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7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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Chemical Analysis andhemical Analysis and
Biological Evaluation of Tropicaliological Evaluation of TropicalPlants:lants: Goetzeaoetzea eleganslegans andnd
Pimentaimenta racemosaacemosaJaymie M. Rivera Gutiérrez
Dr. Claudia Ospina1, Dr. Mayra Pagán1 andProf. Augusto Carvajal2
1
Chemistry Department University of Puerto Rico at Cayey2Biology Department, University of Puerto Rico at Cayey
7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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Aimsims:{ Main:
z The development and application of bioassay for theresearch of antitumor activity on tropical plant’s
extract.
{
Specifics:z Studying cytotoxic properties of tropical species.
z Chemotaxonomic study of tropical plants in Puerto
Rico
7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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Introductionntroduction• Since ancient times, in Puerto Rico someplants have been used to treat diseases likecolds, coughs, diarrhea, respiratory infections,
skin lesions and others. Unfortunately, it isestimated that only a few group of endemicand native plants have been studied around
entire world.
• In Puerto Rico, we are at risk of losing the
ancestral knowledge of the healing propertiesof plants.Meléndez, 2006
7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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Plant-based anticancer moleculesO
O
O
O
O H
OO
O
P o d o p h y l l o t o x i n
L ig n a n is o l a t e d f r o m P o d o p h y llu m p e l ta t u m
O
O
O
O
OO
O
E t o p o s i d e
O G l u
S y n t h e t ic D r u g
N
N
O
O
OO H
C a m p t o t h e c i n
A lk a lo id e x t ra c te d fro m C a m p o th e c a a c u m in a ta
N
NO
O
OO H
H O
N
T o p o t e c a n
S y n t h e tic D r u g
Srivastava, 2005
7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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GENERAL STRATEGY:
Selection of the Organism
GENERAL STRATEGY:
Selection of the OrganismThe species that we chose for this study were:
•Pimenta Racemosa (P.R.) – antiseptic and astringentproperties; against headache, sore throat, stomach painreliever and it helps to regulate the digestion.
•Goetzea Elegans (G.E. - endemic) – Has not been reportedany healing power yet.
Alain, 2006
7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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Goetzea Elegans
Pimenta Racemosa
Cidra, Puerto Rico
Río Piedras, Puerto Rico
February 23, 2008:
A collection expeditionwas organized takingin consideration the
distribution andabundance of the
organisms in aspecific area
GENERAL STRATEGY:ENERAL STRATEGY:
Collectionollection
7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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GENERAL STRATEGY:ENERAL STRATEGY:
MethodologyethodologyP l a n t
D r y i n g a n d M a c e r a t i o nw i t h a m i x t u r e o f C H 2 C l 2 / M e O H ( 1 : 1 )
C r u d e E x t r a c t
S u s p e n d e d i n W a t e r a n dE x t r a c t e d w i t hs o l v e n t s o f d i f f e r e n t p o l a r i t y
H e x a n e C h l o r o f o r m E t h y l A c e t a t e B u t a n o l W a t e r*
S o m e t i m e s , b u t a n o l e x t r a c t i o n i s r e u i r e d
7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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GENERAL STRATEGY:ENERAL STRATEGY:
Methodologyethodology{ Rotovaporation of the samples
7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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Results and Discussionesults and Discussion
-Generaleneral-
Plant
Leaves dry
Weight (g)
±0.02g
Solvent name Solvent
(mL)
Crude extract
dry weight (g)
±0.02
Hexane 400
Chloroform 300
Ethyl Acetate 300
Buthanol 300
P.R. 279.66
Aqueous 175
17.76
Hexane 400
Chloroform 450
Ethyl Acetate 300
G.E. 270.10 25.10
Aqueous 200
7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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GENERAL STRATEGY:ENERAL STRATEGY:
Methodologyethodology• Biological evaluation:
•Bioassay
• Serial dilutions of extracts are made in wells of96-wells microplates in triplicate in 100μL seawater. A suspension of nauplii containing 10-15
brine shrimp larvae are added to each microwellplate incubated at room temperature for 24 hours.
7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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GENERAL STRATEGY:ENERAL STRATEGY:
Methodologyethodology
Bioassay
7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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Results and Discussionesults and Discussion
-Biological evaluationiological evaluation-Extract LC50 (µg/mL)
P.R. Crude 1.6x104
P.R. Hexane 68.9
P.R. CHCl3
92.0
G.E. Crude 6.2x104
G.E. Hexane 171.0
G.E. CHCl3 4.1x102
Three of the six extracts were active with a LC50 ≤200µg/mL. These extracts were obtained after partition with
solvents of different polarities. The crude extracts of thesespecies were inactive with a LC50 ≥200 µg/mL.
7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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Results and Discussionesults and Discussion
-Chemical analysishemical analysis-1H NMR spectrum of Pimenta Racemosa in CD3OD
The functional groups for the crude extract of P.R. are
aliphatic carbons under the 1-2 ppm region, aromatics in 6-7.1ppm, and double bonds at 5pmm. The high peaks under3.5-4.8ppm is the methanol-d used as solvent.
7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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Results and Discussionesults and Discussion-Chemical analysishemical analysis-
1H NMR of chloroform extract of Pimenta Racemosa
The corresponding compound for this NMR is eugenol
and the corresponding molecule is:
7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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Results and Discussionesults and Discussion
-Chemical analysishemical analysis-1H NMR spectrum of Goetzea Elegans in CDCl3
The functional groups for the crude extract of G.E.
are aliphatic protons under the 0.6-2.4 ppm region,and double bonds at 5pmm.
7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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GENERAL STRATEGY:ENERAL STRATEGY:
Methodologyethodology•Chemical analysis:
•TLC Analysis:
•A mixture of different concentrations ofsolvents was prepared as the liquid phase.We took drops of the corresponding samples
of G.E. crude, G.E. hexane extract and G.E.chloroform extract and settled in thestationary phase.
7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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GENERAL STRATEGY:ENERAL STRATEGY:
Methodologyethodology
TLC
7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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Results and Discussionesults and Discussion-Chemical analysishemical analysis-
•TLC analysis:
A B C
Where A is the
crude extract, Bis the hexaneextract and C isCHCl3 extract
for G.E.
•Better mixture of
solvents to purify thecompounds 9.0mLHexane with 1.0mLAcoEt
7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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GENERAL STRATEGY:ENERAL STRATEGY:
Methodologyethodology{ Chemical analysis:
z Separation by column:{
We made a separation by columns using the bestmixture of solvents with different polarities basedon the results of the TLC.
7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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GENERAL STRATEGY:ENERAL STRATEGY:
Methodologyethodology
Column and the six fractions.
7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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Results and Discussionesults and Discussion
-Chemical analysishemical analysis-•TLC analysis:
A B C D
Where A is thecrude extract, B
is fraction #1 , Cis the fraction #2and D is thefraction #3 for
the hexane extractfor G.E.
Mixture of solvents:9.5mL Hexane with0.5mL AcoEt
7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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Results and Discussionesults and Discussion
-Chemical analysishemical analysis-1H NMR of the third fraction of G.E. hexane extract.
The possible functional groups for the hexane extract ofG.E. are aliphatic carbons under the 1-2 ppm region,
aromatics in 7-8ppm, and double bonds at 5pmm.
7/30/2019 Chemical analysis and biological evaluation of tropical plants: goetzea elegans and pimienta racemosa
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Future Work:
{ Extracts purification and separation will be basedon the bioactivity results.
{13C NMR for the third fraction of G.E. hexane
extract.
Sam, et Al, 2007
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Acknowledgments:{ Dr. Claudia Ospina
{ Isamar Ortiz, Karla Claudio andJanibeth Hernandez
{
Biominds Program{ Chemistry and Technicians
Department
{ Interdisciplinary InvestigationInstitute
{ RISE Program
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Chemical Analysis andhemical Analysis and
Biological Evaluation of Tropicaliological Evaluation of TropicalPlants:lants: Goetzeaoetzea eleganslegans andnd
Pimentaimenta racemosaacemosaJaymie M. Rivera Gutiérrez
Dr. Claudia Ospina1, Dr. Mayra Pagán1 andProf. Augusto Carvajal2
1Chemistry Department University of Puerto Rico at Cayey2Biology Department, University of Puerto Rico at Cayey