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Introduction
Starch is a polysaccharide widely used in industry due to its high availability, low cost, and great versatility in its applications, highlighting its use as a thickening, stabilizing, and emulsifying agent in the food and paper industry (Schmiele et al., 2019; Koev et al., 2020). Industrially, starches are extracted from potato (Solanum tuberosum L.), corn (Zea mays L.), rice (Oryza sativa L.), and wheat (Triticum aestivum L.), cataloged as conventional sources (Carvalho, 2013). Given the growing demand for alternative and innovative food products with nutraceutical characteristics beneficial to health and the increase in the consumption of ultra-processed foods, research has been conducted to evaluate the use of banana starches (Musa paradisiaca L.), mango (Mangifera indica L.), cherimoya (Annona cherimola L.) and soursop (Annona muricata L.) because these starches have resistance to enzymatic hydrolysis and a low tendency to syneresis, as well as content of phytochemical compounds (Martínez-Ortiz et al., 2022; Espinosa-Solís et al., 2009). Phytochemical compounds are secondary metabolites produced by plants, these have antioxidant activities mainly (Zubaidi et al., 2023). Soursop fruits have phenols, flavonoids, tannins, alkaloids, phytosterols, and acetogenins (Correa-Gordillo et al. 2012; Terán-Erazo et al., 2019; Aguilar-Hernández et al., 2020a), the latter has been identified qualitatively in soursop fruit starches (Ramírez-Balboa et al., 2021) and have been studied by determining the content of this compound in seeds and pulp, as well as its biological activity (Grba et al., 2022; Prasad et al., 2021; Mutakin et al., 2022). Phytochemical compounds are present in soursop fruit starches forming binary (amylose-lipid), tertiary (amylose-lipid-protein), and quaternary (amylose-lipid-protein-phenol) interactions (Wang et al., 2019; Krishnan et al., 2021), in this regard, the phytochemical compound content of starches is directly proportional to the amylose percentage of starches and lipids (Krishnan et al., 2021). These compounds present in starches can exert beneficial health effects directly, mainly in the digestive tract, or indirectly through their prebiotic activities on the intestinal microbiota and the formation of bioactive bacterial metabolites that act both locally and systemically (Gasaly et al., 2020). In this sense, this study aimed to determine the content of phytochemical compounds in starches extracted from soursop fruits by conventional method and ultrasound.
Materials and Methods
Biological material
Soursop fruits were harvested, considering the harvest index (160 days after anthesis) reported by Balois-Morales et al. (2019), the harvest was in the ejido of Venustiano Carranza, Tepic, Nayarit (21° 32' 2.77'' N, 104° 58'37.73'' W, 893 masl). The fruits were transported to the Food Technology Unit, selecting those that did not present mechanical and microbiological visual damage, and washed with a solution of sodium hypochlorite 1% (v / v).
Starch extraction
The research consisted of extracting starches from whole soursop fruits without separating their structures (epicarp, mesocarp, and endocarp) using two extraction methods (conventional and ultrasound):
Conventional starch extraction
It was performed according to the technique of Flores-Gorosquera et al. (2004). The fruits were ground in an industrial blender (International LI-5a, Mexico) for 3 min with a solution of citric acid (1%) in a ratio of 1:4 (w / v), the resulting paste was filtered with a sieve No. 100 (150 mics) then in a sieve of No. 200 (75 mics) and 270 (53 mics), respectively. Subsequently, a second grinding with distilled water was carried out in a ratio of 1:2 (m / v) for 3 min and a second filtering.
Ultrasound-assisted starch extraction
The technique described by Flores-Gorosquera et al. (2004) and Ramírez-Balboa et al. (2021) were used. Fruits were ground for 3 min with a solution of citric acid (1%) in a ratio of 1:4 (w / v). Subsequently, the resulting paste was sonicated at 300 W power in a 750 W ultrasonic processor (Model CPX750 Cole-Parmer Instruments Vernon Hills, U.S.A.) with 20 kHz probe for 10 min at 25 °C monitored by a manual mercury thermometer (Ramírez-Balboa et al., 2021). The sonicated paste was filtered using sieves of the numbers 100, 200, and 270. Subsequently, a second grinding was carried out using distilled water in a ratio of 1:2 (m / v) and then the precipitate (starch) was recovered.
To remove excess water in starches, a centrifuge was used (Hermle Z326K Wehingen, Germany). During this process, it was observed that the starches when centrifuged were separated into two phases (phase 1 and phase 2), so it was decided to evaluate both phases to determine their physicochemical and phytochemical characteristics.
The starch extracted by conventional method and ultrasound was centrifuged at 1537 g for 5 min at 25 °C to precipitate and separate the phases that were identified as phase 1 (brown color) and phase 2 (white color). Phase 1 was separated with a spatula, leaving phase 2 at the bottom of the tube. Subsequently, 10 mL of water was added to the tube and stirred (first wash), then centrifuged at 1537 g for 5 min at 25 °C repeating the phase separation process. This process was done in triplicate. The starches (phase 1 and phase 2) were dried in a recirculating furnace (LSIS-B2V/VC 55, Germany) for 24 h at 35 °C, pulverized, and sieved (No.100). Finally, they were weighed for quantification and stored. The following nomenclature was used to better interpret the results: ACF1, ACF2, AUF1, and AUF2. The letter A (starch), C (conventional extraction), U (ultrasound-assisted extraction), F (phases); e.g., starch extraction by conventional method phase 1 (ACF1).
Variables evaluated
Lipid content (%)
It was determined using the AOAC technique (2005). 4 g of starch was added to cellulose cartridges and placed in a grease extractor. 130 mL of petroleum ether in a flask was added at a constant weight. It was left to boil for 4 h. The calculation was determined by the weight difference in the flask.
Amylose content (%)
It was performed using the Hoover & Ratnayake technique (2001), 100 mg of dry and defatted starch was diluted in 10 mL of dimethyl sulfoxide (DMSO) (90% w/v) and heated in a water bath (Thermo Scientific, TSGP10, Canada) at 85 ° C/15 min in stirring (solution 1). Subsequently, 100 mL was measured with distilled water (solution 2) and 20 mL of the sample was taken at 25 mL (solution 3). Then 1 mL of solution 3 was taken and 5 mL of iodine were added, adjusting the solution to 50 mL. It was allowed to stand 15 min in darkness and the absorbance was read at 600 nm. A standard potato amylose and amylopectin curve was used.
Analysis of phytochemical compounds
Obtaining methanolic starch extract
In 100 mL of methanol, 20 g of starch (ACF1, AUF1, ACF2, and AUF2) were added. The resulting solution was sonicated in an ultrasonic bath (Luzerner, CD-4820, China) at 35 kHz and 160 W for eight cycles of 8 min each, it was stirred at the end of each cycle, then centrifuged at 6147 g / 10 min and the liquid phase was recovered, the extracts were filtered with a vacuum pump (EVAR model EV-40, Mexico) in a porcelain funnel, using closed-pore filter paper and subsequently concentrating on a rotary vapor (IKA© model RV-10 Digital V, North Carolina, USA) at 40 °C at reduced pressure (65 cmHg), the concentrated extracts were subjected to 35 ± 2 °C in an oven (LSIS-B2V/VC 55, Germany) for 48 h to evaporate the solvent residues (León-Fernández et al., 2021).
Qualitative analysis of phytochemical compounds of the starch extract
The extracts were diluted to a concentration of 4 mg / mL in methanol. Subsequently, the procedures of Sofowara (1993), Harborne (1998) & Evans (2009) were used to determine the presence of the following compounds:
Phytosterols
100 μL of extract, 1 mL of chloroform, and 100 μL concentrated sulfuric acid were added to a test tube. The reddish-brown precipitate at the bottom of the tube indicated the presence (Trease & Evans 1989; Evans, 2002).
Phenols and tannins
In a test tube, 300 μL of extract and 300 μL of ferric chloride (FeCl3) (10%) were placed. Blue or green precipitate indicated presence (Trease & Evans 1989; Evans, 2002).
Alkaloids
In a test tube were placed 300 μL of extract and 300 μL of hydrochloric acid (HCl) at 2 N and 300 μL of Mayer's reagent, the presence of a pale precipitate at the bottom of the tube, indicates the presence of alkaloids (Trease & Evans 1989; Evans, 2002).
Flavonoids
In a test tube, 300 μL of extract were placed, 333 μL of 10% ammonia (NH4OH), and 333 μL of H2SO4 were added. When the yellow color disappears in the sample, the presence is indicated (Trease & Evans 1989; Evans, 2002).
Acetogenins
1 mL of extract was added to a tube and Kedde's reagent was added (Gu et al., 1995) which was prepared with the mixture of solution A (3-5 dinitrobenzoic acid) and solution B (potassium hydroxide KOH 5.7%), 2 mL each. The pink coloration determined the presence of acetogenins (León-Fernández et al., 2021).
The results were expressed with the presence (+) or absence (-) of the phytochemical compound evaluated.
Quantitative analysis of phytochemical compounds
Total soluble phenols
They were determined according to the methodology of Stintzing et al. (2005). 50 μL of the sample was added in Eppendorf vials, then 250 μL of Folin-Ciocalteu solution (v/v 1:10 in deionized water) and 200 μL of sodium carbonate solution (7.5%) were added, the solution was stirred in a vortex (Genie® G650, U.S.A) and incubated at 23 °C for 30 min in the absence of light. Absorbance was measured in a microplate reader (Power Wave XS, Biotek, U.S.A) at a wavelength of 765 nm. The results obtained were expressed in mg gallic acid equivalents per gram of dry base starch (mg GAE/g of d.b.s).
Total soluble tannins
It was determined by the Folin-Ciocalteu + PVPP technique (polyvinylpolypidorrilone) according to the method of Makkar (2003), 100 mg of PVPP was diluted in 1 mL of distilled water and left to stand for 2 h. 200 μL of sample was added and stored at 4 °C for 15 min, then centrifuged at 6147 g for 10 min and the supernatant containing non-tannin phenols was recovered. 50 μL of supernatant was taken and the total phenol content was evaluated using the method of Stintzing et al. (2005). Absorbance was measured in a microplate reader (Power Wave XS, Biotek, U.S.A.) at a wavelength of 765 nm. The result obtained was subtracted from the content of total soluble phenols found in the sample without PVPP. The results obtained were expressed in mg gallic acid equivalents per gram of dry base starch (mg GAE/g of d.b.s).
The calculation of total soluble tannins was performed using the following equation (equation 1):
Equation 1:
Where:
TT= total soluble tannins.
Fn= total soluble phenols.
Fnt= soluble phenols, not tannins.
Total soluble flavonoids
It was performed using the methodology developed by Zhishen et al. (1999), 50 μL of the sample was added in Eppendorf tubes, and 100 μL of deionized water. Subsequently, 10 μL of NaOH (15%) was added, stirred in a vortex (Genie G650, U.S.A.), and rested for 6 min in darkness at 23 °C. After a time, 15 μL of AlCl3 (10%) was added and stirred in a vortex (Genie G650, U.S.A.) tag. The solution was rested for 6 min in darkness at 23 °C. Finally, 200 μL of NaOH (4%) was added and stirred. Absorbance was measured at 510 nm (Power Wave XS, Biotek, U.S.A.). The results obtained were expressed in mg of quercetin equivalent per gram of dry base starch (mg QE/g of d.b.s).
Total phytosterols
The total phytosterol content was determined using the Lieberman-Buchard method (Lobato et al., 2008). In a glass reaction tube, 50 μL of sample and 50 μL of chloroform were added, the mixture was stirred and an aliquot of 50 μL was taken by adding 50 μL of chloroform and 1 mL of Lieberman-Buchard reagent prepared with acetic anhydride and sulfuric acid (20:1 v/v) cooled to 4 °C. The mixture was subjected to a water bath (Thermo Scientific, TSGP10, U.S.A.) at 35 ± 2 °C for 25 min. After a time, 200 μL was taken and the absorbance was read at 550 (Power Wave XS, Biotek, U.S.A.). The results obtained were expressed in mg cholesterol equivalents per gram of dry base starch (mg CE/g d.b.s).
Total alkaloids
It was performed using the technique of Fazel et al. (2010). 1 mL of extract was dissolved in 1 mL of HCl 2 N (1:1 v/v) and 3 mL of chloroform were added. The precipitate was recovered, and the pH was adjusted to 7 with NaOH 0.1 N. The solution was deposited inside a separation funnel and 5 mL of bromocresol green solution and 5 mL of phosphate buffer at pH 4.7 were added; the mixture was stirred and the alkaloid-green complex of bromocresol was extracted by adding 1 mL of chloroform, subsequently, 7 mL were added recovering the complex for each mL added. The yellow complex was adjusted to 10 mL with chloroform. 200 μL was taken and absorbance was measured at 416 nm in the microplate reader (Metash, UV-5100 UV/VIS, China). Results were expressed in mg atropine equivalents per gram of dry base starch (mg AE / g d.b.s).
Total acetogenins
It was performed using the methodology of Aguilar-Hernández et al. (2022). 250 μL of sample and 4 mL of Kedde reagent (Aguilar-Hernández et al., 2020a) were added in quartz cells. Absorbance kinetics were performed at 505 nm for 30 s with readings every 5 s and the highest absorbance peak was recorded on a spectrophotometer (Metash, UV-5100 UV/VIS, China). Results were expressed as mg annonacin equivalents per gram of dry base starch (mg ANE/g d.b.s).
Statistical analysis
A complete randomized design was used with a 2 x 2 factorial arrangement (extraction methods and starch phases). Four treatments were established where starches (phase 1 and phase 2) obtained from conventional and ultrasound-assisted extraction were evaluated. Results were analyzed with an ANOVA and a comparison of means per Tukey test (p ≤ 0.05) using the Statistical Analysis System software (SAS® V. 9.2).
Results and Discussion
Physicochemical analysis
Lipid content (%)
The lipid content (%) in starch extractions was ACF2 (0.016), AUF2 (0.018), ACF1 (2.4), and AUF1 (4.03). The results obtained showed statistical differences in the extractions (p ≤ 0.05) (Table 1). Krishnan et al. (2021) mention that starches can interact with some compounds of a lipid nature through amylose-lipid complexes. Concerning this characteristic, starches can form binary interactions with some lipid compounds of soursop fruits such as phytosterols and acetogenins, such compounds have been reported by Haykal et al. (2019) and Aguilar-Hernández et al. (2022), respectively. Also, starches with high lipid content have been reported to have lower hydrophilic properties, because lipid compounds form complexes with amylose, limiting the availability of hydroxyl groups to interact with water molecules (Henning et al., 2021).
Amylose content (%)
The content of amylose (%) in the starch extractions was ACF2 (24.48), AUF2 (23.14), ACF1 (11.95), and AUF1 (11.05), with a difference in the results (p ≤ 0.05). For the estimation of amylose, the starches were defatted, eliminating the amylose-lipid complex to form an iodine-amylose complex, which is used for the estimation of amylose (Hoover & Ratnayake, 2001); in this research, the high content of amylose in the starches of the ACF1 and AUF1 extractions could be related to the formation of quaternary interactions with other components such as proteins and phenolic compounds (Krishnan et al., 2021). As reported by Oliveira-Bernardo et al. (2018), this result decreases the water absorption capacity of starch granules. Studies in soursop fruit starches have reported 24.2 and 35.1% amylose in starches extracted by conventional and ultrasound-assisted methods (Ramírez-Balboa et al., 2021). Nwokocha & Williams (2009) reported 19% amylose in starches of Annona squamosa L. and Anonna muricata L. using conventional extraction; they also reported that the ideal range of amylose is 18-30% in native starches. The starches of soursop fruits have a low content of amylose (waxy) and low syneresis (slow retrogradation), so they can be used in the preparation of baked and frozen products (Martínez-Ortiz et al., 2022), in addition to their content of phytochemical compounds they can be considered beneficial for health (Cereceres-Aragón et al., 2019; Gasaly et al., 2020).
Table 1 Physicochemical analysis of soursop fruit starch
| Extraction | Lipids (%) | Amylose (%) |
|---|---|---|
| ACF2 | 0.016 ± 01c | 24.48 ± 0. 56a |
| AUF2 | 0.018±0.01c | 23.14 ± 1.67a |
| ACF1 | 2.4 ± 0.8b | 11.95 ± 0.64b |
| AUF1 | 4.03 ± 0.4a | 11.05 ± 0.3b |
| DMS | 1.24 | 2.49 |
| CV | 29.25 | 5.39 |
The same letters per column do not differ statistically (Tukey, p ≥ 0.05). The values are the means of three repetitions ± standard deviation. LSD: least significant difference. CV: coefficient of variation.
Qualitative analysis of phytochemical compounds
In the starches extracted by the conventional method and ultrasound, the presence (+) of phytosterols, phenols, tannins, alkaloids, flavonoids, and acetogenins was observed (Table 2). Phytochemical compounds found in soursop fruit starch have been reported by León-Fernández et al. (2021) and Aguilar-Hernández et al. (2020a), who conducted a study where they evaluated the phytochemical profile of epicarp, mesocarp, and endocarp, however, there is only one indication of acetogenins present in soursop fruit starches (Ramírez-Balboa et al. 2021). Lipid compounds such as acetogenins can bind starches by covalent bonds with amylose, forming an amylose-lipid complex (Krishnan et al., 2021; León-Fernández et al., 2021); in addition, a high amylose content increases the ability to form interactions with other compounds such as polyphenols (Dueñas et al., 2018; Wang et al., 2020). Likewise, there are reports that starches contain dietary fiber, which has bioactive compounds (Bello-Pérez et al., 2006), these are linked to fiber through hydrophobic interactions and covalent bonds (Saura-Calixto, 2011; Agredano-De la Garza et al., 2021).
Table 2 Qualitative analysis of starch extracts from soursop fruits.
| Phytochemical compound | Extraction | |||
|---|---|---|---|---|
| ACF2 | ACF1 | AUF2 | AUF1 | |
| Steroids | + | + | + | + |
| Terpenes | - | - | - | - |
| Phenols and tannins | + | + | + | + |
| Alkaloids | + | + | + | + |
| Flavonoids | + | + | + | + |
| Acetogenins | + | + | + | + |
ACF1: phase 1 conventional starch, ACF2: phase 1 conventional starch, AUF1: phase 1 ultrasound starch, AUF2: phase 2 ultrasound starch, Presence (+), absence (-).
Quantitative analysis of phytochemical compounds
Total soluble phenols
Phenolic compounds are characterized by having aromatic or benzene rings, with unsaturations and OH groups (Abarca-Vargas & Petricevich, 2021). These compounds interact with starch forming hydrogen bonds, hydrophobic, electrostatic, and ionic interactions (Dueñas et al., 2018), binding to amylose by hydrogen bonds between the hydroxyl groups of polyphenols and the oxygen atoms of the glycosidic bonds of polysaccharides (Fernandes et al., 2014). In this research, the content of total soluble phenols (mg GAE/g of d.b.s) in the starches of the extractions was ACF2 (0.02), AUF2 (0.06), ACF1 (1.44), and AUF1 (2.26). The results showed statistical difference (p ≤ 0.05) (Table 3). The starches from AUF1 presented a high content of phenols concerning the rest of the extractions, this result could be because, in ultrasound-assisted extraction, the cavitation effect generates microjets that break the cell walls allowing the release of intracellular compounds (Aguilar-Hernández et al., 2022). Zárate-Martínez et al. (2021) mention that the shell (epicarp) and seeds (endocarp) of the fruits have a high content of phenolic compounds, in this sense, according to the results obtained in AUF1, compounds of other structures (shell and seeds) may be present.
Total tannins
Tannins are hydrophobic compounds that present hydroxyl groups close to each other in their structure, which strengthens the interaction with carbohydrates such as starch through hydrogen bonds (Amoako & Awika, 2016). Table 3 shows the results of the total tannin content obtained from conventional and ultrasound extractions. The starches obtained in phase 1 by both extraction methods presented statistical differences in the content of total tannins concerning phase 2 (p ≤ 0.05). In the case of phase 1 (ACF1 and AUF1), the result could be attributed to ultrasonic waves that allow the release and solubilization of compounds in the extraction solvent, allowing the interaction of these with starches (Aguilar-Hernández et al., 2019). In the present investigation, it was observed that the starches extracted by ACF1 and AUF1 had lower content of amylose and high content of tannins; in this regard, Dueñas et al. (2018) mention that the formation of the amylose-phenol complex is directly proportional to the amount of amylose that composes it. In this sense, Barros et al. (2012) mention that this result could be due to the presence of tannins with high molecular weight such as ellagitannins (e.g., catechin-gallic acid) (Olivas-Aguirre et al., 2015), which have a greater affinity for amylose.
Total soluble flavonoids
Flavonoids are polyphenols with a hydrophobic character and could bind strongly to starch through covalent interactions with amylose (Barros et al. 2012). These compounds are water-soluble and interact with starches during the extraction process (Sarria-Villa et al., 2021). In this research, the total flavonoid content (mg QE/g of d.b.s) in the extractions was ACF2 (0.022), AUF2 (0.049), ACF1 (0.99) and AUF1 (1.55). The results obtained showed statistical difference (p ≤ 0.05). ACF1 and AUF1 starches were high in flavonoids (Table 3), as reported by Zárate-Martínez et al. (2021), these compounds are present in soursop peel, seed, and pulp. So, the high content of flavonoids in these starches (ACF1 and AUF1) may come from husk and seed. While the low concentrations of flavonoids in the starches AUF2 and ACF2, could be related to starches from the pulp, in addition to being polar they are highly soluble in water according to what was reported by Soto-García & Rosales-Castro (2016). Takahama & Hirota (2018), mention that starches with high flavonoid content can be considered resistant because when consumed they reach the intestinal tract intact and serve as a substrate (prebiotic) for the intestinal microbiota.
Total phytosterols
Phytosterols are secondary metabolites produced by plants, they act as a defense against biotic and abiotic stress (Silva et al., 2016). Phytosterols due to their lipid structure can interact with amylose and amylopectin (Krishnan et al., 2021; Takahama & Hirota, 2018). The content of total phytosterols (mg EC/g d.b.s) was ACF2 (0.86), AUF2 (1.6), ACF1 (2.09), and AUF1 (3.55), with statistical difference (p ≤ 0.05) (Table 3). In the present study, it was observed that the method of extraction of starches influenced the content of phytosterols, where AUF1 presented high content of phytosterols (3.55 mg EC/g of d.b.s) concerning the rest of the extractions, whose results coincide with the high content of lipids present (Table 1), these characteristics decrease their water absorption capacity (Henning et al., 2021). According to these results, starches from AUF1 could present rancidity due to lipid oxidation (Villarroel et al., 2018), however, the consumption of starches high in phytosterols reduces the absorption of cholesterol in the intestine by competition, since they have a chemical structure similar to cholesterol (Fernandes & Cabral, 2007). A diet rich in phytosterols can reduce total cholesterol and LDL cholesterol (harmful to health) (Silva et al., 2016). The estimated daily intake of phytosterols varies between 160 and 500 mg/day, however, its beneficial action is achieved with consumptions of 1500 mg to 2400 mg daily (Laitinen & Gylling, 2012). In the seeds of fruits of anonaceas, phytosterols (β-sitosterols and β-stigmasterols) with anticancer effects have been reported (Haykal et al., 2019; Aguilar-Hernández et al., 2022).
Table 3 Bioactive compounds in soursop fruit starches.
| Extraction | TSP | TST | TSF | TPh | TA | TAc |
|---|---|---|---|---|---|---|
| mg GAE / g of d.b.s | mg QE / g of d.b.s | mg CE / | mg AE / | mg ANE / | ||
| g of d.b.s | g of d.b.s | g of d.b.s | ||||
| ACF1 | 1.44 ± 0.08b | 0.706 ± 0.08b | 0.99 ± 0.02b | 2.09 ± 0.04b | 3.580 ± 0.13b | 1.527 ± 0.003b |
| AUF1 | 2.26 ± 0.07a | 0.993 ± 0.05a | 1.55 ± 0.13a | 3.55 ± 0.07a | 8.429 ± 0.13a | 1.818 ± 0.017a |
| ACF2 | 0.023 ± 0.001c | 0.007 ± 0.0002c | 0.022 ± 0.001c | 0.86 ± 0.07d | 0.012 ± 0.002c | 0.034 ± 0.002c |
| AUF2 | 0.060 ± 0.007c | 0.022 ± 0.002c | 0.049 ± 0.003c | 1.60 ± 0.04c | 0.044 ± 0.005c | 0.039 ± 0.0005c |
| CV | 6.04 | 11.82 | 10.61 | 3.03 | 3.16 | 0.99 |
| LSD | 0.15 | 0.13 | 0.18 | 0.16 | 0.24 | 0.02 |
The same letters per column do not differ statistically (Tukey, p ≤ 0.05). The values are the means of three repetitions ± standard deviation. TSP: total soluble phenols, TST: total soluble tannins, TSF: total soluble flavonoids, TPh: total phytosterols, TA: total alkaloids, TAc: total acetogenins. CV: coefficient of variation. LSD: least significant difference. ACF1: phase 1 conventional starch, AUF1: phase 1 ultrasonic starch, ACF2: phase 2 conventional starch, AUF2: phase 2 ultrasonic starch.
Total alkaloids
Alkaloids are natural compounds containing basic nitrogen atoms (Coria-Téllez et al., 2018), and have been reported in Annona muricata L. mainly isoquinoline, aporfin, and protoberberine type (Riley-Saldaña et al., 2017), these compounds may have in their structure a preformed molecule of a phytosterol (Aguilar-Hernández et al., 2020b; Xiang et al., 2022), allowing the formation of binary amylose-lipid interactions with starches (Krishnan et al., 2021 ). According to De la Cruz-Chacón et al. (2011), these compounds are present in seed endosperm, as well as in the shell, and to a lesser extent in the pulp of soursop fruits. In this research, the total alkaloid content (mg AE/g of d.b.s) was ACF2 (0.012), AUF2 (0.044), ACF1 (3.58), and AUF1 (8.42). The results obtained showed statistical differences (p ≤ 0.05) (Table 3). In this sense, the high content of alkaloids in AUF1 starches can be attributed to ultrasound-assisted extraction, because the cavitation effect breaks cell membranes allowing the release of phytochemical compounds (Aguilar-Hernández et al., 2022). Alkaloids from soursop fruits may help reduce blood pressure by blocking calcium ion channels instead of activating endothelium- and nitric oxide-dependent mechanisms (Zubaidi et al., 2023). Isoquinoline, coreximina, and anomurin have been shown to have a transient depressive effect on blood pressure (Nwokocha et al., 2012). The starches extracted from soursop fruits from this research could have the same effect as reported by these researchers.
Total acetogenins
Acetogenins are secondary metabolites that contain tetrahydrofuran rings and a lipophilic side chain that allows the formation of binary complexes with amylose (Bachan et al., 2013; Krishnan et al., 2021). In starches extracted from soursop fruits, the content of total acetogenins (mg ANE/g of d.b.s) was ACF2 (0.034), AUF2 (0.039), ACF1 (1.527) and AUF1 (1.818). The results obtained showed statistical differences (p ≤ 0.05) (Table 3). In this regard, high content of ACGs was observed in AUF1 starches, this could be due to the presence of seed and husk starches, as well as ultrasound, whose results coincide with what was reported by Aguilar-Hernández et al. (2020a), who reported that soursop seeds and shells contain high content of acetogenins and ultrasound-assisted extraction of these compounds increases yield. On the other hand, ACF1 and ACF2 starches presented low content of acetogenins, which could be attributed to the fact that the conventional extraction method has lower extraction efficiency compared to the ultrasound-assisted method (López-Romero et al., 2022; Aguilar-Hernández et al., 2022). The starches of soursop fruits, due to their acetogenin content, can be used in the development of new innovative products with nutraceutical characteristics, in such a way that they are healthy for the consumer by reducing the problems caused by degenerative diseases. Studies indicate that the consumption of acetogenins has anticancer (Mutakin et al., 2022), and antidiabetic (Son et al., 2021) effects, in addition, it has been proven that these compounds have antimicrobial activity against E. faecalis, S. paratyphi, E. coli and L. monocytogenes, which are the most prevalent harmful bacteria found in food (Siderakou et al., 2021).
