Isolation of Martinella obovata via flash column chromatography

Naomi Bryner

Dr. David Singleton (advisor)Dr. Kathleen Halligan (advisor)

Plant Information Bignoniaceae family

› 120 genera, 800 species

Found in Central and South America› Neotropical liana

Versatile plant› Horticulture, food,

crafts, timber, dyes, rituals, and medicine

Arevalo, et al. 2011, Gentry and Cook 1984,Gentry 1992, Witherup, et al. 1995

Overview M. Obovata and M.

iquitosensis› 2 most known

species› 9 species total for

Martinella M. iquitosensis

known to produce martinelline and martinellic acid

NH

N

HO

O

NH

NH

HN

HN

NH

Martinellic acid

NH

N

O

O

NH

NH

HN

HN

NH

HN

HN

NH

Martinelline

Martinelline & Martinellic acid Chemically unique

› Natural pyrroloquinoline ring structures

› Synthesis is sought after

Biologically unique› Natural product

(not peptide) that potently antagonize bradykinin receptors (GPCRs)

Pyrroloquinoline quinone

Davies, et al. 2013, Lovely and Mahmud 1999, , Ma, et al. 2001Ma, et al. 2003Powell and Batey 2002, Shirai, et al. 2008, Yee, et al. 2006, Zhang, et al. 2007

Summer 2013 Bark removal and

extraction

TLC and FCC› Four fractions

IR spectra

1H and 13C NMR› 2D more helpful…

Solvents Used

Extraction: CH2Cl2/MeOH

FCC: Hex/EtOAC gradient

NB1-6-7 IR spectrum

NH

N

HO

O

NH

NH

HN

HN

NH

Martinellic acid

NH

N

O

O

NH

NH

HN

HN

NH

HN

HN

NH

Martinelline

Fall 2013 CHM (1 credit) This proved to be more of a learning

experience than productive HPLC difficulties

› Long method time for little purified product› Inconsistent results for first duplicate

Fall 2013 BIO Solubility tests

› H2O – no, EtOH (95%) – somewhat, DMSO (99.5%) – yes

Disk diffusion antibiotic sensitivity testing and MIC microtiter assays

Bacteria used› Escherichia coli, Pseudomonas aeruginosa,

Bacillus cereus, and Staphylococcus aureus

Fall 2013 BIO Disk diffusion showed that only NB1-6-1,

NB1-6-6, and NB1-6-7 merited further investigation

Clear results:› EtOH (10.5%) + 1 mg/mL NB1-6-7 - S. aureus› EtOH (5.25%) + .5 mg/mL NB1-6-7 - B. cereus› DMSO (24.9%) + 1.5 mg/mL NB1-6-7 E. coli› DMSO (24.9%) + 1.5 mg/mL NB1-6-7 S.

aureus

Loose ends Will run 2D NMRs overnight this week

2/3/14› Hope to ID NB1-6-7› Pyrroloquinoline ring in any sample?

If I had another semester› Bradykinin receptor antagonism

investigation› LC-MS analysis to determine mass (no

preexisting method)

References (1) Arevalo, C.; Ruiz, I.; Piccinelli, A.; Campone, L.; Rastrelli, L. Phenolic derivatives from the leaves of

Martinella obovata (Bignoniaceae). Natural Product Communications, 2011, 6:7, 957-960. (2) Gentry, A. H. A synopsis of Bignoniaceae ethnobotany and economic botany. Annals of the Missouri

Botanical Garden, 1992, 79, 53-64. (3) Gentry, A.H.; Cook, K. Martinella (Bignoniaceae): A widely used eye medicine of South America.

Journal of Ethnopharmacology, 1984, 11, 337-343. (4) Witherup, K.; Ransom, R.; Graham, A.; Bernard, A.; Salvatore, M.; Lumma, W.; Anderson, P.; Pitzenberger,

S.; Varga, S. Martinelline and martinellic acid, novel G-protein linked receptor antagonists from the tropical plant Martinella iquitosensis (Bignoniaceae). Journal of the American Chemical Society, 1995, 117, 6682-6685.

(5) Ma, D.; Xia, C.; Jiang, J.; Zhang, J. First Total Synthesis of Martinellic Acid, a Naturally Occurring Bradykinin Receptor Antagonist. Organic Letters, 2001, 3:14, 2189-2191.

(6) Zhang, Z.; Zhang, Q.; Yan, Z.; Liu, Q. One-Step Synthesis of the Tricyclic Core of Martinellic Acid from 2-(Cyanomethyl)-3-oxo-N-arylbutanamides. Journal of Organic Chemistry, 2007, 72, 9808-9810.

(7) Ma, D.; Xia, C.; Jiang, J.; Zhang, J.; Tang, W. Aromatic Nucleophilic Substitution or CuI-Catalyzed Coupling Route to Martinellic Acid. Journal of Organic Chemistry, 2003, 68, 442-451.

(8) Davies, S.; Fletcher, Ai.; Lee, J.; Lorkin, T.; Roberts, P.; Thomson, J. Asymmetric Synthesis of (-)-Martinellic Acid. Organic Letters, 2013, 15:8, 2050-2053.

(9) Powell, D.; Batey, R. Total Synthesis of the Alkaloids Martinelline and Martinellic Acid via a Hetero Diels-Alder Multicomponent Coupling Reaction. Organic Letters, 2002, 4:17, 2913-2916.

(10) Yee Ng, Pui.; Masse, C.; Shaw, J. Cycloaddition Reactions of Imines with 3-Thiosuccinic Anhydrides: Synthesis of the Tricyclic Core of Martinellic Acid. Organic Letters, 2006, 8:18, 3999-4002.

(11) Shirai, A.; Miyata, O.; Tohnai, N.; Miyata, M.; Procter, D.; Sucunza, D.; Naito, T. Total Synthesis of (-)-Martinellic Acid via Radical Addition-Cyclization-Elimination Reaction. Journal of Organic Chemistry, 2008, 73, 4464-4475.

(12) Lovely, C.; Mahmud, H. An approach to the pyrroloquinoline core of martinelline and martinellic acid. Tetrahedron Letters, 1999, 40, 2079-2082.