Genome-wide diversity analysis suggests divergence among Upper Guinea and the Dahomey Gap populations of the Sisrè berry (Syn: miracle fruit) plant (Synsepalum dulcificum [Schumach. & Thonn.] Daniell) in West Africa.

Although Synsepalum dulcificum is viewed as one of the most economically promising orphan tree crops worldwide, its genetic improvement and sustainable conservation are hindered by a lack of understanding of its evolutionary history and current population structure. Here, we report for the first time the application of genome-wide single nucleotide polymorphism genotyping to a diverse panel of S. dulcificum accessions to depict the genetic diversity and population structure of the species in the Dahomey Gap (DG) and Upper Guinea (UG) regions to infer its evolutionary history. Our findings suggest low overall genetic diversity but strong population divergence within the species. Neighbor-joining analysis detected two genetic groups in the UG and DG regions, while STRUCTURE distinguished three genetic groups, corresponding to the UG, Western DG, and Central DG regions. Application of Monmonier's algorithm revealed the existence of a barrier disrupting connectivity between the UG and DG groups. The Western DG group consistently exhibited the highest levels of nucleotide and haplotype diversities, while that of the Central DG exhibited the lowest. Analyses of Tajima's D, Fu's Fs, and Achaz Y* statistics suggest that while both UG and Central DG groups likely experienced recent expansions, the Western DG group is at equilibrium. These findings suggest a geographical structuring of genetic variation which supports the conclusion of differential evolutionary histories among West African groups of S. dulcificum. These results provide foundational insights to guide informed breeding population development and design sustainable conservation strategies for this species.

Quantification of lectins in Synsepalum dulcificum and comparison with reference foods.

A healthy life means a balance between physical activity and a diet rich in fruits and vegetables, however, some plant-based foods can have certain adverse effects due to the presence of anti-nutritional factors, such as lectins, capable of binding molecules and preventing their normal assimilation. The level of lectins in Synsepalum dulcificum fruit was determined by hemagglutination assays in human blood, and its comparison with foods characterized as having high and low lectin content. The relative hemagglutinating activity of berries from Synsepalum dulcificum compared to our positive high lectin content food reference (Pinto bean) corresponds to 3.13-6.25%, representing safe levels for nutritional food.

Antihyperglycemic and hepatoprotective properties of miracle fruit (Synsepalum dulcificum) compared to aspartame in alloxan-induced diabetic mice.

OBJECTIVE: This study was undertaken to investigate the antihyperglycemic potential of miracle fruit (MF) as well as its hepatic safety as compared to aspartame in alloxan-induced diabetic mice. METHODS: MF extracts were prepared and screened for their phytochemical composition using high-performance liquid chromatography (HPLC). Total phenolic, flavonoid and tannin contents and antioxidant potential were also determined. Additionally, MF was evaluated for its sensory attributes. For in vivo work, MF ethanol extract at high (MFH: 500 mg/kg body weight [BW]) and low (MFL: 250 mg/kg BW) doses as well as aspartame were injected intraperitoneally into alloxan-induced diabetic mice. Blood glucose levels were determined following acute and subchronic treatment. At the end of the study, animals were sacrificed, serum was collected for biochemical analysis and liver tissues were obtained for histopathological examination. RESULTS: MF ethanol extract contained more flavonoids and tannins, and had higher 1,1-diphenyl-1-picrylhydrazyl radical-scavenging activity (79.61%) compared to MF aqueous extract (P < 0.05). HPLC analysis of MF ethanol extract also revealed the presence of 10 antioxidants with quercetin comprising the major polyphenol. Additionally, sensory analysis of MF showed that its intake is effective in masking undesirable sourness. Subchronic administration of MFH proved amelioration of hyperglycemia in mice as compared to aspartame. Moreover, aspartame treatment significantly elevated (P < 0.05) the level of alanine aminotransferase and had destructive effects on the liver histopathology; however, hepatic architecture was restored by low and high doses of MF. CONCLUSION: MF is an effective antihyperglycemic with hepatoprotective properties that can be used as a healthier alternative sweetening agent in place of aspartame for sour beverages.

Safety assessment of miraculin using in silico and in vitro digestibility analyses.

Miraculin is a glycoprotein with the ability to make sour substances taste sweet. The safety of miraculin has been evaluated using an approach proposed by the Food and Agriculture Organization of the United Nations and the World Health Organization for assessing the safety of novel proteins. Miraculin was shown to be fully and rapidly digested by pepsin in an in vitro digestibility assay. The proteomic analysis of miraculin's pepsin digests further corroborated that it is highly unlikely that any of the protein will remain intact within the gastrointestinal tract for potential absorption. The potential allergenicity and toxigenicity of miraculin, investigated using in silico bioinformatic analyses, demonstrated that miraculin does not represent a risk of allergy or toxicity to humans with low potential for cross-reactivity with other allergens. The results of a sensory study, characterizing the taste receptor activity of miraculin, showed that the taste-modifying effect of miraculin at the concentration intended for product development has a rapid onset and disappearance with no desensitizing impact on the receptor. Overall, the results of this study demonstrate that the use of miraculin to impact the sensory qualities of orally administered products with a bitter/sour taste profile is not associated with any safety concerns.

Microencapsulation of Lactococcus lactis Gh1 with Gum Arabic and Synsepalum dulcificum via Spray Drying for Potential Inclusion in Functional Yogurt.

There has been an explosion of probiotic incorporated based product. However, many reports indicated that most of the probiotics have failed to survive in high quantity, which has limited their effectiveness in most functional foods. Thus, to overcome this problem, microencapsulation is considered to be a promising process. In this study, Lactococcus lactis Gh1 was encapsulated via spray-drying with gum Arabic together with Synsepalum dulcificum or commonly known as miracle fruit. It was observed that after spray-drying, high viability (~108 CFU/mL) powders containing L. lactis in combination with S. dulcificum were developed, which was then formulated into yogurt. The tolerance of encapsulated bacterial cells in simulated gastric juice at pH 1.5 was tested in an in-vitro model and the result showed that after 2 h, cell viability remained high at 1.11 × 106 CFU/mL. Incubation of encapsulated cells in the presence of 0.6% (w/v) bile salts showed it was able to survive (~104 CFU/mL) after 2 h. Microencapsulated L. lactis retained a higher viability, at ~107 CFU/mL, when incorporated into yogurt compared to non-microencapsulated cells ~105 CFU/mL. The fortification of microencapsulated and non-microencapsulated L. lactis in yogurts influenced the viable cell counts of yogurt starter cultures, Lactobacillus delbrueckii subs. bulgaricus and Streptococcus thermophilus.

A combination of approaches evidenced seed storage behaviour in the miracle berry Synsepalum dulcificum (Schumach. et Thonn.) Daniell.

BACKGROUND: Knowledge on seed storage behaviour is crucial for planning conservation strategies of plant genetic resources particularly in economically promising but endangered species like Synsepalum dulcificum, viewed as recalcitrant-seeded species albeit sound evidence was lacking. In this study, we combined an experimental approach based on critical moisture content and storage environment analysis, and the seed-coat ratio-seed dry mass (SCR-SM) model to clarify the seed storage behaviour in the species. Seed moisture content at shedding was determined and effects of dehydration and cold storage on seed viability, germination and subsequent seedling vigour were analysed. The probability for dessication-senstivity [P(D-S)] was also determined. RESULTS: Our findings indicated that S. dulcificum seed moisture content at shedding was 36.60% with nearly 100% viability. Seed dehydration below 20% moisture content induced a total loss of viability whereas low temperature storage (at 10 °C or 4 °C reduced shelf life to a maximum of 7 days. More importantly, S. dulcificum seed storage at 0 °C was highly detrimental and resulted in a total loss of viability whatever the storage duration. Only a storage at 25 °C helped expand the shelf life to 28 days. However, at 28 days storage the viability was extremely low with almost no germination. The probability for dessication-senstivity P(D-S) in the species is largely greater than 0.5. Seed dehydration and storage environment highly affected subsequent germination rate and seedling vigour. While dehydration improved seedling performance storage at low temperature rather inhibited seedling growth. CONCLUSION: Taken together, these findings are the first to set evidence of recalcitrance in S. dulcificum and serve hands-on information for practical handling of the seeds and designing sustainable conservation practices for adequate future breeding programme in the species.

Beneficial effects of an investigational wristband containing Synsepalum dulcificum (miracle fruit) seed oil on the performance of hand and finger motor skills in healthy subjects: A randomized controlled preliminary study.

Miracle fruit (Synsepalum dulcificum) seed oil (MFSO) contains phytochemicals and nutrients reported to affect musculoskeletal performance. The purpose of this study was to assess the safety and efficacy of a compression wristband containing MFSO on its ability to measurably improve the hand and finger motor skills of participants. Healthy right-handed participants (n = 38) were randomized in this double-blind, placebo-controlled study of MFSO and vehicle wristbands. Subjects wore the wristband on their left hand 4-6 weeks and then only on their right hand 2-4 weeks; the contralateral untreated hand served as an additional control. Twelve hand/finger motor skills were measured using quantitative bio-instrumentation tests, and subject self-assessment questionnaires were conducted. With each hand, in 9/12 tests, the MFSO group showed a clinically meaningful average improvement compared with an average worsening in the vehicle group. Statistical superiority to the control treatment group was exhibited in 9/12 tests for each hand (p < .01). After discontinuing the MFSO wristband on the left hand, test values regressed toward baseline levels. Subjects favored the MFSO wristband over the control, rating it as effective in improving their motor skills. Use of the MFSO wristband may improve an individual's manual dexterity skills and ability to maintain this performance.

Miracle Fruit (Synsepalum dulcificum) Exhibits as a Novel Anti-Hyperuricaemia Agent.

Miracle fruit (Synsepalum dulcificum) belongs to the Sapotaceae family. It can change flavors on taste buds, transforming acidic tastes to sweet. We evaluated various miracle fruit extracts, including water, butanol, ethyl acetate (EA), and hexane fractions, to determine its antioxidant effects. These extracts isolated from miracle fruit exerted potential for reduction of uric acid and inhibited xanthine oxidase activity in vitro and in monosodiumurate (MSU)-treated RAW264.7 macrophages. Moreover, we also found that the butanol extracts of miracle fruit attenuated oxonic acid potassium salt-induced hyperuricaemia in ICR mice by lowering serum uric acid levels and activating hepatic xanthine oxidase. These effects were equal to those of allopurinol, suggesting that the butanol extract of miracle fruit could be developed as a novel anti-hyperuricaemia agent or health food.

Optimization of conditions for the single step IMAC purification of miraculin from Synsepalum dulcificum.

In this study, the methods for extraction and purification of miraculin from Synsepalum dulcificum were investigated. For extraction, the effect of different extraction buffers (phosphate buffer saline, Tris-HCl and NaCl) on the extraction efficiency of total protein was evaluated. Immobilized metal ion affinity chromatography (IMAC) with nickel-NTA was used for the purification of the extracted protein, where the influence of binding buffer pH, crude extract pH and imidazole concentration in elution buffer upon the purification performance was explored. The total amount of protein extracted from miracle fruit was found to be 4 times higher using 0.5M NaCl as compared to Tris-HCl and phosphate buffer saline. On the other hand, the use of Tris-HCl as binding buffer gave higher purification performance than sodium phosphate and citrate-phosphate buffers in IMAC system. The optimum purification condition of miraculin using IMAC was achieved with crude extract at pH 7, Tris-HCl binding buffer at pH 7 and the use of 300 mM imidazole as elution buffer, which gave the overall yield of 80.3% and purity of 97.5%. IMAC with nickel-NTA was successfully used as a single step process for the purification of miraculin from crude extract of S. dulcificum.

Inhibitory effect of aqueous extracts from Miracle Fruit leaves on mutation and oxidative damage.

This study investigated the inhibitory effects of aqueous extracts from Miracle Fruit leaves (AML) on mutation and oxidative damage. The results showed that AML in the range of 1-5mg/plate inhibited the mutagenicity of 2-aminoanthracene (2-AA), an indirect mutagen, and 4-nitroquinoline-N-oxide (4-NQO), a direct mutagen toward Salmonella typhimurium TA 98 and TA 100. On the other hand, AML in the range of 0.05-0.2mg/ml showed radical scavenging, reducing activities, liposome protection as well as decreased tert-butyl hydroperoxide (t-BHP) induced oxidative cytotoxicity in HepG2 cells. High performance liquid chromatography (HPLC) analysis suggested that the active phenolic constituents in AML are p-hydroxybenzoic acid, vanillic acid, syringic acid, trans-p-coumaric acid and veratric acid. These active phenolic components may contribute to the biological protection effects of AML in different models. The data suggest that AML exhibiting biological activities can be applied to antimutation as well as anti-oxidative damage.

The clinical effects of Synsepalum dulcificum: a review.

Synsepalum dulcificum or the "miracle fruit" is well known for its taste-modifying ability. The aim of this review was to assess the published medically beneficial as well as potential characteristics of this fruit. A search in three databases, including PubMed, ScienceDirect, and Google Scholar, was made with appropriate keywords. The resulting articles were screened in different stages based on the title, abstract, and content. A total of nine articles were included in this review. This review summarized the findings of previously published studies on the effects of miracle fruit. The main studied characteristic of the fruit was its effect on the taste receptors, resulting in the sweet sensation when substances with acidic content were ingested. This effect was shown to be related to a glycoprotein called "miraculin." Other beneficial characteristics of this fruit were its antioxidant and anticancer abilities that are due to the various amides existing in the miracle fruit. Apart from the above, the other observed effect of this fruit was its antidiabetic effect that was tested in rats. Further studies should be conducted to establish the findings. The miracle fruit can be a healthy additive due to its unique characteristics, including sour taste sensation modification as well as its antioxidant and antidiabetic effects.

Antioxidant-rich phytochemicals in miracle berry (Synsepalum dulcificum) and antioxidant activity of its extracts.

Miracle berry is known for its unique characteristic of modifying sour flavours to sweet. Twelve phenolics were identified and quantified in the miracle berry flesh at a level from 0.3 for kaempferol to 17.8 mg/100g FW for epicatechin. Lutein and α-tocopherol were also quantified at a level of 0.4 and 5.8 mg/100g FW, respectively. The TP and TF contents were 1448.3 GA and 9.9 QR mg Equiv/100g FW for the flesh, respectively, compared with 306.7 GA and 3.8 mg QR mg Equiv/100g FW of the seeds. The free radical scavenging and reducing percentage of the flesh extract was 96.3% and 32.5% in DPPH and ABTS assays, respectively. Additionally, the flesh extract had a high FRAP of 22.9 mmol/100g. It significantly inhibited the oxidation of PUFA in fish oil as well. Thus, miracle berry could also serve as an antioxidant-rich fruit to provide health promoting function.

Differential sweetness of commercial sour liquids elicited by miracle fruit in healthy young adults.

Miracle fruit (Synsepalum dulcificum) contains the glycoprotein miraculin which turns a sour taste into a sweet one. Chemical analyses and sensory evaluation experiments were conducted to examine the sweetening effect of miracle fruit with regard to five different commercial sour liquids which were diluted until they were subjectively equally sour. HPLC-based analyses revealed that (1) the predominating acids in two and three of the liquids were citric acid and acetic acid, respectively and (2) all five liquids contained fructose and glucose. Healthy young adults (eight males and 10 females) in the sensory evaluation experiments were asked to chew a miracle fruit and apply their saliva to the oral mucosae. They were asked to score the sweetness elicited by the five liquids relative to a sucrose standard at 0, 15, 25 and 35 min thereafter. The citric acid-based liquids were perceived as being sweeter than the acetic acid-based liquids at all timepoints. Thus, commercial sour liquids that mainly contain citric acid are more effective than acetic acid-based liquids in eliciting a perception of sweetness after the miracle fruit application, while the sugars in the liquids seemed to play a minimal role as determinants of sweetness.

Contents of phenolics and flavonoids and antioxidant activities in skin, pulp, and seeds of miracle fruit.

Miracle Fruit (Synsepalum dulificum) has been studied because of its unique taste modifying properties. This study investigated contents of phenolics, flavonoids, and antioxidant activities in skin, pulp, and seeds of Miracle Fruit. The free phenolic content in skin was almost 3 times of that in pulp and 4 times of that in seeds. Skin contributed 43.96% of free phenolic compounds with 15.91% of freeze-dried solids due to its high phenolic content. As the trend observed for phenolic content, the free flavonoid content in the skin was tremendously higher than that in the seed and pulp. The skin contributed about 52% of total flavonoid with 15.91% of dried solids. On other hand, the differences in the bound phenolic contents were not so distinct among the 3 components. The free antioxidant activities in skin and pulp were comparable, and were significantly higher than that in seeds. Although the antioxidant activities in seeds was considerably lower than that in skin, 49.45% free antioxidant activity, 76.41% bound antioxidant activity, and 58.56% of total antioxidant activity were contributed by seeds due to about 66% of solid of total solids. In general, the results of antioxidant activities using sequential methods were higher than that using direct method. This study suggests that Miracle Fruit is a good source not only for flavor and color, and also antioxidant activity for functional food applications.

Isolation of a 2-oxetanone from the fruits of Synsepalum dulcificum.

Ficumone, a 2-oxetanone isolated from the fruits of Synsepalum dulcificum, has been characterised as (R*)-4-hydroxy-2-oxetanone by means of spectroscopic methods.

Single-step purification of native miraculin using immobilized metal-affinity chromatography.

Miraculin is a taste-modifying protein that can be isolated from miracle fruit ( Richadella dulcifica ), a shrub native to West Africa. It is able to turn a sour taste into a sweet taste. The commercial exploitation of this sweetness-modifying protein is underway, and a fast and efficient purification method to extract the protein is needed. We succeeded in purifying miraculin from miracle fruit in a single-step purification using immobilized metal-affinity chromatography (IMAC). The purified miraculin exhibited high purity (>95%) in reverse-phase high-performance liquid chromatography. We also demonstrated the necessity of its structure for binding to the nickel-IMAC column.

Improvement of insulin resistance by miracle fruit (Synsepalum dulcificum) in fructose-rich chow-fed rats.

In an attempt to probe a new target to improve insulin resistance, miracle fruit (Synsepalum dulcificum) was employed to investigate the effect on insulin resistance induced by fructose-rich chow in rats. Single oral administration of the powder of this miracle fruit decreased the plasma glucose in a dose-dependent manner for 150 min in rats fed fructose-rich chow for 4 weeks. Insulin action on the glucose disposal rate was measured using the glucose-insulin index, the value of the areas under the curve of glucose and insulin during the intraperitoneal glucose tolerance test. Oral administration of miracle fruit (0.2 mg/kg) to fructose-rich chow fed rats, three times daily for 3 days, reversed the raised value of the glucose-insulin index, indicating that miracle fruit has the ability to improve insulin sensitivity. The plasma glucose lowering action of tolbutamide, induced by secretion of endogenous insulin, is widely used to characterize the formation of insulin resistance. The time for the loss of the plasma glucose lowering response to tolbutamide (10.0 mg/kg, i.p.) in fructose-rich chow fed rats was markedly delayed after treatment with miracle fruit compared with the vehicle-treated group. Thus providing supportive data that oral administration of miracle fruit could delay the development of insulin resistance in rats. Also, the in vivo insulin sensitivity was markedly raised by miracle fruit. In conclusion, the results suggest that miracle fruit may be used as an adjuvant for treating diabetic patients with insulin resistance because this fruit has the ability to improve insulin sensitivity.