Presentation Title

Exploration of Natural Remedies: The fractionation of Annona cherimola seeds and its effects on mammalian cancer cells

Faculty Mentor

Sylvine Deprèle PhD, Luiza Nogaj PhD

Start Date

17-11-2018 2:00 PM

End Date

17-11-2018 2:15 PM

Location

C304

Session

Oral 3

Type of Presentation

Oral Talk

Subject Area

health_nutrition_clinical_science

Abstract

The Annona cherimola is a fruit originating from South America, commonly used as a natural remedy due to its several medicinal properties such as being: anti-parasitic, antioxidant, anti-diabetic, antimicrobial, and anti-carcinogenic. This family of plants is also said to contain a classification of polyketides, known as acetogenins, that are thought to play a role in its anti-carcinogenic trait. This overall project aims to examine the toxicity of the A. cherimola on cancer cell lines. A previous study conducted focused on comparing the toxicity of A. cherimola seeds and leaves, revealing the seeds to be much more toxic towards HeLa cells at 0.02% concentrations. This study aims to examine the effect of seed extract fractions, based on increasing polarity, on the viability and cytotoxicity of HeLa cells; as well as, attempt to analyze the extracts to identify compounds responsible for their toxic effects towards cancer cells. In obtaining the seed extract, the seed was first homogenized and lyophilized, then extracted via Soxhlet extraction with ethyl acetate for four hours. The sample was then rotary evaporated to dryness, re-suspended in minimal ethyl acetate and further speed vacuumed. A partition of the overall extract was then subject to a flash column chromatography with gradient elution mobile phase comprising of hexane, ethyl acetate, and methanol. Seven fractions were obtained and speed vacuumed to dryness to use for a series of viability (MTT) and cytotoxicity assays on HeLa cells. It is believed that an increase in polarity will show a congruent increase in the toxicity of the fractions. Results show that fractions eluted using ethyl acetate show to be most toxic of the seven fractions. Furthermore, initial qualitative analysis by means of thin layer chromatography reveals the possible presence of acetogenins in crude extracts. Future studies aim to analyze these fractions through GC-MS to determine their chemical compositions, as well as isolating acetogenins and testing their effects on HeLa cells.

Key words: Natural Remedies, Annona cherimola seeds, Fractionation, Acetogenins, Cancer, HeLa cells, Viability

Summary of research results to be presented

Results show that fractions extracted in ethyl acetate are most toxic to HeLa cells, decreasing viability by approximately 40% in comparison to controls. Further, results also show that fractions extracted in a solution of ethyl acetate and 5% methanol decrease viability by at least 20%. Moreover, in analyzing the effects on the cytotoxicity of HeLa cells, the fractions do not show a significant effect at their tested concentrations. Future directions aim to test fractions at higher concentrations to analyze possible effects of cells.

Upon analysis of the thin layer chromatography through the use of a UV light and Kedde’s Reagent stain, it is believed that three possible acetogenins have been detected through the comparison of Rf values of actual spots to literature Rf values: asimicine, rolliniastatine, and squamocine. Future experiments will be conducted to extract these compounds from the silica and analyze the samples by means of GC-MS.

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Nov 17th, 2:00 PM Nov 17th, 2:15 PM

Exploration of Natural Remedies: The fractionation of Annona cherimola seeds and its effects on mammalian cancer cells

C304

The Annona cherimola is a fruit originating from South America, commonly used as a natural remedy due to its several medicinal properties such as being: anti-parasitic, antioxidant, anti-diabetic, antimicrobial, and anti-carcinogenic. This family of plants is also said to contain a classification of polyketides, known as acetogenins, that are thought to play a role in its anti-carcinogenic trait. This overall project aims to examine the toxicity of the A. cherimola on cancer cell lines. A previous study conducted focused on comparing the toxicity of A. cherimola seeds and leaves, revealing the seeds to be much more toxic towards HeLa cells at 0.02% concentrations. This study aims to examine the effect of seed extract fractions, based on increasing polarity, on the viability and cytotoxicity of HeLa cells; as well as, attempt to analyze the extracts to identify compounds responsible for their toxic effects towards cancer cells. In obtaining the seed extract, the seed was first homogenized and lyophilized, then extracted via Soxhlet extraction with ethyl acetate for four hours. The sample was then rotary evaporated to dryness, re-suspended in minimal ethyl acetate and further speed vacuumed. A partition of the overall extract was then subject to a flash column chromatography with gradient elution mobile phase comprising of hexane, ethyl acetate, and methanol. Seven fractions were obtained and speed vacuumed to dryness to use for a series of viability (MTT) and cytotoxicity assays on HeLa cells. It is believed that an increase in polarity will show a congruent increase in the toxicity of the fractions. Results show that fractions eluted using ethyl acetate show to be most toxic of the seven fractions. Furthermore, initial qualitative analysis by means of thin layer chromatography reveals the possible presence of acetogenins in crude extracts. Future studies aim to analyze these fractions through GC-MS to determine their chemical compositions, as well as isolating acetogenins and testing their effects on HeLa cells.

Key words: Natural Remedies, Annona cherimola seeds, Fractionation, Acetogenins, Cancer, HeLa cells, Viability