Animal Ethical Committee contents were also measured in our study20,21. (Ref. No. is UIP/IAEC/2014/FEB/15). Statistical analysis Induction of esophagitis by pylorous ligation Statistical analysis was carried out using Six (n=6) groups of rats fasted for 24 h were GraphPad Prism 5.0 (Graph Pad Software, San received carboxymethyl cellulose (CMC) as Sham Diago, CA). All results were expressed as Mean ± control or toxic control (0.25% CMC, 3ml/kg, orally) Standard deviation (SD). The data were analyzed by or plant extracts (100, 200, 300 mL/kg, orally, for one-way ANOVA followed by Bonferroni multiple 7 days) or pantoprazole (30 mg/kg, orally, in single comparison test. For biochemical estimations, dose). Next day, coeliotomy was performed using statistical significance differences were considered thiopentone sodium anesthesia (50 mg/kg, intra- with respect to toxic control (aP <0.05, bP <0.01, peritonially). The abdomen of the animal was opened cP <0.001). by midline incision about 2 cm and esophagitis was induced by ligating the fore stomach and corpus with Results and Discussion silk suture. The incised regions were sutured and the GERD is a condition when stomach reflux to the animals were kept in recovery chamber and returned esophagus, leading to various pathological symptoms to their home cages10. The animals were finally like heartburn and regurgitation accompanied by other sacrificed by cervical dislocation after 6 h and complications. When the laryngopharyngeal mucosa stomach was opened along the greater curvature and is damaged by excessive stomach contents, a different esophagus was removed by extending the dissection condition known as laryngopharyngeal reflux (LPR) line of the major axis to determine the esophagitis arises. It appears that LPR would be the extension of 22 index as described for “ulcer index” under the section GERD . Oral administration of Q. indica flower “Gastric secretion in pylorous ligated rats11. The extract significantly inhibited the esophagitis in a dose number of ulcer is noted and the severity of dependent manner. Ulcer was observed in esophageal esophagitis was recorded and calculated with the region in toxic control group. However, Q. indica following scores: 0 = no ulcers. 1=superficial ulcers flower extract decreased the total acidity, gastric juice (with erosion of <1 mm), 2=deep ulcers (erosion 1-2 mm); secretion, esophagitis index and raised the gastric pH and 3=perforation (erosion >2 mm). as compared to toxic control. The sum of scores was divided by a factor of ten Determination of pH, total and free acidity and esophagitis which was designated as the esophagitis index12. The index volume of gastric juice was measured as described Necrosis and ulceration in the esophagus was subsequently under “Gastric secretion in pylorus found in all animals, which was also prominent during ligated rats’’12. The pH measurement of gastric juice microscopic observation. Oral administration of was done using a pH meter. Total acidity, free acidity Q. indica extract (100, 200, 300 mg/kg) significantly and volume of gastric juice were also measured reduced the esophagitis in a dose dependent manner. during the experiment. Q. indica (300 mg/kg) reduced the esophagitis index Estimation of free radical generation and scanning electron (25%), gastric volume (52.98%), and total acidity microscopy (SEM) of esophageal tissues (48.18 %) as compared to normal control (Table 1). Esophageal tissue was homogenized in ice-cold Q. indica extract inhibited the esophagitis index, 0.01 (M) Tris–HCl buffer, pH 7.4 and subjected decreases the volume of gastric juices, total acidity to estimation of tissue glutathione (GSH)13, and increases the gastric pH, which suggested malonaldehyde (MDA)14, catalase (CAT)15, superoxide that ethanolic extract of Q. indica are active dismutase