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Article

Bio-Prospecting of Crude Leaf Extracts from Thirteen Plants of Brazilian Cerrado Biome on Human Glioma Cell Lines

by
Viviane A. O. Silva
1,2,3,
Marcela N. Rosa
1,
Izabela N. F. Gomes
1,
Patrik da Silva Vital
1,
Ana Laura V. Alves
1,
Adriane F. Evangelista
1,
Giovanna B. Longato
1,4,
Adriana C. Carloni
1,
Bruno G. Oliveira
5,
Fernanda E. Pinto
5,
Wanderson Romão
5,
Allisson R. Rezende
6,
Arali A. C. Araújo
6,
Lohanna S. F. M. Oliveira
7,
Alessandra A. M. Souza
7,
Stephanie C. Oliveira
7,
Rosy Iara Maciel A. Ribeiro
7 and
Rui M. Reis
1,8,9,*
1
Molecular Oncology Research Center, Barretos Cancer Hospital, São Paulo 14784-400, Brazil
2
Department of Pathology, School of Medicine of the Federal University of Bahia, Salvador 40170-110, Brazil
3
Gonçalo Moniz Institute, Oswaldo Cruz Foundation (IGM-FIOCRUZ/BA), Salvador 40296-710, Brazil
4
Research Laboratory in Cellular and Molecular Biology of Tumors and Bioactive Compounds, San Francisco University, Bragança Paulista, São Paulo 12916-900, Brazil
5
Petroleomic and Forensic Laboratory, Chemistry Department, Federal University of Espírito Santo, Vitória 29075-910, Brazil
6
Ituiutaba Unit, Department of Agrarian and Natural Sciences (DECAN), State University of Minas Gerais (UEMG), Divinopolis 38302-192, Brazil
7
Laboratory of Experimental Pathology, Federal University of São João del Rei—CCO/UFSJ, Divinópolis 35501-296, Brazil
8
Life and Health Sciences Research Institute (ICVS), School of Health Sciences, University of Minho, 4710-057 Braga, Portugal
9
ICVS/3B’s—PT Government Associate Laboratory, 4710-057 Braga, Portugal
*
Author to whom correspondence should be addressed.
Molecules 2023, 28(3), 1394; https://doi.org/10.3390/molecules28031394
Submission received: 18 November 2022 / Revised: 14 December 2022 / Accepted: 16 December 2022 / Published: 1 February 2023
(This article belongs to the Special Issue Advances on Natural Products: Isolation, Identification and Biological Activity)
Browse Figures
Review Reports Versions Notes

Abstract

:
(1) Background: Malignant gliomas are aggressive tumors characterized by fast cellular growth and highly invasive properties. Despite all biological and clinical advances in therapy, the standard treatment remains essentially palliative. Therefore, searching for alternative therapies that minimize adverse symptoms and improve glioblastoma patients’ outcomes is imperative. Natural products represent an essential source in the discovery of such new drugs. Plants from the cerrado biome have been receiving increased attention due to the presence of secondary metabolites with significant therapeutic potential. (2) Aim: This study provides data on the cytotoxic potential of 13 leaf extracts obtained from plants of 5 families (Anacardiaceae, Annonaceae, Fabaceae, Melastomataceae e Siparunaceae) found in the Brazilian cerrado biome on a panel of 5 glioma cell lines and one normal astrocyte. (3) Methods: The effect of crude extracts on cell viability was evaluated by MTS assay. Mass spectrometry (ESI FT-ICR MS) was performed to identify the secondary metabolites classes presented in the crude extracts and partitions. (4) Results: Our results revealed the cytotoxic potential of Melastomataceae species Miconia cuspidata, Miconia albicans, and Miconia chamissois. Additionally, comparing the four partitions obtained from M. chamissois crude extract indicates that the chloroform partition had the greatest cytotoxic activity against the glioma cell lines. The partitions also showed a mean IC50 close to chemotherapy, temozolomide; nevertheless, lower toxicity against normal astrocytes. Analysis of secondary metabolites classes presented in these crude extracts and partitions indicates the presence of phenolic compounds. (5) Conclusions: These findings highlight M. chamissois chloroform partition as a promising component and may guide the search for the development of additional new anticancer therapies.

Graphical Abstract

1. Introduction

Malignant gliomas are aggressive tumors characterized by fast cellular growth and highly invasive properties [1,2]. Among gliomas, glioblastoma is the most common and aggressive subtype [2]. The standard treatment option for glioblastoma consists of surgical resection, as wide as possible or partial, followed by a combined regimen of radiotherapy and adjuvant chemotherapy, mainly temozolomide (TMZ) [3,4,5,6]. Chemotherapy drugs approved for glioblastomas are limited. In addition to TMZ, there are carmustine, lomustine, and bevacizumab. However, clinically, TMZ is the standard drug for these tumors [7]. TMZ is a DNA alkylating chemotherapy drug that easily penetrates the blood-brain barrier [8,9]. It is important to emphasize due primarily to the over-expression of O6-methylguanine methyltransferase (MGMT) and/or lack of a DNA repair pathway in GBM cells, some patients do not respond to TMZ [10]. In addition, the severe side effects during the treatment turned the standard therapy, TMZ, still essentially palliative [1,11]. Therefore, research focusing on exploring novel therapeutic strategies for treatment that minimize adverse symptoms and improve glioblastoma patients’ outcomes is urgently needed [11,12].
Natural products are an important source for discovering new drugs [13]. Examples of medicinal plant-derived clinical antitumor agents include irinotecan, paclitaxel, etoposide, topotecan, camptothecin, and vincristine [13,14]. Many plant-derived antitumor drugs have been found through large-scale screening studies [15,16]. Brazil has been designated the world’s richest biodiversity, and Cerrado is considered the second largest biome, exhibiting a vast diversity of natural plants [17,18,19]. These plants bring particular properties that allow them to adapt to the hostile environment and the presence of herbivores and pathogens, reflecting the diversity of their secondary compounds [20,21].
Several studies have pointed out the biological effects of extracts from medicinal plants from the Cerrado, including anti-inflammatory, antioxidant, antiparasitic, and antitumor properties, as well as their use for gastrointestinal and respiratory disorders and wound healing [22,23,24,25,26,27,28,29].
De Melo et al. described a total of 84 plant species reported for use in the prevention or treatment of cancer in Brazil; 69.05% of these were mentioned as being used for the treatment of cancer in general and 30.95% for specific tumors. The most frequently mentioned plants were Aloe vera, Euphorbia tirucalli, and Tabebuia impetiginosa. At least one pharmacological study, in vivo or in vitro, was found for 35.71% of the species [30]. In addition, the authors showed that plants from the families Euphorbiaceae, Fabaceae, Apocynaceae, and Asteraceae are highlighted and are of great importance in ethnopharmacology studies aimed at experimental trials [30].
According to de Mesquita et al. (2009) [24], several compounds of Brazilian plant species from the cerrado have been studied with promising results. In their study, 28 out of 412 extracts tested demonstrated a substantial antiproliferative effect, with at least 85% inhibition of cell proliferation in one or more cell lines, using extracts obtained from different parts of Anacardiaceae, Annonaceae, Apocynaceae, Clusiaceae, Flacourtiaceae, Sapindaceae, Sapotaceae, Simaroubaceae, and Zingiberaceae. Furthermore, 50 of the 412 tested compounds are used in traditional medicine, and 21 families show an antineoplastic effect on tumor cell lines in vitro [24]. Despite these therapeutic activities, there were few reports about the effect of extracts isolated from cerrado plants [31], resulting in the cerrado biome as a potential candidate for bioprospecting efforts in cancer research [24,27,28,29].
Considering the potential of plants from Brazilian cerrado, in this study, we performed a phytochemical screening and provided data on the cytotoxic potential of leaf extracts from 13 plants of 5 families (Anacardiaceae, Annonaceae, Fabaceae, Melastomataceae, and Siparunaceae) found in Brazilian cerrado biome. Among these families described, there has been a research focus on Melastomataceae due to its biological potential therapeutic [32]. Therefore, the cytotoxic potential was evaluated on a panel of five glioma cell lines (including two adult glioblastomas, two pediatric gliomas, and one established primary glioblastoma cell culture) and one non-tumoral astrocyte. Our results indicate Miconia chamissois crude extracts and its chloroform partition as perspectives to develop new anticancer therapies.

2. Results

2.1. Cytotoxicity Profile Analysis of Crude Extracts

The in vitro cytotoxicity analysis of crude extracts was evaluated through the cell viability of five commercial cell lines: two adult glioblastomas (GAMG and U251-MG), two pediatric gliomas (SF188 and RES259), one normal human astrocyte (NHA) and one primary glioblastoma culture (HCB151). Initially, a screening was performed with 13 crude extracts of the Brazilian cerrado flora, which showed that the glioma cells presented a heterogeneous profile of cytotoxic response after 72 h of treatment (Table 1). Most extracts evaluated exhibited a dose-dependent response in glioma cells (Figure 1).
From the cell viability curves obtained of the effect of crude extracts concentrations on glioma cells, means of IC50 values ± standard deviation (SD) values for each cell line were calculated (Table 1). Some samples had their IC50 undetermined once it was below the lowest concentration evaluated (<1.5 µg/mL) or above the highest concentration evaluated (>200 µg/mL). Among the 13 extracts, crude extracts numbers 17, 18, and 19 from Melastomataceae (M. cuspidata, M. albicans, and M. chamissois, respectively) presented the highest cytotoxicity and a dose-dependent effect, with IC50 values close to the NCI criteria, which recommends IC50 values <30 μg/mL for compounds to be evaluated as promising and used for future purification tests [33]. The lowest in vitro inhibitory activity was observed for the Bauhinia ungulata (16-I), Astronium fraxinifolium (2), Siparuna guianensis (8), Bauhinia variegata candida (15-I) and Xylopia aromatica (3) crude extracts.
From the IC50 values obtained, it was also possible to evaluate the SI (Selectivity Index) for both crude extracts and the chemotherapy TMZ of glioma cells in relation to the normal astrocyte (NHA) (Table 2). Only crude extract 8 (S. guianensis) presented an expressive SI of 5.4 for the GAMG cell line (Table 2). It is important to emphasize that even the chemotherapy, TMZ, did not show selectivity for the tumor cell lines.

2.2. Cytotoxicity Profile Analysis of Partitions from Miconia Chamissois Crude Extracts

Among the 13 extracts, crude extracts 17, 18, and 19 from Melastomataceae were of the highest cytotoxicity; few studies published on the biological activity and chemical composition of M. chamissois have supported its therapeutic importance. Therefore, in order to identify the cytotoxic potential from secondary compounds of crude extract 19 (M. chamissois) in glioma cells, it was partitioned into HDP (hydroalcoholic partition), HP (hexane partition), CP (chloroform partition), and AEP (ethyl acetate partition). The result revealed that most partitions showed a higher cytotoxic response to glioma cells when compared to crude extract (Table 3). Among the partitions, CP was more cytotoxic than partitions HP and AEP after 72 h of treatment. However, for the HDP partition, it was not possible to determine the IC50 values for all cell lines evaluated, suggesting that they are greater than 200 µg/mL (Table 3).
Moreover, the results obtained from SI of partitions showed that AEP was 12.1 more selective for GAMG cells than normal astrocytes (NHA) (Table 4).
Finally, in order to identify the cell viability kinetics, glioma cells were exposed to M. chamissois crude extract and its partitions HP and CP at 6, 12, 24, and 48 h. The results showed that most glioma cells had a dose-and-time-dependent cytotoxic response to crude extracts and partitions (Table 5). Some cells had their IC50 undetermined once it was more resistant (>200 µg/mL or >300 µg/mL) than the highest concentration. Depending on the cell line, the greatest cytotoxic activity was obtained at 48 h of treatment.

2.3. Characterization of the Crude Extracts and Partitions from Miconia Chamissois

Analysis of the electrospray ionization fourier transform ion cyclotron resonance mass spectrometry (ESI (-) FT-ICR MS) profile of M. chamissois compounds mainly indicated the presence of phenolic compounds at a magnification of 100 to 700 m/z regions [34,35,36,37,38,39,40,41,42,43,44,45,46,47,48,49,50,51,52,53,54,55]. Among the compounds were Palmitic acid, linolenic acid, 5,7-Dihydroxy-2-(4-hydroxy-3,5-dimethoxyphenyl)-5,6-dihydro-4H-chromen-4-one, Methyl 4-[(3,4-dimethoxybenzoyl)oxy]-3-methoxybenzoate. The m/z values of the main molecules found in the M. chamissois extracts are shown in Table 6.
In addition, the analysis of partition HP showed the presence of Ethyl gallate, Palmitic acid, and 6,7-Dimethoxy-1-(4-methoxy-phenyl)-isochroman-3-one. For CP, compounds such as Ethyl gallate, 2-(2-Ethoxy-2-oxoethyl)-2-hydroxysuccinic acid, 6,7-Dimethoxy-1-(4-methoxy-phenyl)-isochroman-3-one, Palmitic, and linoleic acid. Finally, AEP partition revealed Ethyl gallate, (2R,3R)-3,5-Dihydroxy-2-(3-hydroxy-4-methoxyphenyl)-7-methoxy-2,3-dihydro-4H-chromen-4-one,2-(2-Ethoxy-2-oxoethyl)-2-hydroxysuccinic acid, Kaempferol-7-neohesperidoside, 2,6-Bis-O-(3,4,5-trihydroxybenzoyl)-D-glucopyranose, 1,3,6-Trigalloyl glucose, 1,3,6-Trigalloyl glucose, quercitrin and Vitexin (Supplementary Materials (Figure S1)).

3. Discussion

The Brazilian cerrado biome is a major source of plants, which has been explored for cancer treatment in traditional medicine [24,27,29,56]. Based on this, we performed a phytochemical screening. We provided data on the cytotoxic potential of leaf extracts from 13 plants of five families (Anacardiaceae, Annonaceae, Fabaceae, Melastomataceae, and Siparunaceae) on a panel of five glioma cell lines, indicating perspectives to developing new anticancer therapies.
The present study observed the poorest in vitro inhibitory activity for the B. a ungulata, A. fraxinifolium, S. guianensis, B. variegata candida, and X. aromatica crude leaf extracts toward the glioma cell lines. Therefore, although we have not observed cytotoxic effects for these crude extracts in gliomas, many may have action on other tumor types [57,58,59,60,61,62]. Recent studies published by our group revealed the importance of the diversity of secondary compounds in different crude extracts and their selective antitumor potential. Our group reported the antineoplastic effect of the Annonaceae family on cervical cancer [57]. The crude extract 7 and hexane partition from Annona crassiflora Mart. induced several biological responses, such as DNA damage and apoptosis, and also promoted a decrease in in vivo tumor perimeter in cervical cancer cell lines [63]. In addition, the hexane fraction of X. aromatica leaves constituted by phenolic acids, flavonoids, and alkaloids revealed antitumor potential on Ehrlich ascites carcinoma cell lines in vitro and in vivo [59]. Generally, the major components identified in Annonaceae are, typically, acetogenins [64], which exhibited in vitro anticancer activities [65,66], which could explain the anticancer activity observed for A. crassiflora, and other Annonaceae like X. aromatica. We also reported the cytotoxic potential of Tapirira guianensis Aubl extracts on a panel of head and neck squamous cell carcinoma cell lines [60]. In addition, we have previously demonstrated potential antimetastatic activity for B. ungulata extract fractions [58] and have described the mechanism of action of B. variegata candida fraction on cell viability and inhibitory activity against metalloproteinases in human cervical carcinoma (HeLa) and human peripheral blood mononuclear cells [61]. The present study emphasizes the importance of screening studies and provides a basis for further investigation on the antineoplastic activity of cerrado crude extracts antineoplastic activity on anticancer therapy.
Importantly, M. cuspidata, M. albicans, and M. chamissois revealed the greatest cytotoxic activity potential, and it is in agreement with the findings of the literature since their crude extracts belong to the Melastomataceae family, which is composed of important secondary compounds whose use in folk medicine has been focused on analgesics, anti-inflammatories, and even as antineoplastics [24,31,67]. These secondary compounds include simple phenolics, terpenoids, quinones, lignans, and their glycosides, as well as numerous tannins or polyphenols, some flavonoids, and anthocyanins [68]. The specie M. albicans from which crude extract 18 was obtained stands out for its anti-inflammatory, antimicrobial, and antiparasitic actions, as well as cytotoxic effects in some malignant neoplasms, as already evidenced [24,31]. Analgesic effects of M. albicans (Sw.) Triana of crude leaf extracts and their partitions (hexane, methylene chloride, and hydroalcoholic) present in leaves have also been described [67]. Despite the literature findings on the cytotoxic potential of some species of Miconia, to the best of our knowledge, no data related to M. cuspidata (extract 17) and M. chamissois (extract 19) has been previously published.
In this context, we selected crude extract number 19 from M. chamissois to continue the biological characterization for presenting the highest cytotoxic potential and for being one of the least cytotoxic to the normal NHA cells in this study. A better efficacy in the study of natural compounds and/or the effect of antineoplastic agents has been obtained from the partitioning and consequent fractionation of the crude extract to obtain pure components responsible for their bioactivity [69]. The cellular cytotoxicity studies of crude extract 19 and partitions (HDP (hydroalcoholic), HP (hexanic), CP (chloroform), and AEP (ethyl acetate) revealed that CP presented greater cytotoxic potential when compared to the crude extracts and the other partitions in glioma cells. A better characterization of the exposure time required for cytotoxic activity was also performed in the glioma cells through cell viability kinetics at 6, 12, 24, and 48 h. Crude extract 19, and its partitions, exhibited a dose and time-dependent cytotoxic response profile at 24 h of exposure, indicating acute cytotoxicity for these treatments. Studies with natural extracts from other plants on human glioma and colon tumor cells have also shown cytotoxic and antiproliferative effects at 24 h of treatment [70,71,72,73]. A comparison of IC50 values obtained through cell viability kinetics revealed that the CP partition was more cytotoxic than the crude extract and the HP and AEP partitions in most of the cell lines evaluated.
Understanding and exploiting the similarities and differences between adult and pediatric gliomas is an important strategy for developing new targeted therapies [2,3,4]. Here, we highlight the difference in cytotoxic effect between crude extracts in glioma cells, a heterogeneous response profile among adult and pediatric gliomas, and primary culture to crude extract and partitions exposition. It is known that pediatric GBM has a high molecular heterogeneity compared to adult GBM and within its group. All four high-grade lines have highly complex genomic profiles and harbored amplifications and deletions at several known cancer genes dysregulated in pediatric glioma. Thus, these factors may influence the aggressiveness of these tumors and the response to therapies [63,74,75,76]. Our results revealed GAMG more sensitive cells to treatments than other lines. According to the cosmic database, more than 280 mutations have already been found on the GAMG cell, revealing that different pathways may contribute to its sensitivity. However, our study constitutes an initial bioprospection cytotoxicity screening, and the tests carried out do not allow us to conclude the cell lines’ molecular response. This fact indicates a limitation in our study, indicating the need for molecular characterization and a proof concept that allows us to state the exact mechanisms underlying GAMG cells, making them more sensitive. Finally, we emphasize that 2D models are a starting point for studying new drugs and do not recapitulate the microenvironment and GBM’s complex landscape [7].
FT-ICR MS analysis revealed the phenolic compounds as the main molecules found in crude extract 19 and its partitions (HP, CP, and AEP), which were confirmed by similarity in a database [34,35,36,37,38,39,41,42,43,44,45,46,47,48,49,50,51,52,53,54,55]. Phenolic compounds are the biggest group of phytochemicals, and their antitumor potential and biological properties have been evaluated during the past years [77]. Phenolic compounds are widespread in cerrado plants, especially tannins, which are directly responsible for the therapeutic activity of plants in the cerrado biome [24,27,29,56]. Some of the compounds identified in the spectrum of partitions have been described concerning cytotoxicity activity and potential anticancer effects. Among them, oleic acid, the main ingredient of Brucea javanica oil), and widely known to have anticancer effects in many tumors [78]. Palmitic acid is well known for its antitumor effect in prostate cancer [79]. Cremastranone (5,7-dihydroxy-3-(3-hydroxy-4-methoxybenzyl)-6-methoxychroman-4-one), a homoisoflavanone isolated from Cremastra appendiculata, and it was reported by its anti-angiogenic and anti-proliferative activity in human umbilical vein endothelial cells and recently identified its property in suppression of growth of colorectal cancer cells through cell cycle arrest and induction of apoptosis [80]. Another compound that is highlighted is the Kaempferol-3-O-rhamnoside due to its properties in cancer and cell death [81]. Finally, although not mentioned here, many others are being actively characterized for their structure and antitumor potential.
We emphasize that most of a crude extract’s observed efficacy could result from a combined effect of more than one component. Therefore, the partition/fraction results may not always give a real efficacy of in vivo observation. Here, CP exhibited IC50 values significantly lower than the crude extract, suggesting a potential tumorigenic activity from CP and that a great part of the metabolites present in the crude extract responsible for the antitumor potential might be present in the chloroform partition.
Finally, recently our group harnessed the knowledge of this bioprospect study to assess the antitumor potential and biological characterization of CP on glioma cell lines [82]. We reported the effects of CP and its sub-fraction, McC1. The McC1 fraction significantly reduced glioma cell migration, invasion, and clonogenicity in vitro and reduced tumor growth and angiogenesis in vivo. Moreover, this component revealed synergistic effects with TMZ, indicating future perspectives on drug development [82].

4. Conclusions

Our study represents an important screening that systematically evaluated the anticancer activity and the selectivity index of 13 different phytochemical extracts from traditional Brazilian cerrado plant species against glioma cells. Furthermore, studies carried out with one of the most effective extracts (number 19 obtained from M. chamissois species) demonstrated that the chloroform partitions derived from this extract presented greater cytotoxicity in glioma cell lines, showing a heterogeneous and dose-time dependent response profile. Further studies are needed to confirm the potential and mechanisms of action of these extracts and partitions, yet, the results presented provided valuable information for the possible development of novel anticancer drugs based on these medicinal plants in the future.

5. Materials and Methods

5.1. Cell Culture

Five established central nervous system cell lines comprising two adults (GAMG and U251-MG) and two pediatric glioma cell lines (SF188 and RES259), and one normal human astrocyte (NHA) were obtained and cultivated, as described in Table 7. In addition, one primary cell culture (HCB151) derived from surgical glioblastoma biopsies at Barretos Cancer Hospital (São Paulo, Brazil) and previously established by our group was grown under the same conditions detailed in Table 7. The identification and confirmation were conducted on blood derived from the same patient. The Ethics Committee of Barretos Cancer Hospital approved this study [83]. Moreover, all glioma cells were analyzed for mycoplasma presence and authenticated by the Diagnostics Laboratory at Barretos Cancer Hospital through short tandem repeat (STR) DNA typing, as previously described [84].

5.2. Plant Extracts and Partitions

Leaves from 13 plant species of the Brazilian cerrado biome were collected in June 2015 (18°54′5066″ S and 48°13′5728″ W). The samples were identified, and voucher specimens of the plants were kept in the herbarium of the Federal University of Uberlândia (44998HUFU, 56558 HUFU, 59592HUFU), in the herbarium of the Federal University of Minas Gerais (143407BHCB, 143403BHCB, 43397BHCB, 143400BHCB, 143404BHCB, 161589BHCB, 161590BHCB, 161588BHCB) and in the herbarium of the Federal University of Grande Dourados (CG/MS) (under number 11486CG/MS) as presented in Table 8. The plant assays, including the collection of plant material, were conducted in accordance with local legislation. Detailed information about vernacular, botanical name, family, and registration code is identified in Table 8. The leaves were ground into powder, subjected to extraction, and then filtered. The extracts were freeze-dried to obtain the crude extracts. The partitioning from M. chamissois preparation was carried out using 4 different solvents with increased polarity: HDP-Hydroalcoholic, HP-Hexane, CP-Chloroform, and AEP-Ethyl acetate. The partitions were also submitted to freeze-drying. The crude extracts were diluted in dimethyl sulfoxide (DMSO) to obtain a final solution of 50 mg/mL and stored at −20 °C. The partitions were also dissolved in DMSO at a concentration of 25 mg/mL and stored at −20 °C. The intermediate dilutions of the crude extracts and partitions were prepared to obtain a concentration of 1% DMSO.

5.3. Cytotoxic Screening

5.3.1. Cell Viability

The biological activity of crude extracts, partitions, and temozolomide (TMZ) (Sigma-T2577), were evaluated by Cell Titer 96 Aqueous cell proliferation assay (MTS assay, PROMEGA, Madison, WI, USA) as previously described [58,82]. Cells (5 × 103) were plated into 96-well plates, exposed to the vehicle (1% DMSO) or increasing concentrations of the samples (crude extracts 1, 2, 3, and 7; 12 μg/mL to 300 μg/mL, crude extracts 8, 17, 18 and 19; 1.5 μg/mL to 300 μg/mL, crude extracts 10, 14I, 15I, 16I and 21I; 3 μg/mL to 300 μg/mL, partitions: 2.5 to 200 μg/mL, and TMZ, 1.21 to 48.54 µg/mL) diluted in DMEM medium (0.5% fetal bovine serum (FBS)) and incubated for 72 h. The samples’ absorbance was measured at 490 nm in an automatic microplate reader Varioskan (Thermo). The data were expressed as mean viable cells relative to DMSO alone (100% viability) ± SD. The experiments were performed in biological and experimental triplicates, and cell viability analyzes were calculated using the GraphPad PRISM version 5 program.

5.3.2. Kinetics Assay

For the kinetics assay, glioma cells (5 × 103) cell were plated into 96-well plates and exposed to the vehicle (1% DMSO) or increasing concentrations of extract 19 partitions (concentrations from 1.5 µg/mL to 100 µg/mL), diluted in DMEM medium (0.5% fetal bovine serum (FBS)) and incubated for different time points (6, 12, 24, 48 h). The sample’s absorbance was measured at 490 nm in an automatic microplate reader Varioskan (Thermo). The data were expressed as mean viable cells relative to DMSO alone (100% viability) ± SD. The experiments were performed in biological and experimental triplicates, and cell viability analyzes were calculated using the GraphPad PRISM version 5 program.

5.3.3. Inhibitory Concentration of 50% of Cells (IC50)

IC50 determination (inhibitory concentration of 50% of cells) was performed by non-linear regression analysis using the GraphPad PRISM software version 5 from the viability results obtained in item 5.3.1. The NCI (American National Cancer Institute) determines that promising crude extracts for subsequent purification studies and cytotoxicity assays have IC50 values < 30 μg/mL, and for the cytotoxicity studies of this work, this same previously established criterion was adopted [33].

5.4. Selectivity Index

Crude extracts and partitions, as well as TMZ, were also evaluated by the selectivity index (SI), which allows identifying the selectivity of the tested compounds and their potential use in clinical studies. In this study, SI was obtained from the following formula: IC50 of non-tumor cell line (NHA)/IC50 of tumor cell lines. For this analysis, an SI value greater than or equal to 2.0 was adopted as significant, as previously described by the NCI (National Cancer Institute) [33].

5.5. Analysis of Secondary Compounds Present in the Crude Extract and Partitions in Miconia Chamissois by FT-ICR MS

The crude extract and partitions were analyzed using the negative ion-mode Electrospray Ionization Fourier Transform Ion Cyclotron Resonance Mass Spectrometer (ESI (−) FT-ICR MS, model 9.4 T Solarix, Bruker Daltonics Bremen). All mass spectra were externally calibrated using NaTFA (m/z from 200 to 2000). The source parameters of the ESI (−) source were: nebulizer gas pressure of 0.5–1.0 bar, capillary voltage of 3–3.5 Kv, and capillary transfer temperature of 250 °C. The resolution power used was m/Δm50% ≅ 200,000 (where Δm50% is the maximum peak width at peak height m/z ≅ 400) and mass accuracy <8 ppm. The degree of unsaturation for each molecule can be deduced directly from its DBE value according to the equation DBE = c − h/2 + n/2 + 1, where c, h, and n are the numbers of carbon atoms, hydrogens, and nitrogen in the molecular formula, respectively. The FT-ICR mass spectrum was acquired and processed using Compass Data Analysis software. The elemental compositions of the present compounds were determined by measuring the m/z ratio values. The (ESI (−) FT-ICR MS and NMR spectrum are presented in the supplementary material (Figure S1).

5.6. Statistical Analysis

The data were expressed as the mean ± standard deviation (SD) of at least three independent experiments. The Student’s t-test was applied to compare two different conditions, whereas a two-way analysis of variance (ANOVA) was used for assessing differences between more groups, and a p value < 0.05 was adopted as a statistically significant difference. All the analyses were performed using the aforementioned GraphPad PRISM version 7 (GraphPad Software, La Jolla, CA, USA).

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/molecules28031394/s1, Figure S1: Spectrum images of crude extract and partitions from M. chamissois are available as supporting information.

Author Contributions

Conceptualization, V.A.O.S. and R.M.R.; Formal analysis, M.N.R., I.N.F.G., P.d.S.V., A.L.V.A., A.F.E. and G.B.L.; Funding acquisition, R.I.M.A.R. and R.M.R.; Investigation, V.A.O.S. and M.N.R.; Methodology, V.A.O.S., M.N.R., A.F.E., A.C.C., F.E.P., W.R. and L.S.F.M.O.; Project administration, R.M.R.; Resources, B.G.O., F.E.P., W.R., A.R.R., A.A.C.A., A.A.M.S. and S.C.O.; Supervision, R.M.R.; Visualization, R.I.M.A.R.; Writing—original draft, V.A.O.S.; Writing—review & editing, V.A.O.S., R.I.M.A.R. and R.M.R. All authors have read and agreed to the published version of the manuscript.

Funding

This study was funded by grants from the FINEP (MCTI/FINEP/MS/SCTIE/DECIT-01/2013-FPXII-BIOPLAT), CAPES, FAPEMIG, UFSJ, and Barretos Cancer Hospital, all from Brazil.

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

Not applicable.

Acknowledgments

The authors gratefully acknowledge Tatiane Honda Morais for her assistance in data acquisition and technics contributions.

Conflicts of Interest

The authors declare no conflict of interest.

Sample Availability

Samples of the compounds are available from the authors.

References

  1. Horbinski, C.; Berger, T.; Packer, R.J.; Wen, P.Y. Clinical implications of the 2021 edition of the WHO classification of central nervous system tumours. Nat. Rev. Neurol. 2022, 18, 515–529. [Google Scholar] [CrossRef] [PubMed]
  2. Lapointe, S.; Perry, A.; Butowski, N.A. Primary brain tumours in adults. Lancet 2018, 392, 432–446. [Google Scholar] [CrossRef] [PubMed]
  3. Siegel, R.L.; Miller, K.D.; Fuchs, H.E.; Jemal, A. Cancer statistics, 2022. CA Cancer J. Clin. 2022, 72, 7–33. [Google Scholar] [CrossRef] [PubMed]
  4. Cohen, A.R. Brain Tumors in Children. N. Engl. J. Med. 2022, 386, 1922–1931. [Google Scholar] [CrossRef]
  5. Little, S.E.; Popov, S.; Jury, A.; Bax, D.A.; Doey, L.; Al-Sarraj, S.; Jurgensmeier, J.M.; Jones, C. Receptor tyrosine kinase genes amplified in glioblastoma exhibit a mutual exclusivity in variable proportions reflective of individual tumor heterogeneity. Cancer Res. 2012, 72, 1614–1620. [Google Scholar] [CrossRef]
  6. Germano, I.; Swiss, V.; Casaccia, P. Primary brain tumors, neural stem cell, and brain tumor cancer cells: Where is the link? Neuropharmacology 2010, 58, 903–910. [Google Scholar] [CrossRef]
  7. Liu, P.; Griffiths, S.; Veljanoski, D.; Vaughn-Beaucaire, P.; Speirs, V.; Bruning-Richardson, A. Preclinical models of glioblastoma: Limitations of current models and the promise of new developments. Expert Rev. Mol. Med. 2021, 23, e20. [Google Scholar] [CrossRef]
  8. Mittal, S.; Pradhan, S.; Srivastava, T. Recent advances in targeted therapy for glioblastoma. Expert Rev. Neurother. 2015, 15, 935–946. [Google Scholar] [CrossRef]
  9. Alrwas, A.; Papadopoulos, N.E.; Cain, S.; Patel, S.P.; Kim, K.B.; Deburr, T.L.; Bassett, R., Jr.; Hwu, W.J.; Bedikian, A.Y.; Davies, M.A.; et al. Phase I trial of biochemotherapy with cisplatin, temozolomide, and dose escalation of nab-paclitaxel combined with interleukin-2 and interferon-alpha in patients with metastatic melanoma. Melanoma Res. 2014, 24, 342–348. [Google Scholar] [CrossRef]
  10. Hermisson, M.; Klumpp, A.; Wick, W.; Wischhusen, J.; Nagel, G.; Roos, W.; Kaina, B.; Weller, M. O6-methylguanine DNA methyltransferase and p53 status predict temozolomide sensitivity in human malignant glioma cells. J. Neurochem. 2006, 96, 766–776. [Google Scholar] [CrossRef]
  11. Weller, M.; van den Bent, M.; Tonn, J.C.; Stupp, R.; Preusser, M.; Cohen-Jonathan-Moyal, E.; Henriksson, R.; le Rhun, E.; Balana, C.; Chinot, O.; et al. European Association for Neuro-Oncology (EANO) guideline on the diagnosis and 719 treatment of adult astrocytic and oligodendroglial gliomas. Lancet Oncol. 2017, 18, e315–e720. [Google Scholar] [CrossRef]
  12. Alves, A.L.V.; Gomes, I.N.F.; Carloni, A.C.; Rosa, M.N.; da Silva, L.S.; Evangelista, A.F.; Reis, R.M.; Silva, V.A.O. Role of glioblastoma stem cells in cancer therapeutic resistance: A perspective on antineoplastic agents from natural sources and chemical derivatives. Stem Cell Res. Ther. 2021, 12, 206. [Google Scholar] [CrossRef]
  13. Cragg, G.M.; Pezzuto, J.M. Natural Products as a Vital Source for the Discovery of Cancer Chemotherapeutic and Chemopreventive Agents. Med. Princ. Pract. 2016, 25 (Suppl. 2), 41–59. [Google Scholar] [CrossRef]
  14. Stupp, R.; Mason, W.P.; van den Bent, M.J.; Weller, M.; Fisher, B.; Taphoorn, M.J.; Belanger, K.; Brandes, A.A.; Marosi, C.; Bogdahn, U.; et al. Radiotherapy plus concomitant and adjuvant temozolomide for glioblastoma. N. Engl. J. Med. 2005, 352, 987–996. [Google Scholar] [CrossRef]
  15. Pezzuto, J.M. Plant-derived anticancer agents. Biochem. Pharm. 1997, 53, 121–133. [Google Scholar] [CrossRef]
  16. Cortelo, P.C.; Demarque, D.P.; Dusi, R.G.; Albernaz, L.C.; Braz-Filho, R.; Goncharova, E.I.; Bokesch, H.R.; Gustafson, K.R.; Beutler, J.A.; Espindola, L.S. A Molecular Networking Strategy: High-Throughput Screening and Chemical Analysis of Brazilian Cerrado Plant Extracts against Cancer Cells. Cells 2021, 10, 691. [Google Scholar] [CrossRef]
  17. Valli, M.; Russo, H.M.; Bolzani, V.S. The potential contribution of the natural products from Brazilian biodiversity to bioeconomy. Acad. Bras. Cienc. 2018, 90 (Suppl. 1), 763–778. [Google Scholar] [CrossRef]
  18. Sano, E.E.; Rodrigues, A.A.; Martins, E.S.; Bettiol, G.M.; Bustamante, M.M.C.; Bezerra, A.S.; Couto, A.F., Jr.; Vasconcelos, V.; Schuler, J.; Bolfe, E.L. Cerrado ecoregions: A spatial framework to assess and prioritize Brazilian savanna environmental diversity for conservation. J. Environ. Manag. 2019, 232, 818–828. [Google Scholar] [CrossRef]
  19. da Silva Lima, L.; Olivares, F.L.; Rodrigues de Oliveira, R.; Vega, M.R.G.; Aguiar, N.O.; Canellas, L.P. Root exudate profiling of maize seedlings inoculated with Herbaspirillum seropedicaeand humic acids. Chem. Biol. Technol. Agric. 2014, 1, 1–18. [Google Scholar] [CrossRef]
  20. Ratter, J.A.; Ribeiro, J.F.; Bridgewater, S. The Brazilian cerrado vegetation and threats to its biodiversity. Ann. Bot. 1997, 80, 223–230. [Google Scholar] [CrossRef] [Green Version]
  21. Santos, L.D.; Viel, A.M.; Tarosso, L.F.; Momesso, L.d.S.; Palmieri, D.A.; Spera, K.D. Medicinal plants of the Brazilian cerrado: Knowing to preserve. Biosci. J. 2020, 556–567. [Google Scholar] [CrossRef]
  22. Malara, F.A.; Matos, D.C.; Ribeiro, L.C.; Falcoski, T.O.; Andrade, T.J.; Santos, V.N.; Lima, N.M.; Carlos, I.Z. Medicinal Plants from Brazilian Cerrado Biome: Potential sources of new anti-inflammatory compounds and antitumor agents on Ehrlich carcinoma. Acad. Bras. Ciênc. 2021, 93, e20191101. [Google Scholar] [CrossRef] [PubMed]
  23. Silva, D.H.S.; Castro-Gamboa, I.; Bolzani, V.D.S. Plant diversity from Brazilian Cerrado and Atlantic Forest as a tool for prospecting potential therapeutic drugs. Compr. Nat. Prod. II Chem. Biol. 2010, 3, 95–133. [Google Scholar]
  24. de Mesquita, M.L.; de Paula, J.E.; Pessoa, C.; de Moraes, M.O.; Costa-Lotufo, L.V.; Grougnet, R.; Michel, S.; Tillequin, F.; Espindola, L.S. Cytotoxic activity of Brazilian Cerrado plants used in traditional medicine against cancer cell lines. J. Ethnopharmacol. 2009, 123, 439–445. [Google Scholar] [CrossRef]
  25. Ribeiro Neto, J.A.; Pimenta Taroco, B.R.; Batista Dos Santos, H.; Thome, R.G.; Wolfram, E.; de A Ribeiro, R.I. Using the plants of Brazilian Cerrado for wound healing: From traditional use to scientific approach. J. Ethnopharmacol. 2020, 260, 112547. [Google Scholar] [CrossRef]
  26. de Giffoni de Carvalho, J.T.; da Silva Baldivia, D.; Leite, D.F.; de Araujo, L.C.A.; de Toledo Espindola, P.P.; Antunes, K.A.; Rocha, P.S.; de Picoli Souza, K.; Dos Santos, E.L. Medicinal Plants from Brazilian Cerrado: Antioxidant and Anticancer Potential and Protection against Chemotherapy Toxicity. Oxid. Med. Cell. Longev. 2019, 2019, 3685264. [Google Scholar] [CrossRef]
  27. Bailao, E.F.; Devilla, I.A.; da Conceicao, E.C.; Borges, L.L. Bioactive Compounds Found in Brazilian Cerrado Fruits. Int. J. Mol. Sci. 2015, 16, 23760–23783. [Google Scholar] [CrossRef]
  28. Fank-de-Carvalho, S.; Somavilla, N.; Marchioretto, M.; Báo, S. Biodiversity in Ecosystems—Linking Structure and Function; BoD -Books on Demand; IntechOpen: London, UK, 2015; 642p. [Google Scholar]
  29. Albuquerque, U.P.; Ramos, M.A.; Melo, J.G. New strategies for drug discovery in tropical forests based on ethnobotanical and chemical ecological studies. J. Ethnopharmacol. 2012, 140, 197–201. [Google Scholar] [CrossRef]
  30. de Melo, J.G.; Santos, A.G.; de Amorim, E.L.; do Nascimento, S.C.; de Albuquerque, U.P. Medicinal plants used as antitumor agents in Brazil: An ethnobotanical approach. Evid. Based Complement. Altern. Med. 2011, 2011, 365359. [Google Scholar] [CrossRef]
  31. Napolitano, D.R.; Mineo, J.R.; de Souza, M.A.; de Paula, J.E.; Espindola, L.S.; Espindola, F.S. Down-modulation of nitric oxide production in murine macrophages treated with crude plant extracts from the Brazilian Cerrado. J. Ethnopharmacol. 2005, 99, 37–41. [Google Scholar] [CrossRef]
  32. Ferreira, J.F.; Lopez, M.H.M.; Gomes, J.V.D.; Martins, D.H.N.; Fagg, C.W.; Magalhaes, P.O.; Davies, N.W.; Silveira, D.; Fonseca-Bazzo, Y.M. Seasonal Chemical Evaluation of Miconia chamissois Naudin from Brazilian Savanna. Molecules 2022, 27, 1120. [Google Scholar] [CrossRef]
  33. Suffness, M. Assays related to cancer drug discovery. Methods Plant Biochem. Assays Bioactivity 1990, 6, 71–133. [Google Scholar]
  34. Wang, J.; Jia, Z.; Zhang, Z.; Wang, Y.; Liu, X.; Wang, L.; Lin, R. Analysis of Chemical Constituents of Melastoma dodecandrum Lour. by UPLC-ESI-Q-Exactive Focus-MS/MS. Molecules 2017, 22, 476. [Google Scholar] [CrossRef]
  35. Xu, J.; Tong, C.; Fu, Q.; Guo, K.; Shi, S.; Xiao, Y. Comprehensive Polyphenolic Profile of Plantago depressa using High-Speed Countercurrent Chromatography Off-line with High-Performance Liquid Chromatography–Diode Array Detector–Quadrupole Time-of-flight Tandem Mass Spectrometry. eFood 2019, 1, 94–105. [Google Scholar] [CrossRef]
  36. Ameachi, N.C.; Chijioke, C.L. Evaluation of bioactive compounds in Pseudarenthemum tunicatum leaves using gas chromatography-mass spectrometry. Evaluation 2018, 18. [Google Scholar]
  37. Abdelkader, A.F.A.; Aldughaish, A.M. Physiological and Chemical Characteristics of Age-Differed Ficus Benjamina L. Trees Cultivated in El-Ahassa, Saudi Arabia. J. Plant Sci. 2016, 4, 63. [Google Scholar]
  38. Mala, M.; Saravanakumar, K. GC-MS Analysis of bioactive compounds in the Methanolicleaf extract of Memecylon edule Roxb. from Authukurichisacred grove, Tamilnadu, India. Life Sci. Arch. 2015, 1, 386–393. [Google Scholar]
  39. Cao, S.-G.; Wu, X.-H.; Sim, K.-Y.; Tan, B.; Pereira, J.; Goh, S.-H. Styryl-lactone derivatives and alkaloids from Goniothalamus borneensis (Annonaceae). Tetrahedron 1998, 54, 2143–2148. [Google Scholar] [CrossRef]
  40. Farag, M.A.; Al-Mahdy, D.A. Comparative study of the chemical composition and biological activities of Magnolia grandiflora and Magnolia virginiana flower essential oils. Nat. Prod. Res. 2013, 27, 1091–1097. [Google Scholar] [CrossRef]
  41. Iheagwam, F.N.; Israel, E.N.; Kayode, K.O.; De Campos, O.C.; Ogunlana, O.O.; Chinedu, S.N. GC-MS Analysis and Inhibitory Evaluation of Terminalia catappa Leaf Extracts on Major Enzymes Linked to Diabetes. Evid.-Based Complement. Altern. Med. Ecam 2019, 2019, 6316231. [Google Scholar]
  42. Seito, L.N.; Ruiz, A.L.; Vendramini-Costa, D.; Tinti, S.V.; de Carvalho, J.E.; Bastos, J.K.; Di Stasi, L.C. Antiproliferative activity of three methoxylated flavonoids isolated from Zeyheria montana Mart. (Bignoniaceae) leaves. Phytother. Res. 2011, 25, 1447–1450. [Google Scholar] [CrossRef] [PubMed]
  43. Miyashita, M.; Suzuki, T.; Yoshikoshi, A. Stereoselective total synthesis of (-)-picrotoxinin and (-)-picrotin. J. Am. Chem. Soc. 1989, 111, 3728–3734. [Google Scholar] [CrossRef]
  44. Kim, H.W.; Choi, S.Y.; Jang, H.S.; Ryu, B.; Sung, S.H.; Yang, H. Exploring novel secondary metabolites from natural products using pre-processed mass spectral data. Sci. Rep. 2019, 9, 17430. [Google Scholar] [CrossRef] [PubMed]
  45. Pang, Y.; Wang, D.; Fan, Z.; Chen, X.; Yu, F.; Hu, X.; Wang, K.; Yuan, L. Blumea balsamifera--a phytochemical and pharmacological review. Molecules 2014, 19, 9453–9477. [Google Scholar] [CrossRef] [PubMed]
  46. Lee, B.; Basavarajappa, H.D.; Sulaiman, R.S.; Fei, X.; Seo, S.Y.; Corson, T.W. The first synthesis of the antiangiogenic homoisoflavanone, cremastranone. Org. Biomol. Chem. 2014, 12, 7673–7677. [Google Scholar] [CrossRef]
  47. Hurtado Fernández, E. Avocado (Persea Americana): Complementarity of Different Omics Technologies for Its Metabolic Characterization; Universidad de Granada: Granada, Spain, 2014. [Google Scholar]
  48. Chen, Y.C.; Chiu, H.L.; Chao, C.Y.; Lin, W.H.; Chao, L.K.; Huang, G.J.; Kuo, Y.H. New Anti-Inflammatory Aromatic Components from Antrodia camphorata. Int. J. Mol. Sci. 2013, 14, 4629–4639. [Google Scholar] [CrossRef] [Green Version]
  49. Miceli, A.; Moncada, A.; Sabatino, L.; Vetrano, F. Effect of Gibberellic Acid on Growth, Yield, and Quality of Leaf Lettuce and Rocket Grown in a Floating System. Agronomy 2019, 9, 382. [Google Scholar] [CrossRef]
  50. Grimbs, A.; Shrestha, A.; Rezk, A.S.; Grimbs, S.; Hakeem Said, I.; Schepker, H.; Hütt, M.; Albach, D.C.; Brix, K.; Kuhnert, N.; et al. Bioactivity in Rhododendron: A Systemic Analysis of Antimicrobial and Cytotoxic Activities and Their Phylogenetic and Phytochemical Origins. Front. Plant Sci. 2017, 8, 551. [Google Scholar] [CrossRef]
  51. Kanchanapoom, T.; Kasai, R.; Yamasaki, K. Iridoid and phenolic diglycosides from Canthium berberidifolium. Phytochemistry 2002, 61, 461–464. [Google Scholar] [CrossRef]
  52. Kenfack, J.N.; Ponou, B.K.; Kühlborn, J.; Teponno, R.B.; Nono, R.N.; Fouedjou, R.T.; Opatz, T.; Park, H.J.; Tapondjou, L.A. A New Flavonol Glycoside from Tristemma hirtum (Melastomataceae). Nat. Prod. Sci. 2018, 24, 213–218. [Google Scholar] [CrossRef]
  53. Al-Zikri, P.N.H.; Taher, M.; Susanti, D.; Rezali, M.F.; Read, R.W.; Sohrab, M.; Hasan, C.M.; Ahmad, F. Cytotoxic tirucallane triterpenes from the stem of Luvunga scandens. Rev. Bras. Farm. 2014, 24, 561–564. [Google Scholar] [CrossRef]
  54. Hooi Poay, T.; Sui Kiong, L.; Cheng Hock, C. Characterisation of galloylated cyanogenic glucosides and hydrolysable tannins from leaves of Phyllagathis rotundifolia by LC-ESI-MS/MS. Phytochem. Anal. 2011, 22, 516–525. [Google Scholar] [CrossRef]
  55. Costa, M.F.; Jesus, T.I.; Lopes, B.R.; Angolini, C.F.; Montagnolli, A.; Gomes, L.P.; Pereira, G.S.; Ruiz, A.L.; Carvalho, J.E.; Eberlin, M.N.; et al. Eugenia aurata and Eugenia punicifolia HBK inhibit inflammatory response by reducing neutrophil adhesion, degranulation and NET release. Bmc Complement. Altern. Med. 2016, 16, 403. [Google Scholar] [CrossRef]
  56. Fank-de-Carvalho, S.M.; Somavilla, N.S.; Marchioretto, M.S.; Báo, S.N. Plant structure in the Brazilian neotropical savannah species. In Biodiversity in Ecosystems—Linking Structure and Function; BoD -Books on Demand; IntechOpen: London, UK, 2015; pp. 425–459. [Google Scholar]
  57. Silva, V.A.O.; Alves, A.L.V.; Rosa, M.N.; Silva, L.R.V.; Melendez, M.E.; Cury, F.P.; Gomes, I.N.F.; Tansini, A.; Longato, G.B.; Martinho, O.; et al. Hexane partition from Annona crassiflora Mart. promotes cytotoxity and apoptosis on human cervical cancer cell lines. Investig. New Drugs 2019, 37, 602–615. [Google Scholar] [CrossRef]
  58. dos Santos, K.M.; de Fatima Nunes, D.A.; Gomes, I.N.F.; da Silva, S.L.; de Azambuja Ribeiro, R.I.M. Inhibition of gelatinase activity of MMP-2 and MMP-9 by extracts of Bauhinia ungulata L. Biosci. J. 2015, 31, 584–590. [Google Scholar] [CrossRef]
  59. Faria, I.; Silva, A.; Lima, G.; Longatti, T.; Carmo, L.; Villar, J.; Araujo, A.; Thomé, R.; Santos, H.; Ribeiro, R. Alkaloid and phenolic compounds of Xylopia aromatica inhibits tumor growth by down-regulating matrix metalloproteinase-2 expression. Pak. J. Pharm. Sci. 2019, 34, 599–606. [Google Scholar]
  60. Silva-Oliveira, R.J.; Lopes, G.F.; Camargos, L.F.; Ribeiro, A.M.; Santos, F.V.; Severino, R.P.; Severino, V.G.; Terezan, A.P.; Thome, R.G.; Santos, H.B.; et al. Tapirira guianensis Aubl. Extracts Inhibit Proliferation and Migration of Oral Cancer Cells Lines. Int. J. Mol. Sci. 2016, 17, 1839. [Google Scholar] [CrossRef] [Green Version]
  61. Santos, K.; Gomes, I.; Silva-Oliveira, R.; Pinto, F.; Oliveira, B.; Chagas, R.; Romão, W.; Reis, R.M.; Ribeiro, R. Bauhinia variegata candida Fraction Induces Tumor Cell Death by Activation of Caspase-3, RIP, and TNF-R1 and Inhibits Cell Migration and Invasion In Vitro. Biomed Res. Int. 2018, 2018, 4702481. [Google Scholar] [CrossRef]
  62. Rosa, M.N.; LRV, E.S.; Longato, G.B.; Evangelista, A.F.; Gomes, I.N.F.; Alves, A.L.V.; de Oliveira, B.G.; Pinto, F.E.; Romao, W.; de Rezende, A.R.; et al. Bioprospecting of Natural Compounds from Brazilian Cerrado Biome Plants in Human Cervical Cancer Cell Lines. Int. J. Mol. Sci. 2021, 22, 3383. [Google Scholar] [CrossRef]
  63. Lemee, J.M.; Clavreul, A.; Menei, P. Intratumoral heterogeneity in glioblastoma: Don’t forget the peritumoral brain zone. Neuro-Oncol. 2015, 17, 1322–1332. [Google Scholar] [CrossRef]
  64. Bermejo, A.; Figadere, B.; Zafra-Polo, M.C.; Barrachina, I.; Estornell, E.; Cortes, D. Acetogenins from Annonaceae: Recent progress in isolation, synthesis and mechanisms of action. Nat. Prod. Rep. 2005, 22, 269–303. [Google Scholar] [CrossRef] [PubMed]
  65. Ishimaru, C.; Takeuchi, T.; Yonezawa, Y.; Kuriyama, I.; Takemura, M.; Kato, I.; Sugawara, F.; Yoshida, H.; Mizushina, Y. Inhibitory effect of acetogenins on mammalian DNA polymerase and human cancer cell growth. Lett. Drug Des. Discov. 2007, 4, 239–245. [Google Scholar] [CrossRef]
  66. Takahashi, S.; Yonezawa, Y.; Kubota, A.; Ogawa, N.; Maeda, K.; Koshino, H.; Nakata, T.; Yoshida, H.; Mizushina, Y. Pyranicin, a non-classical annonaceous acetogenin, is a potent inhibitor of DNA polymerase, topoisomerase and human cancer cell growth. Int. J. Oncol. 2008, 32, 451–458. [Google Scholar] [CrossRef] [PubMed]
  67. Vasconcelos, M.A.; Royo, V.A.; Ferreira, D.S.; Crotti, A.E.; Andrade e Silva, M.L.; Carvalho, J.C.; Bastos, J.K.; Cunha, W.R. In vivo analgesic and anti-inflammatory activities of ursolic acid and oleanoic acid from Miconia albicans (Melastomataceae). Z. Fur Nat. C J. Biosci. 2006, 61, 477–482. [Google Scholar] [CrossRef]
  68. Serna, D.M.; Martinez, J.H. Phenolics and Polyphenolics from Melastomataceae Species. Molecules 2015, 20, 17818–17847. [Google Scholar] [CrossRef]
  69. Sharma, S.B.; Gupta, R. Drug development from natural resource: A systematic approach. Mini Rev. Med. Chem. 2015, 15, 52–57. [Google Scholar] [CrossRef]
  70. Zhang, Y.; Xie, R.F.; Xiao, Q.G.; Li, R.; Shen, X.L.; Zhu, X.G. Hedyotis diffusa Willd extract inhibits the growth of human glioblastoma cells by inducing mitochondrial apoptosis via AKT/ERK pathways. J. Ethnopharmacol. 2014, 158 Pt A, 404–411. [Google Scholar] [CrossRef]
  71. Chang, H.F.; Huang, W.T.; Chen, H.J.; Yang, L.L. Apoptotic effects of gamma-mangostin from the fruit hull of Garcinia mangostana on human malignant glioma cells. Molecules 2010, 15, 8953–8966. [Google Scholar] [CrossRef]
  72. Liu, B.; Gao, Y.Q.; Wang, X.M.; Wang, Y.C.; Fu, L.Q. Germacrone inhibits the proliferation of glioma cells by promoting apoptosis and inducing cell cycle arrest. Mol. Med. Rep. 2014, 10, 1046–1050. [Google Scholar] [CrossRef]
  73. Gentile, M.T.; Ciniglia, C.; Reccia, M.G.; Volpicelli, F.; Gatti, M.; Thellung, S.; Florio, T.; Melone, M.A.; Colucci-D’Amato, L. Ruta graveolens L. induces death of glioblastoma cells and neural progenitors, but not of neurons, via ERK 1/2 and AKT activation. PLoS ONE 2015, 10, e0118864. [Google Scholar] [CrossRef]
  74. Flores, B.C.; Klinger, D.R.; Welch, B.G.; White, J.A.; Batjer, H.H.; Samson, D.S. Management of intracranial aneurysms associated with arteriovenous malformations. Neurosurg. Focus 2014, 37, E11. [Google Scholar] [CrossRef]
  75. Oberoi, R.K.; Parrish, K.E.; Sio, T.T.; Mittapalli, R.K.; Elmquist, W.F.; Sarkaria, J.N. Strategies to improve delivery of anticancer drugs across the blood-brain barrier to treat glioblastoma. Neuro Oncol. 2016, 18, 27–36. [Google Scholar] [CrossRef]
  76. Patel, A.P.; Tirosh, I.; Trombetta, J.J.; Shalek, A.K.; Gillespie, S.M.; Wakimoto, H.; Cahill, D.P.; Nahed, B.V.; Curry, W.T.; Martuza, R.L.; et al. Single-cell RNA-seq highlights intratumoral heterogeneity in primary glioblastoma. Science 2014, 344, 1396–1401. [Google Scholar] [CrossRef]
  77. Carocho, M.; Ferreira, I.C. The role of phenolic compounds in the fight against cancer--a review. Anti-Cancer Agents Med. Chem. 2013, 13, 1236–1258. [Google Scholar] [CrossRef]
  78. Jiang, L.; Wang, W.; He, Q.; Wu, Y.; Lu, Z.; Sun, J.; Liu, Z.; Shao, Y.; Wang, A. Oleic acid induces apoptosis and autophagy in the treatment of Tongue Squamous cell carcinomas. Sci. Rep. 2017, 7, 11277. [Google Scholar] [CrossRef]
  79. Zhu, S.; Jiao, W.; Xu, Y.; Hou, L.; Li, H.; Shao, J.; Zhang, X.; Wang, R.; Kong, D. Palmitic acid inhibits prostate cancer cell proliferation and metastasis by suppressing the PI3K/Akt pathway. Life Sci. 2021, 286, 120046. [Google Scholar] [CrossRef]
  80. Shin, H.E.; Lee, S.; Choi, Y.; Park, S.; Kwon, S.; Choi, J.K.; Seo, S.Y.; Lee, Y. Synthetic Homoisoflavane Derivatives of Cremastranone Suppress Growth of Colorectal Cancer Cells through Cell Cycle Arrest and Induction of Apoptosis. Biomol. Ther. 2022, 30, 576–584. [Google Scholar] [CrossRef]
  81. Diantini, A.; Subarnas, A.; Lestari, K.; Halimah, E.; Susilawati, Y.; Supriyatna, S.; Julaeha, E.; Achmad, T.H.; Suradji, E.W.; Yamazaki, C.; et al. Kaempferol-3-O-rhamnoside isolated from the leaves of Schima wallichii Korth. inhibits MCF-7 breast cancer cell proliferation through activation of the caspase cascade pathway. Oncol. Lett. 2012, 3, 1069–1072. [Google Scholar] [CrossRef]
  82. Silva, A.G.; Silva, V.A.O.; Oliveira, R.J.S.; de Rezende, A.R.; Chagas, R.C.R.; Pimenta, L.P.S.; Romao, W.; Santos, H.B.; Thome, R.G.; Reis, R.M.; et al. Matteucinol, isolated from Miconia chamissois, induces apoptosis in human glioblastoma lines via the intrinsic pathway and inhibits angiogenesis and tumor growth in vivo. Investig. New Drugs 2020, 38, 1044–1055. [Google Scholar] [CrossRef]
  83. Teixeira, S.A.; Luzzi, M.C.; Martin, A.C.B.M.; Duarte, T.T.; Leal, M.O.; Teixeira, G.R.; Reis, M.T.; Junior, C.R.A.; Santos, K.; Melendez, M.E.; et al. The Barretos Cancer Hospital Animal Facility: Implementation and Results of a Dedicated Platform for Preclinical Oncology Models. Vet. Sci. 2022, 9, 636. [Google Scholar] [CrossRef]
  84. Cruvinel-Carloni, A.; SilvaOliveira, R.; Torrieri, R.; Bidinotto, L.T.; Berardinelli, G.N.; Oliveira-Silva, V.A.; Clara, C.A.; de Almeida, G.C.; Martinho, O.; Squire, J.A.; et al. Molecular characterization of short-term primary cultures and comparison with corresponding tumor tissue of Brazilian glioblastoma patients. Transl. Cancer Res. 2017, 6, 332–345. [Google Scholar] [CrossRef] [Green Version]
Molecules 28 01394 g001 550
Figure 1. Time and concentration cytotoxic effect of the crude extract’s exposure on the human cancer cell line. Cells (5 × 103) were plated into 96-well plates, exposed to the vehicle (1% DMSO) or increasing concentrations of the samples (A) crude extracts (1, 2, 3 and 7) 12 μg/mL to 300 μg/mL, (B) crude extracts (8, 17, 18 and 19) 1.5 μg/mL to 300 μg/mL and (C) crude extracts (10, 14I, 15I, 16I and 21I) 3 μg/mL to 300 μg/mL. Cell viability was measured with an MTS assay after 72 h of extract treatment. The cell viability of the untreated cells was regarded as 100%. The results shown are the means ± S.D of three independent experiments.
Figure 1. Time and concentration cytotoxic effect of the crude extract’s exposure on the human cancer cell line. Cells (5 × 103) were plated into 96-well plates, exposed to the vehicle (1% DMSO) or increasing concentrations of the samples (A) crude extracts (1, 2, 3 and 7) 12 μg/mL to 300 μg/mL, (B) crude extracts (8, 17, 18 and 19) 1.5 μg/mL to 300 μg/mL and (C) crude extracts (10, 14I, 15I, 16I and 21I) 3 μg/mL to 300 μg/mL. Cell viability was measured with an MTS assay after 72 h of extract treatment. The cell viability of the untreated cells was regarded as 100%. The results shown are the means ± S.D of three independent experiments.
Molecules 28 01394 g001
Table
Table 1. Mean and standard deviation of IC50 (µg/mL) values for crude leaf extracts from plants of the Brazilian cerrado biome and the chemotherapy temozolomide, evaluated at 72 h.
Table 1. Mean and standard deviation of IC50 (µg/mL) values for crude leaf extracts from plants of the Brazilian cerrado biome and the chemotherapy temozolomide, evaluated at 72 h.
IC50 (µg/mL) Mean ± Standard Deviation (SD)
Crude Extract/ChemotherapyNHAGAMGU251-MGSF188RES259HCB151
136.38 ± 11.3<1.5146.1 ± 35.872.72 ± 8.389.92 ± 71.525.93 ± 12.9
213.4 ± 2.3<1.5>300118.5 ± 1.7124.7 ± 188.172.85 ± 26.9
3>200<1.5120.6 ± 10.957.45 ± 13.770.93 ± 4054.78 ± 22
741.30 ± 18.21<1.5117.2 ± 53.543.75 ± 1.450.40 ± 3928.69 ± 13.4
836.19 ± 20.16.66 ± 4.3176.5 ± 74.8866.20 ± 5.386.80 ± 43.5125.7 ± 10.9
1021.46 ± 1.918.83 ± 4.6770.52 ± 6118.45 ± 16.944.15 ± 43.9145.7 ± 19.7
14-I22.84 ± 14.05<1.577.68 ± 20.126.55 ± 12.474.76 ± 1.555.30 ± 0
15-I35.68 ± 37.1<1.5136.4 ± 33.634.31 ± 8106.3 ± 5.281.95 ± 0
16-I62.67 ± 50.2<1.5296.9 ± 23.2100.5 ± 16.6>200208.1 ± 30.1
172.9 ± 1.425.7 ± 12.928.9 ± 5.513.6 ± 5.729.3 ± 17.430.2 ± 10.6
1812.05 ± 1.119.94 ± 0.470.21 ± 7.423.08 ± 3.638.07 ± 5.730.05 ± 3.9
1912.11 ± 5.38.37 ± 11.889.85 ± 519.28 ± 8.333.04 ± 12.027.43 ± 6.2
21-I3.73 ± 3.7<1.5132.5 ± 32.56.56 ± 0.637.04 ± 8.447.52 ± 0
TMZ1.28 ± 26.267.06 ± 31.8439.78 ± 23.6730.74 ± 27.9116.93 ± 27.5538.22 ± 26.02
TMZ (temozolomide).
Table
Table 2. Selectivity index for crude leaf extracts from plants of Brazilian cerrado biome and chemotherapy temozolomide.
Table 2. Selectivity index for crude leaf extracts from plants of Brazilian cerrado biome and chemotherapy temozolomide.
Crude Extract/ChemotherapyGlioma Cell Lines
GAMGU251-MGSF188RES259HCB151
1UD0.20.50.41.4
2UDUD0.10.10.2
3UDIDUDUDUD
7UD0.40.90.81.4
85.40.20.50.40.3
101.10.31.20.50.1
14-IUD0.30.90.30.4
15-IUD0.31.00.30.4
16-IUD0.20.6UD0.3
170.10.10.20.10.1
180.60.20.50.30.4
191.40.10.60.40.4
21-IUD0.030.60.10.1
TMZ0.20.030.00.10.03
UD—undetermined values. TMZ (temozolomide).
Table
Table 3. IC50 values for crude extract and partitions from Miconia chamissois and chemotherapy, temozolomide, evaluated at 72 h.
Table 3. IC50 values for crude extract and partitions from Miconia chamissois and chemotherapy, temozolomide, evaluated at 72 h.
IC50 (µg/mL) Mean ± Standard Deviation (SD)
Crude Extract/Partitions/ChemotherapyNHAGAMGU251-MGSF188RES259HCB151
CE12.11 ± 5.358.37 ± 11.8189.85 ± 5.0919.28 ± 8.2833.04 ± 12.0827.43 ± 6.20
HDP>200>200>200>200>200>200
HP9.17 ± 0.9937.39 ± 5.2234.66 ± 5.455.35 ± 2.2718.22 ± 4.5821.76 ± 6.74
CP14.06 ± 2.4134.31 ± 5.5021.54 ± 1.807.28 ± 3.4020.83 ± 4.9115.00 ± 1.00
AEP31.22 ± 10.402.57 ± 3.34102.4 ± 2.6141.69 ± 6.0627.40 ± 4.2534.26 ± 4.20
TMZ1.28 ± 26.267.06 ± 31.8439.78 ± 23.6730.74 ± 27.9116.93 ± 27.5538.22 ± 26.02
CE (crude extract); HDP (hydroalcoholic partition); HP (hexane partition); CP (chloroform partition); AEP (ethyl acetate partition); temozolomide (TMZ).
Table
Table 4. Selectivity index for crude extracts and partitions from Miconia chamissois and chemotherapy, temozolomide.
Table 4. Selectivity index for crude extracts and partitions from Miconia chamissois and chemotherapy, temozolomide.
Crude Extract/Partitions/ChemotherapyCell Lines
GAMGU251-MGSF188RES259HCB151
CE1.40.10.60.40.4
HDPUDUDUDUDUD
HP0.20.31.70.50.4
CP0.40.71.90.70.9
AEP12.10.30.71.10.9
TMZ0.20.030.040.10.03
UD—undetermined values. CE (crude extract); HDP (hydroalcoholic partition); HP (hexane partition); CP (chloroform partition); AEP (ethyl acetate partition); temozolomide (TMZ).
Table
Table 5. IC50 values for crude leaf extract and partitions from Miconia chamissois.
Table 5. IC50 values for crude leaf extract and partitions from Miconia chamissois.
IC50 (µg/mL) Mean ± Standard Deviation (SD)
Cell LinesCrude Extract/Partition6 h12 h24 h48 h
NHACE>300>300>30064.94 ± 23.29
HP>20077.66 ± 24.0237.64 ± 32.2130.55 ± 37.02
CP>20061.39 ± 21.9835.13 ± 26.5820.93 ± 32.62
GAMGCE>300>30089.29 ± 20.3335.98 ± 26.61
HP>20049.20 ± 21.8555.95 ± 26.2434.78 ± 32.96
CP51.52 ± 18.4332.17 ± 24.6440.11 ± 26.1339.14 ± 23.52
U251-MGCE>300>300>300>300
HP>200>200>200>200
CP>200>200>20069.03 ± 30.89
SF188CE>30091.88 ± 17.8656.87 ± 24.3815.42 ± 32.78
HP>20051.88 ± 24.8426.54 ± 31.3516.46 ± 34.98
CP>20041.58 ± 22.6122.35 ± 30.206.34 ± 32.81
RES259CE>300>300>30020.45 ± 24.31
HP>20093.91 ± 19.8966.21 ± 30.0722.89 ± 32.60
CP>200>20064.72 ± 25.959.16 ± 35.78
CE (crude extract), HP (hexane partition) and CP (chloroform partition).
Table
Table 6. Proposed structures by ESI (-) FT-ICR MS for the main molecules of Miconia chamissois crude extract.
Table 6. Proposed structures by ESI (-) FT-ICR MS for the main molecules of Miconia chamissois crude extract.
m/z Measuredm/z TheoreticalError (ppm)DBE[M-H]Proposed CompoundReference
191.05620191.05602−0.452[C7H11O6]chlorogenic acid[34]
197.04549197.165110.275[C9H9O5]syringic acid[35]
219.05098219.169070.233[C8H11O7]2-hydroxypropane-1,2,3-tricarboxylic acid, dimethyl ester[36]
229.07151229.071301.114[C10H13O6]D-(-) Erythrose[37]
255.23294255.232950.061[C16H31O2]palmitic acid[38]
265.10783266.107891.196[C14H17O5]goniothalesdiol[39]
277.21771277.25421−1.464[C19H29O2]linolenic acid[40]
281.24860281.454100.012[C18H33O2]oleic acid[41]
299.09247299.298640.1110[C17H15O5]5-hydroxy-6,7-dimethoxyflavanone[42]
309.09756311.378501.357[C15H17O7]picrotin[43]
313.10808313.325260.2010[C18H17O5]agrimonolide[44]
331.08231331.297450.0610[C17H15O7]dihydroquercetin-7,4’-dimethylether[45]
345.09851345.32300−1.5510[C18H17O7]cremastranone[46]
351.12974351.32698−0.193[C14H23O10]methyl 4-(6.7-dideoxy-galacto-hept-6-enopyranosyl)-galactopyranoside[47]
363.10909363.10800−1.529[C18H19O8]4,4’-Dihydroxy-3,3’-dimethoxy-2,2’-dimethyl-5,6,5’,6’-bimethylenedioxybiphenyl[48]
379.13921375.134101.678[C19H23O8]gibberellic Acid[49]
431.09823431.370370.3312[C21H19O10]kaempferol-o-rhamnoside[50]
445.13427445.134001.977[C19H25O12]canthoside A[51]
447.09333447.36978−0.0912[C21H19O11]6-hydroxyapigenin-7-o-β-d-glucopyranoside[52]
455.35390455.35100−1.827[C30H47O3]3-Oxotirucalla-7,24-dien-21-oic acid[53]
483.07802483.357260.0211[C20H19O14]3, 6-di-o-galloyl-d-glucose[54]
593.15123593.51122−0.0613[C27H29O15]rutinosylkaempferol[55]
635.08917635.46179−0.2816[C27H23O18]1,2,3-tri-o-galloyl-β-d-glucose[54]
DBE: Double bond equivalent; m/z: mass-to-charge ratio.
Table
Table 7. Culture conditions and origin of the cell lines used.
Table 7. Culture conditions and origin of the cell lines used.
Cell LinesTypeSubtypeGrade (OMS)DescriptionSexAgeMainly MutationsOriginCulture Conditions
GAMGEstablishedGBMIVAdultFemale42 yearsTP53 and TERTDSMZDMEM + 10% FBS + 1% P/S
U251-MGEstablishedGBMIVAdultMale75 yearsCDKN2A/B, EGFR, TP53, PTEN and TERTKindly provided by Dr. Joseph CostelloDMEM + 10% FBS + 1% P/S
SF188EstablishedGBMIVPaediatricMale8 yearsTP53 and NF1Kindly provided by Dr. Chris JonesDMEM + 10% FBS + 1% P/S
RES259EstablishedDAIIPaediatricFemale4 yearsTERTKindly provided by Dr. Chris JonesDMEM + 10% FBS + 1% P/S
HCB151PrimaryGBMIVAdultMale59 yearsCDKN2A and PTENBarretos Cancer HospitalDMEM + 10% FBS + 1% P/S
NHAEstablished--NANANA-ECACCDMEM + 10% FBS + 1% P/S
GBM: Glioblastoma, DA: Diffuse astrocytoma; NA = Not available; NHA: Normal Human Astrocyte, DSMZ—German Collection of Microorganisms and Cell Cultures. ECACC (European Collection of Authenticated Cultures). FBS: Fetal Bovine Serum; P/S Penicillin/Streptomycin solution.
Table
Table 8. Classification of natural extracts from the Brazilian cerrado biome.
Table 8. Classification of natural extracts from the Brazilian cerrado biome.
Crude Extract IdentificationVernacular NameBotanical NameFamilyRegistration Code
1Pau PomboTapirira guianensisFabaceae143407BHCB
2Gonçalo-AlvesAstronium fraxinifoliumAnacardiaceae143403BHCB
3Pimenta-de-MacacoXylopia aromaticaAnnonaceae43397BHCB
7AraticumAnnona crassifloraAnnonaceae143400BHCB
8NegraminaSiparuna guianensisSiparunaceae143404BHCB
10MarcelaAchyrocline alataAsteraceae11486CG/MS
14-IPata-de-vacaBauhinia variegataFabaceae161589BHCB
15-IPata-de-vaca brancaBauhinia variegata candidaFabaceae161590BHCB
16-IPata-de-viadoBauhinia ungulataFabaceae161588BHCB
17Pixirica-da-mataMiconia cuspidataMelastomataceae44998HUFU
18Canela de velhoMiconia albicansMelastomataceae56558 HUFU
19Pixirica-açuMiconia chamissoisMelastomataceae59592HUFU
21-IBarbatimãoStryphnodendron adstringensFabaceae169871BHCB
HUFU—herbarium of the Federal University of Uberlândia; BHCB—herbarium of the Federal University of Minas Gerais; CG/MS—herbarium of the Federal University of Grande Dourados.
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MDPI and ACS Style

Silva, V.A.O.; Rosa, M.N.; Gomes, I.N.F.; Vital, P.d.S.; Alves, A.L.V.; Evangelista, A.F.; Longato, G.B.; Carloni, A.C.; Oliveira, B.G.; Pinto, F.E.; et al. Bio-Prospecting of Crude Leaf Extracts from Thirteen Plants of Brazilian Cerrado Biome on Human Glioma Cell Lines. Molecules 2023, 28, 1394. https://doi.org/10.3390/molecules28031394

AMA Style

Silva VAO, Rosa MN, Gomes INF, Vital PdS, Alves ALV, Evangelista AF, Longato GB, Carloni AC, Oliveira BG, Pinto FE, et al. Bio-Prospecting of Crude Leaf Extracts from Thirteen Plants of Brazilian Cerrado Biome on Human Glioma Cell Lines. Molecules. 2023; 28(3):1394. https://doi.org/10.3390/molecules28031394

Chicago/Turabian Style

Silva, Viviane A. O., Marcela N. Rosa, Izabela N. F. Gomes, Patrik da Silva Vital, Ana Laura V. Alves, Adriane F. Evangelista, Giovanna B. Longato, Adriana C. Carloni, Bruno G. Oliveira, Fernanda E. Pinto, and et al. 2023. "Bio-Prospecting of Crude Leaf Extracts from Thirteen Plants of Brazilian Cerrado Biome on Human Glioma Cell Lines" Molecules 28, no. 3: 1394. https://doi.org/10.3390/molecules28031394

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