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Effect of Different Extraction Methods on Anthocyanin Content in Hibiscus sabdariffa L. and their Antiplatelet and Vasorelaxant Properties

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Abstract

Hibiscus sabdariffa L. is a worldwide component for tea and beverages, being a natural source of anthocyanins, which are associated with cardiovascular activities. To investigate this relationship, we explored different methods of aqueous extraction on the anthocyanin content and antioxidant activity of H. sabdariffa L. calyx extract (HSCE). Pharmacological effects via platelet aggregation, calcium mobilization, cyclic nucleotide levels, vasodilator-stimulated phosphoprotein Ser157 and Ser239, and on the vasomotor response of aortic rings isolated from mice are studied herewith. We found that the application of ultrasonic turbolization, 20 min, combined with acidified water was significantly more effective in the extraction process, providing extracts with the highest levels of anthocyanins (8.73 and 9.63 mg/100 g) and higher antioxidant activity (6.66 and 6.78 μM trolox/g of sample). HSCE significantly inhibited (100–1000 μg/mL) arachidonic acid-induced platelet aggregation, reduced calcium mobilization, and increased cAMP and cGMP levels with VASPSer157 and VASPSer239 phosphorylation. Vasorelaxation reduction was confirmed by the aortic rings and endothelium assays treated with nitric oxide synthase inhibitors, soluble guanylyl cyclase (sGC) oxidizing agent, or Ca2+-activated K+ channel inhibitor. The increasing of cGMP levels could be understood considering the sGC stimulation by HSCE compounds in the specific stimulus domain, which allows an understanding of the observed antiplatelet and vasorelaxant properties of H. sabdariffa L. calyx extract.

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Data Availability

All data analyzed and generated in the study is presented in the article, any additional data or information can be made available on request to the authors.

Abbreviations

ABTS :

2,2′-Azino-bis (3-ethylbenzothiazoline-6-sulfonic acid)

Ach :

Acetylcholine

AW :

Acidified water

AA :

Arachidonic acid

ACD-C :

Acid citrate-dextrose anticoagulant

BSA :

Bovine serum albumin

cAMP :

Cyclic adenosine monophosphate

cGMP :

Cyclic guanosine monophosphate

CRCs :

Cumulative concentration–response curves

DPPH :

2,2-Diphenyl-1-picryl-hidrazil

DW :

Distilled water

EGTA :

Ethylene glycol-bis(2-aminoethylether)-N,N,N′,N′-tetraacetic acid

EPG :

Epigallocatechin-3-gallate

Fura-2 AM :

Fura-2-acetoxymethyl ester

GBD :

Glibenclamide

GTP :

Guanosine-5′-triphosphate

HSCE :

H. sabdariffa calyx extract

IBMX :

3-isobutyl-l-methyl-xanthine

L-NAME :

Nω-Nitro-L-arginine methyl ester hydrochloride

ON :

nitric oxide

ODQ :

1H-[1,2,4]Oxadiazolo[4,3-a]quinoxalin-1-one

PE :

Phenylephrine

PDB :

Protein data bank

PGI2 :

Prostaglandin I2

PKA :

Protein kinase A

PKC :

Protein kinase C

PRP :

Platelet-rich plasma

RMSD :

Root mean square deviations

sGC :

Soluble guanylyl cyclase

SD :

Standard deviations

SDS-PAGE :

Sodium dodecyl sulfate–polyacrylamide gel electrophoresis

SEM :

Standard error of the mean

TEA :

Tetraethylammonium

WP :

Washed platelets

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Acknowledgements

The authors would like to thank the financial support of Fundect, FAPESP (D.R.S.M process number 59/300.549/2016 and C.H.L postdoc fellowship, process number 2017/26687-3), and FAPEMIG (Project 29628, APQ-01868-22 and FUNDEP Project 30201).

Funding

This work was supported by the FAPEMIG, grant number Project 29628, APQ-01868-22.

Author information

Author notes

  1. Denise Rubinho dos Santos Martins and Caroline Honaiser Lescano contributed equally to this work.

Authors and Affiliations

  1. Faculty of Biological and Environmental Sciences, Federal University of Grande Dourados, Dourados, MS, 79804-970, Brazil

    Denise Rubinho dos Santos Martins

  2. Department of Pharmacology, University of Campinas, Campinas, SP, 13083-881, Brazil

    Caroline Honaiser Lescano, Alberto Fernando Oliveira Justo & Julia Modesto Vicente

  3. Institute of Agricultural Sciences, Federal University of Minas Gerais, Montes Claros, MG, 39404-547, Brazil

    Caroline Honaiser Lescano, Sérgio Henrique Sousa Santos, Charles Martins Aguilar & Ivan Pires de Oliveira

  4. Department of Internal Medicine, University of São Paulo Medical School, São Paulo, SP, Brazil

    Alberto Fernando Oliveira Justo

  5. Faculty of Medicine, UNIFUNEC University Center, Santa Fé do Sul, SP, 15775-000, Brazil

    Alexandre Borges

  6. Faculty of Engineering, Federal University of Grande Dourados, Dourados, MS, 79804970, Brazil

    Eliana Janet San**ez-Argandoña

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  1. Denise Rubinho dos Santos Martins
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Contributions

D.R.S.M.: Investigation, Data curation, Resources, Conceptualization, Methodology, Writing – original draft. C.H.L.: Investigation, Data curation, Resources, Conceptualization, Methodology, Writing – original draft. A.F.O.J.: Conceptualization, Methodology, Investigation. J.M.V.: Conceptualization, Methodology, Investigation. S.H.S.S.: Writing – review & editing. C.M.A.: Writing – review & editing. A.B.: Conceptualization, Writing – review & editing. I.P.O.: Methodology, Supervision, Writing – review & editing. E.J.S.A.: Supervision, Writing – review & editing.

Corresponding author

Correspondence to Ivan Pires de Oliveira.

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Ethics Approval and Consent to Participate

Protocol approved by the Human Ethics Committee of the University of Campinas (UNICAMP) (Number: 99775118.5.0000.5404).

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Martins, D.R., Lescano, C.H., Justo, A.F.O. et al. Effect of Different Extraction Methods on Anthocyanin Content in Hibiscus sabdariffa L. and their Antiplatelet and Vasorelaxant Properties. Plant Foods Hum Nutr 78, 342–350 (2023). https://doi.org/10.1007/s11130-023-01067-5

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