title: Rabbit conjunctivae edema and release of NO, TNF-α, and IL-1β from macrophages induced by fractions and esculentosides isolated from Phytolacca americana.
abstract: BACKGROUND
  The roots of Phytolacca americana L. (Phytolaccaceae) may be toxic. Despite heated controversy over the toxic compounds of P. americana, especially esculentosides, relevant studies remain scarce.
  
  
  OBJECTIVE
  The objective of this study is to screen the toxic fractions and compounds of P. americana, to determine the controlling indices, and to provide evidence for unraveling the mechanism.
  
  
  METHODS
  Petroleum ether (PE), CH2Cl2, n-BuOH, and water fractions were isolated from 70% ethanol extract of P. americana. The n-BuOH fraction was dissolved in 50% ethanol and precipitated by adding ethyl ether. The resultant supernatants and precipitates were referred to as SUPs and SEDs fractions, respectively. SUPs fraction was separated by column chromatography into four main stimulating esculentosides that were identified by HR-ESI/MS and NMR as EsA, EsB, EsC, and EsF. The irritating effects of esculentosides on rabbit conjunctivae (500 μg/eye) was observed by pathological examination and those on macrophages (5, 25, 50 and 100 μg/mL) were evaluated by detecting changes of NO, TNF-α, and IL-1β levels.
  
  
  CONCLUSIONS
  n-BuOH, SUP fractions, and EsC induced severe conjunctival edema. The four esculentosides induced dose-dependent releases of proinflammatory mediators NO, TNF-α, and IL-1β from macrophages, and releasing amounts peaked after 2 h of treatment. EsC and EsF induced macrophages to release mediators most significantly. EsC (50 μg/mL) functioned more effectively than EsF did, and similarly n-BuOH and SUPs fractions functioned more effectively than the esculentoside mixture. Thus, the four esculentosides exerted proinflammatory effects synergistically.
  
  
  CONCLUSIONS
  All extracted esculentosides, especially EsC, induced inflammatory stimulation. Phytolacca americana-induced irritation of the gastrointestinal tract may be associated with esculentosides such as EsC.
authors.affiliation: a School of Pharmacy, Nanjing University of Chinese Medicine , Nanjing , PR China .|b Jiangsu Key Laboratory of Chinese Medicine Processing , Nanjing University of Chinese Medicine , Nanjing , PR China , and.|c Engineering Center of State Ministry of Education for Standardization of Chinese Medicine Processing , Nanjing , PR China.
authors.forename: Hongli
authors.initials: H
authors.lastname: Yu
authors.affiliation: a School of Pharmacy, Nanjing University of Chinese Medicine , Nanjing , PR China .
authors.forename: Le
authors.initials: L
authors.lastname: Gong
authors.affiliation: a School of Pharmacy, Nanjing University of Chinese Medicine , Nanjing , PR China .
authors.forename: Xinzhi
authors.initials: X
authors.lastname: Wang
authors.affiliation: a School of Pharmacy, Nanjing University of Chinese Medicine , Nanjing , PR China .|b Jiangsu Key Laboratory of Chinese Medicine Processing , Nanjing University of Chinese Medicine , Nanjing , PR China , and.|c Engineering Center of State Ministry of Education for Standardization of Chinese Medicine Processing , Nanjing , PR China.
authors.forename: Hao
authors.initials: H
authors.lastname: Wu
authors.affiliation: a School of Pharmacy, Nanjing University of Chinese Medicine , Nanjing , PR China .
authors.forename: Tengfei
authors.initials: T
authors.lastname: Zhao
authors.affiliation: a School of Pharmacy, Nanjing University of Chinese Medicine , Nanjing , PR China .
authors.forename: Kuilong
authors.initials: K
authors.lastname: Wang
authors.affiliation: a School of Pharmacy, Nanjing University of Chinese Medicine , Nanjing , PR China .
authors.forename: Xiaobing
authors.initials: X
authors.lastname: Cui
authors.affiliation: a School of Pharmacy, Nanjing University of Chinese Medicine , Nanjing , PR China .
authors.forename: Lin
authors.initials: L
authors.lastname: Chen
mesh_terms: D000818:Animals; D003228:Conjunctiva / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism; D003231:Conjunctivitis / Q000139:chemically induced* / Q000276:immunology / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004487:Edema / Q000139:chemically induced* / Q000276:immunology / Q000378:metabolism; D018836:Inflammation Mediators / Q000378:metabolism*; D053583:Interleukin-1beta / Q000378:metabolism*; D008264:Macrophages / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism; D008297:Male; D008813:Mice, Inbred ICR; D009569:Nitric Oxide / Q000378:metabolism*; D029604:Phytolacca americana / Q000737:chemistry / Q000633:toxicity*; D008517:Phytotherapy; D010936:Plant Extracts / Q000737:chemistry / Q000302:isolation & purification / Q000633:toxicity*; D018517:Plant Roots; D010946:Plants, Medicinal; D011817:Rabbits; D018570:Risk Assessment; D012503:Saponins / Q000737:chemistry / Q000302:isolation & purification / Q000633:toxicity*; D012997:Solvents / Q000737:chemistry; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*
chemical_list: D018836:Inflammation Mediators; D053583:Interleukin-1beta; D010936:Plant Extracts; D012503:Saponins; D012997:Solvents; D014409:Tumor Necrosis Factor-alpha; D009569:Nitric Oxide
keywords: Inflammation; stimulation; toxin; traditional Chinese medicine