title: Isolation, characterization and HPLC quantification of compounds from Aquilegia fragrans Benth: Their in vitro antibacterial activities against bovine mastitis pathogens.
abstract: BACKGROUND
  The underground parts of Aquilegia fragrans are traditionally used for the treatment of wounds and various inflammatory diseases like bovine mastitis. However, there are no reports on the phytochemical characterization and antibacterial studies of A. fragrans.
  
  
  OBJECTIVE
  To isolate compounds from the methanol extract of the underground parts of A. fragrans and determine their antibacterial activity against the pathogens of bovine mastitis. The study was undertaken in order to scientifically validate the traditional use of A. fragrans.
  
  
  METHODS
  Five compounds were isolated from the methanol extract of the underground parts of A. fragrans using silica gel column chromatography. Structural elucidation of the isolated compounds was done using spectral data analysis and comparison with literature. High performance liquid chromatography (HPLC) was used for the qualitative and quantitative determination of isolated compounds in the crude methanol extract. The methanol extract and isolated compounds were evaluated for antibacterial activities against mastitis pathogens using broth micro-dilution technique.
  
  
  RESULTS
  The five isolated compounds were identified as (1) 2, 4-dihydroxyphenylacetic acid methyl ester (2) β-sitosterol (3) Aquilegiolide (4) Glochidionolactone-A and (5) Magnoflorine. A quick and sensitive HPLC method was developed for the first time for qualitative and quantitative determination of four isolated marker compounds from A. fragrans. The crude methanol extract and compound 5 exhibited weak antibacterial activities that varied between the bacterial species (MIC=500-3000 µg/ml).
  
  
  CONCLUSIONS
  The above results show that the crude methanol extract and isolated compounds from A. fragrans exhibit weak antibacterial activities. Further phytochemical and pharmacological studies are required for proper scientific validation of the folk use of this plant species in the treatment of various inflammatory diseases like bovine mastitis.
authors.affiliation: Microbial Biotechnology Division, CSIR - Indian Institute of Integrative Medicine, Sanatnagar, Srinagar, Jammu & Kashmir 190005, India; Centre of Research for Development (CORD), University of Kashmir, 190006, India.
authors.forename: Saleem
authors.initials: S
authors.lastname: Mushtaq
authors.affiliation: Bioorganic Chemistry Division, CSIR - Indian Institute of Integrative Medicine, Sanatnagar, Srinagar, Jammu & Kashmir 190005, India.
authors.forename: Mushtaq A
authors.initials: MA
authors.lastname: Aga
authors.affiliation: Microbial Biotechnology Division, CSIR - Indian Institute of Integrative Medicine, Sanatnagar, Srinagar, Jammu & Kashmir 190005, India. Electronic address: pervaizqazi@yahoo.com.
authors.forename: Parvaiz H
authors.initials: PH
authors.lastname: Qazi
authors.affiliation: Centre of Research for Development (CORD), University of Kashmir, 190006, India.
authors.forename: Md Niamat
authors.initials: MN
authors.lastname: Ali
authors.affiliation: Microbial Biotechnology Division, CSIR - Indian Institute of Integrative Medicine, Sanatnagar, Srinagar, Jammu & Kashmir 190005, India.
authors.forename: Aabid Manzoor
authors.initials: AM
authors.lastname: Shah
authors.affiliation: Microbial Biotechnology Division, CSIR - Indian Institute of Integrative Medicine, Sanatnagar, Srinagar, Jammu & Kashmir 190005, India.
authors.forename: Sajad Ahmad
authors.initials: SA
authors.lastname: Lone
authors.affiliation: Microbial Biotechnology Division, CSIR - Indian Institute of Integrative Medicine, Sanatnagar, Srinagar, Jammu & Kashmir 190005, India.
authors.forename: Aiyatullah
authors.initials: A
authors.lastname: Shah
authors.affiliation: Microbial Biotechnology Division, CSIR - Indian Institute of Integrative Medicine, Sanatnagar, Srinagar, Jammu & Kashmir 190005, India.
authors.forename: Aehtesham
authors.initials: A
authors.lastname: Hussain
authors.affiliation: Medicinal Chemistry Division, CSIR - Indian Institute of Integrative Medicine, Sanatnagar, Srinagar, Jammu & Kashmir 190005, India.
authors.forename: Faheem
authors.initials: F
authors.lastname: Rasool
authors.affiliation: Medicinal Chemistry Division, CSIR - Indian Institute of Integrative Medicine, Sanatnagar, Srinagar, Jammu & Kashmir 190005, India.
authors.forename: Hafizullah
authors.initials: H
authors.lastname: Dar
authors.affiliation: Microbial Biotechnology Division, CSIR - Indian Institute of Integrative Medicine, Sanatnagar, Srinagar, Jammu & Kashmir 190005, India.
authors.forename: Zeeshan Hamid
authors.initials: ZH
authors.lastname: Shah
authors.affiliation: Bioorganic Chemistry Division, CSIR - Indian Institute of Integrative Medicine, Sanatnagar, Srinagar, Jammu & Kashmir 190005, India.
authors.forename: Shabir H
authors.initials: SH
authors.lastname: Lone
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000737:chemistry* / Q000494:pharmacology*; D001060:Aporphines / Q000737:chemistry; D031884:Aquilegia / Q000737:chemistry*; D001572:Benzofurans / Q000737:chemistry; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D005260:Female; D008414:Mastitis, Bovine / Q000188:drug therapy*; D000432:Methanol / Q000737:chemistry; D008826:Microbial Sensitivity Tests / Q000379:methods; D010936:Plant Extracts / Q000737:chemistry* / Q000494:pharmacology*; D012855:Sitosterols / Q000737:chemistry
chemical_list: D000900:Anti-Bacterial Agents; D001060:Aporphines; D001572:Benzofurans; D010936:Plant Extracts; D012855:Sitosterols; C438746:aquilegiolide; C025473:gamma-sitosterol; C001670:magnoflorine; D000432:Methanol
keywords: Antibacterial activity; Aquilegia fragrans; Bovine mastitis; HPLC; Magnoflorine; Magnoflorine (PubChem CID: 73337); Minimum inhibitory concentration (MIC); β-sitosterol (PubChem CID: 222284)