Previous Article in Journal
High Diagnostic Performance of the Indicator Plaster Neuropad for the Detection of Established Diabetic Autonomic Neuropathy
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
Diabetology
Volume 6
Issue 6
10.3390/diabetology6060056
diabetology-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Article

Effect of Chronic Administration of Justicia secunda Vahl in Mice Diabetized with Streptozotocin

by
Tomás Fregoso-Aguilar
1,†,
Perla Xóchitl Escamilla-Ramírez
1,†,
Dulce E. Nicolás-Álvarez
1,
Jorge Alberto Mendoza-Pérez
2,
Eduardo Osiris Madrigal-Santillán
3,
Judith Margarita Tirado-Lule
4,
Elda Victoria Rodríguez-Negrete
3,
Eduardo Madrigal-Bujaidar
5,
Isela Álvarez-González
5,
Gabriela Ibáñez-Cervantes
6,
Ángel Morales-González
4,* and
José A. Morales-Gonzalez
3,*
1
Escuela Nacional de Ciencias Biológicas, Instituto Politécnico Nacional, Departamento de Fisiología, Av. Wilfrido Massieu s/n, Col. Nueva Industrial Vallejo, Alcaldía Gustavo A. Madero, Ciudad de México C.P. 07700, Mexico
2
Escuela Nacional de Ciencias Biológicas, Instituto Politécnico Nacional, Departamento de Ingeniería en Sistemas Ambientales, Av. Wilfrido Massieu s/n, Col. Nueva Industrial Vallejo, Alcaldía Gustavo A. Madero, Ciudad de México C.P. 07700, Mexico
3
Laboratorio de Medicina de Conservación, Escuela Superior de Medicina, Instituto Politécnico Nacional, Plan de San Luis y Díaz Mirón, Col. Casco de Santo Tomás, Alcaldía Miguel Hidalgo, Ciudad de México C.P. 11340, Mexico
4
Escuela Superior de Cómputo, Instituto Politécnico Nacional, Unidad Profesional “A. López Mateos”, Ciudad de México C.P. 07738, Mexico
5
Laboratorio de Genética, Escuela Nacional de Ciencias Biológicas, Instituto Politécnico Nacional, Av. Wilfrido Massieu s/n, Zacatenco, Gustavo A. Madero, Mexico City C.P. 07738, Mexico
6
Sección de Estudios de Posgrado e Investigación, Escuela Superior de Medicina, Instituto Politécnico Nacional, Plan de San Luis y Díaz Mirón, Col. Casco de Santo Tomás, Alcaldía Miguel Hidalgo, Ciudad de México, C.P. 11340, Mexico
*
Authors to whom correspondence should be addressed.
†
These authors contributed equally to this work.
Diabetology 2025, 6(6), 56; https://doi.org/10.3390/diabetology6060056
Submission received: 24 April 2025 / Revised: 31 May 2025 / Accepted: 11 June 2025 / Published: 18 June 2025
Browse Figures
Versions Notes

Abstract

:
Certain pharmacological properties of the methanolic extract of Justicia secunda Vahl leaves (Acanthaceae) were evaluated in Streptozotocin (STZ)-treated albino mice to confirm whether it could be considered an alternative candidate for the treatment of diabetes. Using qualitative phytochemistry, alkaloids, flavonoids, and tannins were detected. In an in vitro DPPH antioxidant activity test, high extract concentrations inhibited the radical by 90% during the first minutes of the reaction. The extract presented a slight genoprotective effect on mouse peripheral blood during the last days of the micronucleus test. Oral administration of the extract at a high dose every two days for 6 weeks caused a hypoglycemic effect in STZ-treated mice, protection against weight loss, and decreased blood triglyceride levels from week 3 of treatment. These effects could be mediated by the antioxidant activity of the detected metabolites and, perhaps, by an inhibitory effect on intestinal α-glucosidase. This renders J. secunda a good candidate for the long-term alternative treatment of diabetes without abandoning allopathic therapy.

Graphical Abstract

1. Introduction

The use of medicinal plants is widespread in Mexico and other Latin American countries, where many sectors of the population, particularly the poorest, use them as alternative treatments for various chronic degenerative diseases, such as diabetes [1,2]. For this purpose, different parts of the plants can be used, such as the leaves, flowers, bark, roots, or even the entire plant [3]. The use of plants is also increasing in developed countries; on the European continent alone, around 100 million people utilize Traditional Medicine, and an equal or greater number has been estimated for other regions of the world [4].
Worldwide, approximately 70,000 species of plants are used in Traditional Medicine. In this context, during the period from 1991 to 2003, Mexico was in fourth place among countries exporting medicinal and aromatic plants, at 37,600 tons, and it was estimated that in 2022, it ranked second in this regard [5,6,7]. For example, in one study, it was found that the most commonly used plant species in areas such as the Guadalajara City metropolitan zone (Mexico) were Arnica (Heterotheca inoloides), Cuachalalate (Amphypterigium adstringens), Tila (Tila Mexicana), Gordolobo (Gnaphalium spp.), Salvia (Salvia officinalis), and Cola de Caballo (Equisetum hyemale); in addition, 534/1000 inhabitants use nopal (Opuntia ficus-indica), chaya (Cnidoscolus aconitifolius), and matarique (Psacalium peltatum) [8]. This is because people in such areas view medicinal plants from the perspectives of health, financial resources, cultural identity, and the perception of security [9,10,11]. As previously mentioned, many of these species are employed for the treatment of chronic degenerative diseases such as diabetes, and in Mexico, the use of several entities for this purpose has been reported [7,12,13], such as cow’s hoof (Bahuinia forficata) [14,15], Mexican oregano (Lippia graveolens) for its antioxidant properties [16], nopal (Opuntia spp.) [17,18], Neem (Azadirachta indica) [19,20], and many more.
However, many remedies prepared with medicinal plants, such as infusions or teas, are administered without the supervision of medical experts, and in many cases, there is insufficient scientific data to support the use and therapeutic effectiveness of these infusions for the conditions they are intended to treat. In this sense, few studies have focused on elucidating the adverse effects of medicinal preparations used to treat various pathologies. In the majority of studies in this area the authors have reported that, at the doses used (50 mg—2000 mg) in rat, mouse, or rabbit models, no toxic effects are present [21,22,23]. Only certain works have emphasized that some medicinal plants do entail side effects—for example, in the liver [24,25]—while others have found abortive effects in rabbits and mice [26,27].
Justicia secunda Vahl (Acanthaceae) is a species of plant that can flourish in humid, tropical, and temperate climates (Central America and South America) [28,29,30]; in parts of the central and southern regions of Mexico, the use of this plant (and others) as an alternative treatment (herbal) for diabetes is becoming very common; however, apart from a few studies, there is scarce evidence in the scientific literature supporting its therapeutic benefits against that disease. The ethnomedical use of this plant has been reported in the Huasteca Potosina region and the Yucatan Peninsula [31], as well as that of Justicia spicigera Schltdl in the Mexican states of Michoacán, Tabasco, Nayarit, Jalisco, Chiapas, Morelos, Tlaxcala, Veracruz, and Yucatán, where infusions are prepared to treat inflammation, anemia, leukemia, tuberculosis, diarrhea, hemorrhoids, rheumatism, parasitosis, arthritis, and bone and eye diseases. Additionally, evidence of the anticonvulsant, antidepressant, anxiolytic, antinociceptive, and antidiabetic properties of some of its active metabolites is beginning to emerge [32,33]; still, other works have reported its use in women’s reproductive health [34].
Although we are not currently in a declared pandemic, as in the case of COVID-19, it is clear that diabetes represents a global health problem. In May 2025, the number of people diagnosed with this disease will be 589 million, and it has been estimated that this number will reach 852 million people by 2050, representing a 45% increase in reported cases [35]. In Mexico, both type 1 and type 2 diabetes greatly increased in 2019 and 2020 [36,37]. In 2024, Mexico ranked eighth worldwide with 13.6 million adults (20–79 years) with diabetes, while by 2050, it is expected to reach ninth place with 19.9 million diabetics. In addition, it is estimated that 41.3 million Mexicans are not diagnosed [35]. All of this leads to an urgent need to broaden the perspective for implementing alternative treatments for diabetes in developing countries. In this respect, the aim of the present research is to establish whether the methanolic extract of Justicia secunda Vahl, administered chronically in a chemical model of diabetes in mice, exerts antioxidant, genoprotective, and hypoglycemic effects. This will provide scientific data in favor of its use as part of a possible alternative therapy against diabetes, which could be extrapolated to the field of treatment of this pathology in humans in the near future.

2. Materials and Methods

2.1. Acquisition of the Plant Species and Processing in the Laboratory

Justicia secunda Vahl leaves were obtained at an accredited commercial establishment located in Cuemánco in the Xochimilco municipality of Mexico City. These were taken to the Hormones and Behavior Laboratory of the Department of Physiology of the National School of Biological Sciences for complete environmental drying for 1 week. The material was then crushed and macerated in methanol for 1 week. The macerate was subjected to reduced pressure distillation (Rotavaporator Prendo© Model 1750; Prendo, Puebla, México) to remove the methanol; the distillate was then left to air-dry for 1 week to obtain the dry, crude combined extract of the leaves and stems [15].

2.2. Chemicals Used in the Experiments

The chemicals 2,2-Diphenyl-1-picrylhydrazyl (DPPH) and streptozocin (STZ) were purchased from Sigma-Aldrich (Burlington, MA, USA). Giemsa dye was purchased from Hycel (Mexico City, Mexico). Methanol was purchased from Golden Bell Co. (Mexico City, Mexico).

2.3. Animals in Laboratory Settings

As described in a previous study [15], male Swiss albino mice (weighing 25–30 g each) of the NIH strain were used. They were obtained from the official supplier of the National School of Biological Sciences (ENCB) of the National Polytechnic Institute (IPN) and housed in the animal chamber of the ENCB Department of Physiology, Zacatenco Campus, for acclimatization in communal acrylic cages (48 cm long × 22 cm wide × 20 cm high), with water and food ad libitum and a light–dark cycle of 12:00 (lights on at 08:00), as well as a room temperature of 22 ± 2 °C under standard humidity conditions. The animals were handled and subjected to experimentation according to Mexican standards (NOM-033-ZOO-1995, NOM-062-ZOO-1999, and NOM-087-ECOL-1995 [38,39,40]). In addition, all experiments were approved by the Research Ethics Committee of the National School of Biological Sciences (ZOO-005-2022).

2.4. Qualitative Phytochemical Analysis

Samples of the crude extract of the leaves of J. secunda were subjected to various qualitative chemical reactions, as described in other works [41], to determine secondary metabolites, mainly of a polar nature. These were identified based on changes in coloration, the formation of precipitates, foaming, etc.

2.5. Spectroscopic Analysis

Fractions of the dry extract of J. secunda were taken and stored in vials for a series of general spectroscopic analyses. The samples were analyzed at the Center of Nanosciences and Micro and Nanotechnologies of the National Polytechnic Institute (Mexico City, México). The Fourier transform infrared analysis (FT-IR) technique was applied as described in a previous study [15].

2.6. Evaluation of In Vitro Antioxidant Activity

Four concentrations of the crude extract of J. secunda (50, 25, 21.5, and 6.25 mg/mL) were prepared in methanol to determine its antioxidant activity, utilizing the in vitro DPPH (2,2-Diphenyl-1-picrylhydrazyl) method [15,42,43,44]. The absorbance value was recorded at 0 (without the extract sample), 1, 5, 15, 20, 30, and 60 min.

2.7. Assessment of Genoprotective Activity

Three groups of male Swiss albino mice were formed and housed in three cages (35 cm long × 25 cm wide × 12 cm high), each containing six mice. Each group received one of the following treatments: (i) control: administration of vehicle (mineral oil intragastrically [i.g.]) every other day for 1 week, taking a blood smear every other day for 2 weeks; (ii) anthracene (10 mg/kg, i.g.); and (iii) anthracene + J. secunda (500 mg/kg, i.g.) (as described in other works) [15,44].

2.8. Assessment of Hypoglycemic Activity

Four groups of male Swiss albino mice were formed and housed in four cages, each containing six mice. Each group received one of the following treatments, as described in other works [15,44]: (i) Control mice (saline, 0.9%; vol = weight/1000): each week, blood glucose was measured with a commercial device (Optium FreeStyle©; Abbott, Boston, MA, USA), making a small cut in the distal part of the mouse’s tail to drain one drop of blood (prior fasting <12 h) over 36 days. (ii) Diabetic mice were administered a single intraperitoneal dose (i.p.) of Streptozotocin (STZ; 120 mg/kg, dissolved in citrate buffer). (iii) Diabetic + J. secunda extract (500 mg/kg, i.g.). (iv) Diabetic + J. secunda (500 mg/kg) + acarbose (300 mg/kg). Each week, body weight was measured using a granatary balance and triglyceride levels were measured with a commercial device (Accutrent; Roche Diagnostics GMBH; Mannheim, Germany ).

2.9. Statistical Analysis

The data obtained in the animal experiments were analyzed with the SigmaStat ver. 12.0 statistical software, utilizing the repeated measures two-way ANOVA and the Student–Newman–Keuls post hoc tests to determine significant differences between the groups [15,44].

3. Results

3.1. Qualitative Phytochemical Analysis

According to the methodology described in the corresponding section, fresh Justicia secunda leaves were subjected to around 30 chemical reactions and divided into acid, ethanolic, and aqueous extracts. Table 1 lists the secondary metabolites detected with the battery of chemical reactions used for this purpose.

3.2. Spectroscopic Analysis

Figure 1 depicts the spectrum of J. secunda leaf extract obtained by means of infrared analysis with Fourier transform (FTIR). For a wavelength of 600 −1000 cm−1, a =C-H bond vibrations were located; at a wavelength close to 1100 cm−1, the carbonyl functional group was located. Methyl groups were located at a wavelength of 1375 cm−1, and methylene groups were located at a wavelength of 1460 cm−1. In addition, -C-C-H bonds around 2900 cm−1 were found and, at 3300 cm−1, hydroxyl bonds were detected.

3.3. Evaluation of In Vitro Antioxidant Activity

Figure 2 summarizes the antioxidant activity of the four concentrations of Justicia secunda tested using the in vitro DPPH method, with which the percentage of inhibition of DPPH when using the extract was evaluated. Ascorbic acid (2%) was employed as a reference standard, and it reached its maximal inhibition in the presence of DPPH (90.84%) when 10 min of the reaction had elapsed.
In total, the 50 mg/mL and 25 mg/mL concentrations of the extract reached a maximal percentage of inhibition greater than 91% for the DPPH radical during the reaction, highlighting the fact that both concentrations exceeded 90% inhibition minutes prior to when ascorbic acid reached this level. However, the 12.5 mg/mL concentration exhibited a delayed and lower inhibitory capacity than the previous concentrations, only inhibiting 50% of the DPPH radical at 30 min after the start of the reaction. Lastly, the 6.25 mg/mL concentration did not demonstrate good inhibitory activity, in that its value at the end of the reaction was 47.2%.

3.4. Assessment of Genoprotective Activity

Figure 3 presents the number of micronuclei in mouse peripheral blood over 1 week of treatment with anthracene (10 mg/kg) and the methanolic extract of J. secunda leaves (500 mg/kg), as well as the number in the following week without treatment. The group of mice treated with anthracene demonstrated a greater number of erythrocytes with micronuclei than the control group, and this value increased over the last 2 days (when anthracene was no longer administered). On the other hand, the group of mice treated with the J. secunda extract exhibited a number of micronuclei similar to that of the group treated with anthracene, and the presence of micronuclei significantly decreased only during the last 2 days of week 2; this group also being untreated during this time (p < 0.05; two-way repeated measures ANOVA).
Table 2 shows the quantitative data of the number of micronuclei found in 10 microscope fields (100×) of the three groups of mice from which blood smears were taken.

3.5. Assessment of Hypoglycemic Activity

When a person suffers from some type of diabetes, it is common to observe a loss in body weight; for this reason, in this work, the body weight of the mice was also measured over the 36 days of the experiment. Figure 4 presents changes in the body weight of mice under different treatments. Mice in the control group revealed a significant increase in body weight during the last 2 weeks of treatment (p < 0.05; two-way repeated measures ANOVA). Hyperglycemic (diabetic) mice demonstrated a tendency to lose weight from week 1 and a significant decrease in body weight from week 3 compared with the other groups (p < 0.05; two-way repeated measures ANOVA and Student–Newman–Keuls multiple comparisons test). However, in the group that was administered methanolic extract of J. secunda leaves and the group with the same extract plus the reference drug acarbose, body weight was maintained at a constant level throughout the treatment period (±30 g).
Table 3 shows the quantitative data of the quantitative data on the change in body weight of groups of diabetic mice and those that received extracts of J. secunda during 6 weeks of treatment.
Figure 5 depicts changes in the blood glucose values of mice under different treatments. Statistical analysis showed significant differences between treatments (p < 0.001; two-way repeated measures ANOVA) and weeks of measurement (p < 0.001), as well as a significant interaction between treatment factor and weeks of measurement (p < 0.001). The diabetic mice presented high glucose values from week 1, reaching values of 500 mg/dL at week 6, while the control group exhibited blood glucose levels oscillating at 100 mg/dL throughout the experiment. Conversely, diabetic mice treated with the methanolic extract of J. secunda leaves presented a significant decrease in blood glucose values from week 3 of administration (p < 0.05; post hoc Student–Newman–Keuls multiple comparisons test). It is noteworthy that, during week 3 of treatment, the animals’ glucose levels were equal to those of the control group during weeks 3 and 5, but these increased in week 6, though the levels remained below those of the diabetic mice. Finally, the co-administration of the methanolic extract of J. secunda leaves plus the drug acarbose demonstrated a significant protective effect against the effects of Streptozotocin (hyperglycemia) starting in week 4 of treatment; however, these values were above the blood glucose values of the diabetic mice treated with the J. secunda extract. Finally, the co-administration of the methanolic extract of J. secunda leaves plus the drug acarbose demonstrated a significant protective effect against the effects of Streptozotocin (hyperglycemia) starting in week 4 of treatment; however, these values were above the blood glucose values of the diabetic mice treated with the J. secunda extract.
Table 4 shows the quantitative data of blood glucose levels of groups of diabetic mice and those that received extracts of J. secunda during 6 weeks of treatment.

3.6. Evaluation of Effect on Blood Triglycerides

Figure 6 reveals the changes in the blood triglyceride (TG) levels of mice subjected to different treatments for 36 days (6 weeks). Statistical analysis showed significant differences between treatments (p < 0.001; two-way repeated measures ANOVA) and weeks of measurement (p < 0.001), as well as a significant interaction between treatment factor and weeks of measurement (p < 0.001). The control mice presented oscillating blood TG values, with a tendency to decrease in week 5 with respect to the values exhibited in the baseline measurement (week 0). In the group treated with Streptozotocin (diabetic), elevated TG values were found from week 1 to a maximal value (212.4 mg/dL) in week 6; conversely, in the diabetic mice administered with methanolic extract of J. secunda, the extract exerted a protective effect against the elevated TG levels caused by STZ throughout the treatment and even maintained TG levels below the values found in the control group in weeks 2, 3, and 6 (p < 0.05; post hoc multiple comparisons test of Student–Newman–Keuls). Finally, although co-administration of the extract plus acarbose in diabetic mice caused a decrease in blood TG levels, these levels remained, on average, above those found in the diabetic mice treated with the extract only.
Table 5 shows the quantitative data of blood triglyceride levels of groups of diabetic mice and those that received extracts of J. secunda during 6 weeks of treatment.

4. Discussion

The use of medicinal plants is widespread in Mexico and other Latin American countries, where many sectors of the population utilize this resource due to ease of acquisition; however, these individuals typically lack knowledge of the scientific information that supports their therapeutic benefits. It is estimated that 3352 plant species are used in Mexico [45] out of 25,000–30,000 species in total [46,47], representing 12.7% of the world’s plant species. For these reasons, this work focused on evaluating the different pharmacological properties of Justicia secunda Vahl in a chemical model of diabetes in mice.
The qualitative phytochemical tests carried out on the methanolic extract of J. secunda detected alkaloids, tannins, flavonoids, and sterols. Plant secondary metabolites normally form part of the defense mechanisms against predators, but it has been found that several of these compounds possess various pharmacological and therapeutic properties in humans; for example, evidence suggests that alkaloids have an effect in the treatment of diabetes through different mechanisms; for example, (1) the inhibition of digestive enzymes such as alpha-amylases [48], (2) the inhibition of aldose reductase and protein tyrosine phosphatase, (3) increased insulin secretion, (4) the inhibition of advanced glycation end-products, (5) an increase in glucose uptake by extrahepatic tissues [49,50], and (6) phosphodiesterase inhibition [51].
Antioxidant activity was studied using the in vitro DPPH test. The methanolic extract of the J. secunda leaves (50 mg/mL and 25 mg/mL) demonstrated a total antioxidant activity similar to that of the reference chemical compound (2% ascorbic acid), with maximal activity occurring in even less time than that of ascorbic acid and maintaining this activity for 90 min of reaction time. Although this test was qualitative, the number of some of these metabolites seemed low, and they only exhibited antioxidant activity when prepared at the highest concentrations in the extract. In fact, the other three secondary metabolites detected in this work are considered highly antioxidant agents; for example, flavonoids that structurally consist of a 15-carbon skeleton and two aromatic rings (A and B) connected by a three-carbon chain (C ring) are oxidized by interacting with free radicals, and their OH groups stabilize these radicals. These characteristics are associated with their cardioprotective, anticancer, and other useful effects for the treatment of obesity and diabetes [52].
Another set of metabolites detected in the extract of J. secunda that possesses antioxidant activity corresponded to tannins, a group of polyphenolic compounds characterized by macromolecules and high-molecular-weight polymers (500–2000 Da). These prevent the production of free radicals and lipid peroxidation due to the significant number of OH groups in their structures [53]. In several studies, bitter taste, astringent, anti-Parkinson, anti-Alzheimer, antidepressant, anti-inflammatory, antibacterial, anti-apoptotic, and anti-aging properties have been attributed to these compounds [54,55]. The last qualitatively detected secondary metabolite corresponded to the group of sterols, whose basic structure is the same as 1,2-cyclopentaneperhydrophenantrene and whose molecules are derived from isopentenyl pyrophosphate, which is normally part of cell membranes. This aids in maintaining their fluidity, integrity, and permeability [56].
Considering that the crude extract of J. secunda leaves was obtained using a polar solvent (methanol), we assume that it could have contained traces of some non-polar sterol-type metabolites, such as sterols, which could have contributed to the antioxidant activity found at the highest concentrations tested in the in vitro DPPH test. This has been noted in other works, reporting that sterols prevent the redox imbalance that occurs intracellularly by decreasing the production of reactive oxygen species (ROS) [57]. Although antioxidant activity was determined in vitro in this work, the confirmation of this effect agrees with results reported in vivo for rats administered a relatively high dose of Justicia tranquebariensis leaf extract (400 mg/kg) [58].
Regarding the genoprotective effect evaluated using the micronucleus technique in mouse peripheral blood, we were unable to find any work performed directly with the Justicia genus; however, there are very extensive works that have reported the many properties of this plant and related species of the family Acanthaceae [59]. Using this method, we observed the presence of micronuclei in mature erythrocytes obtained from the peripheral blood of mice administered the mutagenic agent anthracene plus the dry extract of J. secunda leaves; thus, a weak genoprotective effect could be concluded at the end of the experiment. Although the J. secunda extract did not prevent the appearance of micronuclei, it did prevent their number from increasing in mice treated with the extract compared with those treated only with anthracene.
As noted, antioxidant activity in vitro was only observed at high concentrations of the extract; conversely, in the in vivo genoprotective activity test, the mice were administered a relatively high dose of the crude extract, and a weak genoprotective effect was observed. Anthracene is a polycyclic aromatic hydrocarbon that does not possess great potency as a carcinogen and exhibits weak mutagenic and genotoxic activity [60]. We also employed anthracene in this in vivo experiment to compare it with the effect of administering J. secunda extract in mice that were diabetized using streptozocin; this compound has several proven mechanisms of action, including causing mutations in the genetic material of cells by methylating DNA [61,62,63]. We believe that the dry extract of J.secunda prevented the methylating action of anthracene in the DNA of erythrocytes from the peripheral mouse blood, especially in week 2 of the experiment. However, the effect was weak due to the time of exposure to anthracene and the extract (2 weeks). Nonetheless, it was sufficient to prevent the number of micronuclei in the erythrocytes of mice treated with the extract from increasing, which was as high as in those treated with anthracene alone. Although we did not determine the in vivo antioxidant activity along with genoprotective activity, there is evidence that much of the genoprotective activity and the inhibition of clastogenicity caused by mutagen agents is linked to the antioxidant effect of secondary metabolites from plants [64,65,66], suggesting that free radicals such as superoxide are linked to genotoxicity [67].
The methanolic extract of J. secunda was orally administered chronically for 6 weeks in groups of albino mice treated with streptozocin, as described in the Methodology section of this paper. One of the most notable aspects of this model was that the hyperglycemic mice lost a significant amount of weight starting in week 3, a sign that usually occurs in humans with diabetes. Once it enters the β-cells, STZ exerts several effects, including acting as a producer of free radicals and causing DNA methylation; this gives rise to necrosis in pancreatic β-cells. In addition, it has been found that the signal transducer and activator of transcription 3 (STAT3) protein is involved in the death of β-cells and the subsequent hyperglycemia caused by the administration of STZ in mice and rats [62,68].
Body weight loss has been observed in mice treated with STZ in other studies and, as previously mentioned, this may be due to the mechanism of action of STZ in causing DNA methylation. However, this weight loss could also be due to the effect that STZ exerts on various organs, in that STZ administered to albino rats at a dose of 45 mg/kg, in addition to causing weight loss, also gave rise to an increase in the relative weight of the livers and kidneys of these animals, without any apparent effect on the weight of the pancreas [69]. We believe that these effects are the result of the generated energy source requirements of these organs, as they do not receive energy from glucose when endocrine pancreas β-cells die. Various plant extracts have also been administered in other studies, improving the body weight of STZ-treated animals. In one study, the methanolic extract of Justicia adhatoda leaves was administered to BALB/c male mice, and results similar to those presented in this study were reported; in particular, the extract prevented weight loss caused by STZ. Moreover, the authors administered different doses of the extract (up to 400 mg/kg), while our team administered 500 mg/kg without finding evidence of toxicity in the mice. It is worth noting that, in that study, sterols such as β-sitosterol were also detected, among other metabolites [70].
We found that the methanolic extract of J. secunda leaves exerted hypoglycemic and hypotriglyceridemic effects in mice treated with diabetes starting in week 3 of treatment. As noted previously, there are few studies on the Justicia plant, although at least one has been mentioned with results similar to those reported herein [e.g., 70]. In the bibliographic search carried out regarding the study of this plant, although around 400–600 species of Justicia have been identified worldwide [71], only certain works were found in which the different effects and activities of some species of the genus Justicia have been reported. There are reports of the activity of certain species of Justicia on the central nervous system; for example, the anxiolytic effects of J. gendarussa [72]; the anxiolytic effects of Justicia spicigera [73]; the anticonvulsant activity of Justicia extensa [74]; activity in peripheral organs and tissues, such as the cytotoxic activity of the species Justicia betonica and Justicia vahlii [75,76]; the anti-inflammatory effects of Justicia adhatoda [77]; and, of course, the antioxidant activity of J. adhatoda, J. gendarussa, and J. secunda [78,79,80,81]. We mentioned that there have been few reports on the toxicity of the J. secunda plant, but some studies have found that the LD50 is more than 3800–5000 mg/kg in rats [81,82]. Therefore, we believe that its effects on glucose and triglyceride levels are, in fact, due to the secondary metabolites present in this species. These effects could be explained by the antioxidant, antigenotoxic, and weight loss-protective contributions of all of the metabolites contained in the dry extract of J. secunda; as such, these metabolites may mitigate the hyperglycemic effect resulting from the mechanism of action of STZ. We should also note that some non-polar metabolites of this steroid type, including non-polar metabolites such as sterols, can contribute to these properties.
It is noteworthy that, in addition to its hypoglycemic effect, the methanolic extract of J. secunda also exerted a marked hypotriglyceridemic effect. This could be related to the hepatoprotective activity of this species, which has been reported in other works, and could contribute to better use of lipids by the livers of hyperglycemic mice [83,84]. However, in this sense, it should be noted that over several weeks of treatment, TG levels were found to be below even those of the control group mice, which might not be beneficial for individuals in the long term and thus needs to be analyzed in more detail. On the other hand, in addition to the antioxidant and genoprotective properties of the administered extract, we sought to explore the possible mechanism of action by which this species promotes these effects, given that some studies have reported that several plants used in the treatment of diabetes possess additional mechanisms of action similar to those mentioned in this work: for example, the inhibition of intestinal brush-border enzyme activity, such as that of α-glucosidase, which ultimately promotes a delay in the absorption of glucose ingested in food into the intestine; contribute to preventing blood glucose levels from rising; and stimulate glucose uptake in the target tissues of insulin, in addition to the liver [25,85]. We administered acarbose plus J. secunda extract to another group of mice previously treated with STZ with the idea that, if the secondary metabolites contained in the methanolic extract of Justicia secunda can also inhibit intestinal α-glucosidase, then a synergistic effect would be caused by this extract in addition to the effect of the reference drug. Furthermore, blood glucose levels (and perhaps those of triglycerides) would decrease more than in the hyperglycemic mice treated only with the extract. However, although decreases in glucose and triglyceride levels were observed in this group, they were not similar to those caused by the extract alone; that is, we can confirm what has been found in other works [86], although it is possible that the dose of acarbose administered (300 mg/kg) was insufficient to observe the expected synergy.
Finally, in addition to the effects we found for the methanolic extract of J. secunda leaves, this work is relevant because we explored the effects of chronic oral administration with a relatively high dose (500 mg/kg) over 6 weeks on the physiological parameters affected by diabetes. Conversely, in several studies, acute evaluations were only carried out with durations ranging from 2 h to 1 week of administration. This is important considering that in Mexican communities, where these species are consumed in the form of infusions, intake takes place on a daily basis while the patient remains in a fasting state. Therefore, the consumption scheme of many of these plants in herbal Traditional Medicine contexts was imitated, and no toxic effects were found in the mice. These results could encourage the inhabitants of these low-income communities to cultivate this species not only for ethnomedical use, as has been discussed in other works utilizing Justicia as an object of study [87,88,89].

5. Limitations and Considerations

There are clinical limitations of this study that must be taken into account. While our results are promising regarding the consideration of J. secunda as an alternative therapy for treating diabetes, they can only be extrapolated to a human context with more in-depth studies regarding the safety of the dosage used. It will also have to be verified as to whether the chronic administration of this species leads to side effects in the future. However, our experimental model presents some similarities to the way in which people use this and other plant species to treat diabetes: (i) The extract was administered chronically, implying a relatively long time interval in the life cycle of the rodents; similarly, in humans, people with limited resources who suffer from diabetes also take infusions of the plant (orally) for long periods of their lives (if not all their lives). (ii) A high dose (500 mg/kg) was administered while, in Mexican communities, people prepare the infusion by taking 20 or more leaves of the plant and boiling them in tap water. (iii) In humans and mice, the baseline blood glucose levels in a fasting state were similar: around 80–100 mg/dL.

6. Conclusions

This work studied Justicia secunda Vahl, which is used frequently in many Latin American countries, including Mexico. We emphasized the chronic and long-term administration of a high oral dose (500 mg/kg) without finding toxic effects. Therapeutic properties, including antioxidant activity, protection against weight loss, a hypotriglyceridemic effect, and hypoglycemic activity, were found in diabetic mice. All of these properties can be mediated by secondary metabolites detected in the fractions of the methanolic extract. Alkaloids, flavonoids, and tannins provided the extract with the ability to reduce the DPPH radical and act as an antioxidant chemical cocktail that could confer therapeutic properties to mitigate the diabetogenic mechanism of action of Streptozotocin. Furthermore, a mechanism of action similar to that of the inhibition of intestinal α-glucosidases may also participate in these therapeutic actions.
We propose that Justicia secunda can be considered as a good alternative candidate for the treatment of diabetes, without dismissing conventional allopathic treatments.

Author Contributions

T.F.-A., Á.M.-G. and J.A.M.-G. conceived the idea and planned the experiments; P.X.E.-R., D.E.N.-Á. and J.A.M.-P. performed parts of the experiments, the description of the results, and part of the figures and tables; E.O.M.-S., J.M.T.-L., E.V.R.-N., E.M.-B., I.Á.-G. and G.I.-C. performed part of the experiments, the statistical analysis, and part of the figures. G.I.-C., T.F.-A., Á.M.-G. and J.A.M.-G. wrote the article. All authors have read and agreed to the published version of the manuscript.

Funding

The study was partially funded by the Research and Postgraduate secretariat with research project SIP20240774 of the Escuela Nacional de Ciencias Biológicas, of Instituto Politécnico Nacional, and SIP 20240267 ESM-IPN; SIP20241869 ESCOM-IPN.

Institutional Review Board Statement

The protocol was approved by the Research Ethics Committee of the National School of Biological Sciences of the National Polytechnic Institute, under folio ZOO-005-2022, dated 22 December 2022.

Informed Consent Statement

Not applicable.

Data Availability Statement

The original contributions presented in this study are included in the article. Further inquiries can be directed to the corresponding author.

Conflicts of Interest

The authors declare no conflicts of interest.

References

  1. Martínez, M. Las Plantas Medicinales de México, 3rd ed.; Botas: Ciudad de México, México, 1944; 630p. [Google Scholar]
  2. Hersch, P. De hierbas y herbolarios en el México actual. Antropol. Méx. 1999, 7, 60–65. [Google Scholar]
  3. Guzmán-Maldonado, S.H.; Díaz-Huacuz, R.S.; González-Chavira, M.M. Plantas medicinales: La realidad de una tradición ancestral. INIFAP 2017, 1, 1–36. [Google Scholar]
  4. WHO. Traditional Medicine Strategy 2014–2023; World Health Organization: Geneva, Switzerland, 2013; 78p.
  5. Lange, D. International trade in medicinal and aromatic planst. Actors, volumes and commodities. In Medicinal and Aromatic Plants; Bogers, R.J., Craker, L.E., Lange, D., Eds.; Springer: Berlin/Heidelberg, Germany, 2006; Volume 11, pp. 155–170. [Google Scholar]
  6. Cruz-Pérez, A.L.; Barrera-Ramos, J.; Bernal-Ramírez, L.A.; Bravo-Avilez, D.; Rendón-Aguilar, D. Actualized inventory of medicinal plants used in traditional medicine in Oaxaca, México. J. Ethnobiol. Ethnomed. 2021, 17, 7. [Google Scholar]
  7. INIFAP. México, Segundo Lugar Mundial en Registro de Plantas Medicinales. Available online: www.gob.mx/agricultura/prensa/mexico-segundo-lugar-mundial-en-registro-de-plantas-medicinales (accessed on 6 February 2022).
  8. García de Alba-García, J.E.; Ramírez-Hernández, B.C.; Robles-Arellano, G.; Zañudo Hernández, J.; Salcedo-Rocha, A.L.; García de Alba-Verduzco, J.E. Conocimiento y uso de las plantas medicinales en la zona metropolitana de Guadalajara. Desacatos 2012, 39, 29–44. [Google Scholar]
  9. Hamilton, A.C. Medicinal plants, conservation and livelihoods. Biodivers. Conserv. 2004, 13, 1477–1517. [Google Scholar] [CrossRef]
  10. He, J.; Yang, B.; Dong, M.; Wang, Y. Crossing the roof of the world: Trade in medicinal plants from Nepal to China. J. Ethnopharmacol. 2018, 224, 100–110. [Google Scholar] [CrossRef]
  11. Eshete, M.A.; Molla, E.L. Cultural significance of medicinal plants in healing human ailments among Guji semi-pastoralist people, Suro Barguda District, Ethiopia. J. Ethnobiol. Ethnomed. 2021, 17, 61. [Google Scholar] [CrossRef] [PubMed]
  12. Acosta-Recalde, P.; Lugo, G.; Vera, Z.; Morinigo, M.; Maidana, G.M.; Samaniego, L. Uso de plantas medicinales y fitoterápicos en pacientes con Diabetes Mellitus tipo 2. Mem. Inst. Investig. Cienc. Salud. 2018, 16, 6–11. [Google Scholar] [CrossRef]
  13. Geck, M.S.; Reyes-García, A.J.; Casu, L.; Leonti, M. Acculturation and ethnomedicine: A regional comparison of medicinal plant knowledge among the zoque of southern México. J. Ethnopharmacol. 2016, 187, 146–159. [Google Scholar] [CrossRef]
  14. Andrade-Cetto, A.; Heinrich, M. Mexican plants with hypoglycaemic effect used in the treatment of diabetes. J. Ethnopharmacol. 2005, 99, 325–348. [Google Scholar] [CrossRef]
  15. Chávez-Bustos, E.A.; Morales-González, A.M.; Liliana Anguiano-Robledo, L.; Madrigal-Santillán, E.O.; Valadez-Vega, C.; Lugo-Magaña, O.; Mendoza-Pérez, J.A.; Fregoso-Aguilar, T.A. Bauhinia forficata Link, Antioxidant, genoprotective, and hypoglycemic activity in a murine model. Plants 2022, 11, 3052. [Google Scholar] [CrossRef] [PubMed]
  16. Bautista-Hernández, I.; Aguilar, C.N.; Martínez-Ávila, G.C.; Torres-León, C.; Ilina, A.; Flores-Gallegos, A.A.; Verma, D.K.; Chávez-González, M.L. Mexican Oregano (Lippia graveolens Kunth) as Source of Bioactive Compounds: A Review. Molecules 2021, 26, 5156. [Google Scholar] [CrossRef]
  17. Andrade-Cetto, A.; Wiedenfeld, H. Anti-hyperglycemic effect of Opuntia streptacantha Lem. J. Ethnopharmacol. 2011, 133, 940–943. [Google Scholar] [CrossRef] [PubMed]
  18. Santos-Díaz, M.S.; Barba-de la Rosa, A.P.; Héliès-Toussaint, C.; Guéraud, F.; Nègre-Salvayr, A. Opuntia spp.: Characterization and benefits in chronic diseases. Oxidative Med. Cell. Longev. 2017, 2017, 8634249. [Google Scholar] [CrossRef] [PubMed]
  19. Pingali, U.; Abid-Ali, M.; Gundagani, S.; Nutalapati, C. Evaluation of the effect of an aqueous extract of Azadirachta indica (Neem) leaves and twigs on Glycemic control, endothelial dysfunction and systemic inflammation in subjects with type 2 diabetes mellitus: A randomized, double-blind, placebo-controlled clinical study. Diabetes Metab. Syndr. Obes. Targets Ther. 2020, 13, 4401–4412. [Google Scholar]
  20. Patil, S.M.; Shirahatti, P.S.; Ramith Ramu, R. Azadirachta indica A. Juss (neem) against diabetes mellitus: A critical review on its phytochemistry, pharmacology, and toxicology. J. Pharm. Pharmacol. 2022, 74, 681–710. [Google Scholar] [CrossRef]
  21. Roman-Ramos, R.; Flores-Saenz, J.L.; Alarcon-Aguilar, F.J. Anti-hyperglycemic effect of some edible plants. J. Ethnopharmacol. 1995, 48, 25–32. [Google Scholar] [CrossRef]
  22. Chattopadhyay, R.R. A comparative evaluation of some blood sugar lowering agents of plant origin. J. Ethnopharmacol. 1999, 67, 367–372. [Google Scholar] [CrossRef]
  23. Ovalle-Magallanes, B.; Déciga-Campos, M.; Mata, R. Antinociceptive and hypoglycaemic evaluation of Conyza filaginoides (D.C.) Hieron Asteraceae. J. Pharm. Pharmacol. 2015, 67, 1733–1743. [Google Scholar] [CrossRef]
  24. Ahangarpour, A.; Heidari, H.; Oroojan, A.A.; Mirzavandi, F.; Esfehani, K.N.; Mohammadi, Z.D. Antidiabetic, hypolipidemic and hepatoprotective effects of Arctium lappa root’s hydro-alcoholic extract on nicotinamide-streptozotocin induced type 2 model of diabetes in male mice. Avicenna J. Phytomed. 2017, 7, 169–179. [Google Scholar]
  25. Valdivia-Correa, B.; Gómez-Gutiérrez, C.; Uribe, M.; Méndez-Sánchez, N. Herbal medicine in Mexico: A cause of hepatotoxicity. A critical review. Int. J. Mol. Sci. 2016, 17, 235. [Google Scholar] [CrossRef] [PubMed]
  26. WHO. Monographs on Selected Medicinal Plants; World Health Organization: Geneva, Switzerland, 2002; Volume 2, pp. 1–358.
  27. EFSA. Compendium of Botanicals Reported to Contain Naturally Occurring Substances of Possible Concern for Human Health When Used in Food and Food Supplements; European Food Safety Authority: Parma, Italy, 2012; Volume 10, p. 2663. [CrossRef]
  28. Zambrano-Mora, P.; Bustamante-Pesantes, K.E. Caracterización y studio fitoquímico de Justicia secunda Vahl (sanguinaria, chingamochila, insulina). Rev. Cub. Plant. Med. 2017, 22, 1–8. [Google Scholar]
  29. Domínguez, I. Justicia Secunda Vahl, Especie Utilizada en la Medicina Indígena COLOMBIANA. Universidad de Sevilla. Colombia. 2019. Available online: https://idus.us.es/server/api/core/bitstreams/c4c83c94-8023-4a83-af48-a1f50315e906/content. (accessed on 14 August 2022).
  30. Raz, L. Agudelo, Justicia Secunda Vahl. Catálogo de Plantas y Líquenes de Colombia. Universidad Nacional de Colombia. 2023. Available online: https://ipt.biodiversidad.co/sib/resource?r=catalogo_plantas_liquenes (accessed on 10 October 2023).
  31. Ortíz-Andrade, R.; Cabañas-Wuan, A.; Arana-Argáez, V.E.; Alonso-Castro, A.J.; Zapata-Bustos, R.; Salazar-Olivo, L.A.; Domínguez, F.; Chávez, M.; Carranza-Álvarez, C.; García-Carranca, A. Antidiabetic effects of Justicia spicigera Schltdl (Acanthaceae). J. Ethnopharmacol. 2012, 143, 455–462. [Google Scholar] [CrossRef]
  32. González-Trujano, M.E.; Domínguez, F.; Pérez-Ortega, G.; Aguillón, M.; Martínez-Vargas, D.; Almazán-Alvarado, S.; Martínez, A. Justicia spicigera Schltdl. and kaempferitrin as potential anticonvulsant natural products. Biomed. Pharmacother. 2017, 92, 240–248. [Google Scholar] [CrossRef]
  33. Quiñonez-Bastidas, G.N.; Navarrete, A. Mexican plants and derivates compounds as alternative for inflammatory and neuropathic pain treatment—A review. Plants. 2021, 10, 865. [Google Scholar] [CrossRef] [PubMed]
  34. Cabada-Aguirre, P.; López-López, A.M.; Ostos-Mendoza, K.C.; Garay-Buenrostro, K.D.; Luna-Vital, D.A.; Mahady, G.B. Mexican traditional medicines for women’s reproductive health. Sci. Rep. 2023, 13, 2807. [Google Scholar] [CrossRef] [PubMed]
  35. IDF. Diabetes Atlas. International Diabetes Federation. 11th ed. 2025. Available online: https://diabetesatlas.org/resources/idf-diabetes-atlas-2025/ (accessed on 27 May 2025).
  36. ENSANUT. Encuesta Nacional de Salud y Nutrición 2018. Presentación de Resultados (insp.mx). Available online: https://ensanut.insp.mx/encuestas/ensanut2018/doctos/informes/ensanut_2018_presentacion_resultados.pdf (accessed on 24 June 2022).
  37. Bello-Chavolla, O.Y.; Antonio-Villa, N.E.; Fermín-Martínez, C.A.; Fernández-Chirino, L.; Vargas-Vázquez, A.; Ramírez-García, D.; Basile-Alvarez, M.R.; Hoyos-Lázaro, A.E.; Carrillo-Larco, R.M.; Wexler, D.J.; et al. Diabetes-related excess mortality in Mexico: A comparative analysis of national death registries between 2017–2019 and 2020. Diab. Care. 2022, 45, 2957–2966. [Google Scholar] [CrossRef]
  38. Norma Oficial Mexicana, NOM-033-ZOO-1995. Sacrificio Humanitario de Los Animales Domésticos Y Silvestres, Publicada en El Dof El 16 de Julio de 1996. Available online: https://www.imss.gob.mx/sites/all/statics/profesionalesSalud/investigacionSalud/cbis/nom-033-zoo-1995.pdf (accessed on 24 June 2022).
  39. Norma Oficial Mexicana, NOM-062-ZOO-1999. Especificaciones Técnicas Para la Producción, Cuidado Y Uso de Los Animales de Laboratorio, Publicada en El Dof El 22 de Agosto de 2001. Available online: https://www.gob.mx/cms/uploads/attachment/file/203498/NOM-062-ZOO-1999_220801.pdf (accessed on 24 June 2022).
  40. Norma Oficial Mexicana, NOM-087-ECOL-1995. Requisitos Para la Separación, Envasado, Almacenamiento, Recolección, Transporte, Tratamiento Y Disposición Final de Los Residuos Peligrosos Biológico-Infecciosos Que SE Generan en Establecimientos Que Prestan Atención Médica, Publicada en El Dof El 7 de Noviembre de 1995. Available online: https://siga.jalisco.gob.mx/assets/documentos/normatividad/nom087semarnat1995.html (accessed on 24 June 2022).
  41. Shafodino, F.S.; Lusilao, J.M.; Mwapagha, L.M. Phytochemical characterization and antimicrobial activity of Nigella sativa seeds. PLoS ONE 2022, 17, e0272457. [Google Scholar] [CrossRef]
  42. Blois, M.S. Antioxidant determinations by the use of a stable free radical. Nature 1958, 181, 1199–1200. [Google Scholar] [CrossRef]
  43. Da Silva-Mendoça, J.; Avellaneda-Guimarães, R.D.; Zorgetto-Pinheiro, V.A.; Di Pietro-Fernandes, C.; Marcelino, G.; Bogo, D.; De Cássia-Freitas, K.; Aiko-Hiane, P.; Silva- De-Pádua-Melo, E.; Brandão-Vilela, M.L.; et al. Natural antioxidant evaluation: A review of detection methods. Molecules 2022, 27, 3563. [Google Scholar] [CrossRef]
  44. Guevara-Martínez, S.J.; Villanueva-Mejía, F.; Fregoso-Aguilar, T.A.; Zamudio-Ojeda, A.; Herrera-Bucio, R.; Morales-Palacios, F.G. Chronic administration of tetrahydrofuran extracts of Cochlospermum vitifolium (Wild) Sprengel in a mouse diabetes model: Hypoglycemic, antioxidant, and genoprotective effects. Biomed. Pharmacother. 2025, 187, 118132. [Google Scholar] [CrossRef] [PubMed]
  45. Bye, R. La intervención del hombre en la diversificación de las plantas en México. In Diversidad Biológica de Mexico: Orígenes y Distribución, 1st ed.; Ramamoorthy, T.P., Bye, R., Lot, A., Fa, J., Eds.; Instituto de Biología, UNAM: Mexico city, Mexico, 1994; pp. 689–714. [Google Scholar]
  46. Aguilar, A.; Camacho, J.R.; Chino, S.; Jácquez, P.; López, M.E. Herbario Medicinal del Instituto Mexicano del Seguro Social: Información Etnobotánica, 1st ed.; Instituto Mexicano del Seguro Social: Mexico city, Mexico, 1994; 253p.
  47. Toledo, V.M.; Ordoñez, M.J. El panorama de la biodiversidad de México: Una revision de los hábitats terrestres. In Diversidad biologica de Mexico: Orígenes y distribución, 1st ed.; Ramamoorthy, T.P., Bye, R., Lot, A., Fa, J., Eds.; Instituto de Biología, UNAM: Mexico city, Mexico, 1998; pp. 739–757. [Google Scholar]
  48. Gao, H.; Huang, Y.N.; Gao, B.; Li, P.; Inagaki, C.; Kawabata, J. Inhibitory Effect on α-Glucosidase by Adhatoda Vasica Nees. Food Chem. 2008, 108, 965–972. [Google Scholar] [CrossRef]
  49. Wang, F.W.; Zhu, N.; Zhou, F.; Lin, D.X. Natural aporphine alkaloids with potential to impact metabolic syndrome. Molecules 2021, 26, 6117. [Google Scholar] [CrossRef]
  50. Behl, T.; Gupta, A.; Albratty, M.; Najmi, A.; Meraya, A.M.; Alhazmi, H.A.; Anwer, K.; Bhatia, S.; Bungau, S.G. Alkaloidal phytoconstituents for diabetes management: Exploring the unrevealed potential. Molecules 2022, 27, 5851. [Google Scholar] [CrossRef] [PubMed]
  51. Bhambhani, S.; Kondhare, K.R.; Giri, A.P. Diversity in chemical structures and biological properties of plant alkaloids. Molecules 2021, 26, 3374. [Google Scholar] [CrossRef]
  52. Al-Ishaq, R.K.; Abotaleb, M.; Kubatka, P.; Kajo, K.; Büsselberg, D. Flavonoids and their anti-diabetic effects: Cellular mechanisms and effects to improve blood sugar levels. Biomolecules 2019, 9, 430. [Google Scholar] [CrossRef]
  53. Watrelot, A.A.; Norton, E.L. Chemistry and reactivity of tannins in Vitis spp.: A review. Molecules 2020, 25, 2110. [Google Scholar] [CrossRef] [PubMed]
  54. Hussain, G.; Huang, J.; Rasul, A.; Anwar, H.; Imran, A.; Maqbool, J.; Razzaq, A.; Aziz, N.; Makhdoom, E.H.; Konuk, M.; et al. Putative roles of plant-derived tannins in neurodegenerative and neuropsychiatry disorders: An updated review. Molecules 2019, 24, 213. [Google Scholar] [CrossRef]
  55. Soares, S.; Brandão, E.; Guerreiro, C.; Soares, S.; Mateus, N.; de Freitas, V. Tannins in food: Insights into the molecular perception of astringency and bitter Taste. Molecules 2020, 25, 2590. [Google Scholar] [CrossRef]
  56. Kopylov, A.T.; Malsagova, K.A.; Stepanov, A.A.; Kaysheva, A.L. Diversity of plant sterols metabolism: The impact on human health, sport, and accumulation of contaminating sterols. Nutrients 2021, 13, 1623. [Google Scholar] [CrossRef]
  57. Pramanik, P.K.; Chakraborti, S.; Bagchi, A.; Chakraborti, T. Bioassay-based Corchorus capsularis L. leaf-derived β-sitosterol exerts antileishmanial effects against Leishmania donovani by targeting trypanothione reductase. Sci. Rep. 2020, 10, 20440. [Google Scholar] [CrossRef] [PubMed]
  58. Sukalingam, K.; Ganesan, K.; Xu, B. Protective effect of aqueous extract from the leaves of Justicia tranquebariesis against thioacetamide-induced oxidative stress and hepatic fibrosis in rats. Antioxidants 2018, 7, 78. [Google Scholar] [CrossRef] [PubMed]
  59. Khandelwal, P.; Wadhwani, B.D.; Rao, R.S.; Mali, D.; Vyas, P.; Kumar, T.; Nair, R. Exploring the pharmacological and chemical aspects of pyrrolo-quinazoline derivatives in Adhatoda vasica. Heliyon 2024, 10, e25727. [Google Scholar] [CrossRef]
  60. Mastandrea, C.; Chichizola, C.; Ludueña, B.; Sánchez, S.; Álvarez, H.; Gutiérrez, A. Hidrocarburos aromáticos policíclicos. Riesgos para la salud y marcadores biológicos. Acta Bioquím. Clín. Latinoam. 2005, 39, 27–36. [Google Scholar]
  61. Szkudelski, T. The Mechanism of alloxan and streptozotocin action in B cells of the rat pancreas. Physiol. Res. 2001, 50, 536–546. [Google Scholar] [CrossRef]
  62. Lenzen, S. The mechanisms of alloxan- and streptozotocin-induced diabetes. Diabetologia 2008, 51, 216–226. [Google Scholar] [CrossRef]
  63. King, A.; Bowe, J. Animal models for diabetes: Understanding the pathogenesis and finding new treatments. Biochem. Pharmacol. 2016, 99, 1–10. [Google Scholar] [CrossRef]
  64. Madrigal-Santillán, E.; Fragoso-Antonio, S.; Valadez-Vega, C.; Solano-Solano, G.; Zúñiga-Pérez, C.; Sánchez-Gutiérrez, M.; Izquierdo-Vega, J.A.; Gutiérrez-Salinas, J.; Esquivel-Soto, J.; Esquivel-Chirino, C.; et al. Investigation on the protective effects of cranberry against the DNA damage induced by Benzo[a]pyrene. Molecules 2012, 17, 4435–4451. [Google Scholar] [CrossRef]
  65. Ho, C.K.; Siu-wai, C.; Siu, P.M.; Benzie, I.F. Genoprotection and genotoxicity of green tea (Camellia sinensis): Are they two sides of the same redox coin? Redox Rep. 2013, 18, 4. [Google Scholar] [CrossRef]
  66. Adil, M.; Dastagir, G.; Quddoos, A.; Naseer, M.; Filimban, F.Z. HPLC analysis, genotoxic and antioxidant potential of Achillea millefolium L. and Chaerophyllum villosum Wall ex. Dc. BMC Complement. Med. Ther. 2024, 24, 91. [Google Scholar] [CrossRef]
  67. Uribe-Hernández, R.; Pérez-Zapata, A.J. Inducción de la fragmentación del DNA por Antraceno y Benzo(a) Pireno en leucocitos polimorfonucleares humanos in vitro. Interciencia 2005, 30, 419–423. [Google Scholar]
  68. Weng, Q.; Zhao, M.; Zheng, J.; Yang, L.; Xu1, Z.; Zhang, Z.; Wang, J.; Wang, J.; Yang, B.; Lu, Q.B.; et al. STAT3 dictates β-cell apoptosis by modulating PTEN in streptozocin-induced hyperglycemia. Cell Death Differ. 2020, 27, 130–145. [Google Scholar] [CrossRef]
  69. Zafar, M.; Naqvi, N.U.H. Effects of STZ-induced diabetes on the relative weights of kidney, liver and pancreas in albino rats: A comparative study. Int. J. Morphol. 2010, 28, 135–142. [Google Scholar] [CrossRef]
  70. Ameer, M.R.; Khalid, Z.M.; Shinwari, M.I.; Ali, H. Correlation among antidiabetic potential, biochemical parameters and GC-MS analysis of the crude extracts of Justicia adhatoda L. Pak. J. Bot. 2021, 53, 2111–2125. [Google Scholar] [CrossRef]
  71. Carneiro, M.R.B.; Sallum, L.O.; Martins, J.L.R.; Peixoto, J.d.C.; Napolitano, H.B.; Rosseto, L.P. Overview of the Justicia genus: Insights into its chemical diversity and biological potential. Molecules 2023, 28, 1190. [Google Scholar] [CrossRef]
  72. Subramanian, N.; Jothimanivannan, C.; Kumar, R.S.; Kameshwaran, S. Evaluation of anti-anxiety activity of Justicia gendarussa Burm. Pharmacoligia 2013, 4, 404–407. [Google Scholar] [CrossRef]
  73. García-Ríos, R.I.; Mora-Pérez, A.; González-Torres, D.; Carpio-Reyes, R.J.; Soria-Fregozo, C. Anxiolytic-like effect of the aqueous extract of Justicia spicigera leaves on female rats: A comparison to diazepam. Phytomedicine 2019, 55, 9–13. [Google Scholar] [CrossRef] [PubMed]
  74. Sowemimo, A.A.; Adio, O.; Fageyinbo, S. Anticonvulsant activity of the methanolic extract of Justicia extensa T. Anders. J. Ethnopharmacol. 2011, 138, 697–699. [Google Scholar] [CrossRef]
  75. Naik, S.K.; Manjula, B.L.; Balaji, M.V.; Marndi, S.; Kumar, S.; Devi, R.S. Antibacterial activity of Justicia betonica Linn. Asian Pac. J. Health Sci. 2022, 9, 227–230. [Google Scholar] [CrossRef]
  76. Basit, A.; Ahmad, S.; Khan, K.U.R.; Naeem, A.; Usman, M.; Ahmed, I.; Shahzad, M.N. Chemical profiling of Justicia vahlii Roth. (Acanthaceae) using UPLC-QTOF-MS and GC-MS analysis and evaluation of acute oral toxicity, antineuropathic and antioxidant activities. J. Ethnopharmacol. 2022, 6, 114942. [Google Scholar] [CrossRef]
  77. Basit, A.; Shutian, T.; Khan, A.; Khan, S.M.; Shahzad, R.; Khan, A.; Khan, S.; Khan, M. Anti-inflammatory and analgesic potential of leaf extract of Justicia adhatoda L. (Acanthaceae) in Carrageenan and Formalin-induced models by targeting oxidative stress. Biomed. Pharmacother. 2022, 153, 113322. [Google Scholar] [CrossRef]
  78. Kaur, A.; Katoch, D.; Singh, B.; Arora, S. Seclusion of vacicine: An quinazoline alkaloid from bioactive fraction of Justicia adhatoda and its antioxidant, antimutagenic and anticancerous activities. J. Global Biosci. 2016, 5, 3836–3850. [Google Scholar]
  79. Zhang, H.X.; Xia, Z.; Xu, T.Q.; Chen, Y.M.; Zhou, G.X. New compounds from the aerial parts of Justicia gendarussa Burm.f. and their antioxidant and anti-inflammatory activities. Nat. Prod. Res. 2021, 35, 3478–3486. [Google Scholar] [CrossRef]
  80. Świątek, Ł.; Sieniawska, E.; Sinan, K.I.; Zengin, G.; Boguszewska, A.; Hryć, B.; Bene, K.; Polz-Dacewicz, M.; Dall’Acqua, S. Chemical characterization of different extracts of Justicia secunda Vahl and determination of their anti-oxidant, anti-enzymatic, anti-viral, and cytotoxic properties. Antioxidants 2023, 12, 509. [Google Scholar] [CrossRef]
  81. Onochie, A.U.; Oli1, A.H.; Oli, A.N.; Ezeigwe, O.C.; Nwaka, A.C.; Okani, C.O.; Okam, P.C.; Ihekwereme, C.P.; Okoyeh, J.N. The pharmacobiochemical effects of ethanol extract of Justicia secunda. Vahl Leaves in Rattus norvegicus. J. Exp. Pharmacol. 2020, 12, 423–437. [Google Scholar] [CrossRef] [PubMed]
  82. Akintimehin, E.S.; Karigidi, K.O.; Omogunwa, T.S.; Adetuyi, F.O. Safety assessment of oral administration of ethanol extract of Justicia carnea leaf in healthy wistar rats: Hematology, antioxidative and histology studies. Clin. Phytosci. 2021, 7, 2. [Google Scholar] [CrossRef]
  83. Mondal, M.; Hossain, M.; Rahman, M.A.; Saha, S.; Uddin, N.; Hasan, R.; Kader, A.; Wahed, T.B.; Kundu, S.K.; Islam, M.T.; et al. Hepatoprotective and antioxidant activities of Justicia gendarussa leaf extract in carbofuran-induced hepatic damage in rats. Chem. Res. Toxicol. 2019, 32, 2499–2508. [Google Scholar] [CrossRef]
  84. Anyasor, G.N.; Moses, N.; Kale, O. Hepatoprotective and hematological effects of Justicia secunda Vahl leaves on carbon tetrachloride induced toxicity in rats. Biotech. Histochem 2020, 95, 349–359. [Google Scholar] [CrossRef] [PubMed]
  85. Theiler, B.A.; Istvanits, S.; Zehl, M.; Marcourt, L.; Urban, E.; Caisad, L.O.E.; Glasl, S. HPTLC bioautography guided isolation of α-Glucosidase inhibiting compounds from Justicia secunda Vahl (Acanthaceae). Phytochem. Anal. 2017, 28, 87–92. [Google Scholar] [CrossRef]
  86. Escandón-Rivera, S.M.; Mata, R.; Andrade-Cetto, A. Molecules isolated from Mexican hypoglycemic plants: A review. Molecules 2020, 25, 4145. [Google Scholar] [CrossRef]
  87. Arogbodo, J.O. Evaluation of the phytochemical, proximate and elemental constituents of Justicia secunda M. Vahl leaf. Int. J. Innov. Sci. Res. Technol. 2020, 5, 1262–1268. Available online: https://www.researchgate.net/publication/342525145_Evaluation_of_the_Phytochemical_Proximate_and_Elemental_Constituents_of_Justicia_secunda_M_Vahl_Leaf (accessed on 13 October 2023).
  88. Ajuru, M.G.; Kpekot, K.A.; Robinson, G.E.; Amutadi, M.C. Proximate and phytochemical analysis of the laeaves of Justicia carnea Lindi. and Justicia secunda Vahl and its taxonomic implications. J. Biomed. Biosens. 2021, 2, 1–12. [Google Scholar]
  89. Alatorre-Cruz, J.M.; Carreño-López, R.; Alatorre-Cruz, G.C.; Paredes-Esquivel, L.J.; Santiago-Saenz, Y.O.; Nieva-Vázquez, A. Traditional Mexican food: Phenolic content and public health relationship. Foods 2023, 12, 1233. [Google Scholar] [CrossRef] [PubMed]
Diabetology 06 00056 g001
Figure 1. Spectrum obtained with infrared analysis with Fourier transform (FTIR) of the methanolic extract of Justicia secunda leaves.
Figure 1. Spectrum obtained with infrared analysis with Fourier transform (FTIR) of the methanolic extract of Justicia secunda leaves.
Diabetology 06 00056 g001
Diabetology 06 00056 g002
Figure 2. Percentage of inhibition of the presence of DPPH of four concentrations of J. secunda during 90 min of the reaction. Each point represents the average of two absorbance measurements (517 nm) as described in Methodology.
Figure 2. Percentage of inhibition of the presence of DPPH of four concentrations of J. secunda during 90 min of the reaction. Each point represents the average of two absorbance measurements (517 nm) as described in Methodology.
Diabetology 06 00056 g002
Diabetology 06 00056 g003
Figure 3. Genoprotective activity measured as number of micronuclei in peripheral mouse blood. Data expressed as mean ± standard error of the mean (SEM). * p < 0.05; comparison of anthracene + J. secunda vs. anthracene. Two-way repeated measures ANOVA. n = 6 mice per group.
Figure 3. Genoprotective activity measured as number of micronuclei in peripheral mouse blood. Data expressed as mean ± standard error of the mean (SEM). * p < 0.05; comparison of anthracene + J. secunda vs. anthracene. Two-way repeated measures ANOVA. n = 6 mice per group.
Diabetology 06 00056 g003
Diabetology 06 00056 g004
Figure 4. Changes in the body weight of the mice under different treatments for 36 days (6 weeks). Data are expressed as mean ± standard error of the mean (SEM). * Denotes a p < 0.05; comparison of diabetic vs. all groups; t denotes p < 0.05, comparison vs. all groups (two-way repeated measures ANOVA and Student—Newman—Keuls tests). n = 6 mice per group.
Figure 4. Changes in the body weight of the mice under different treatments for 36 days (6 weeks). Data are expressed as mean ± standard error of the mean (SEM). * Denotes a p < 0.05; comparison of diabetic vs. all groups; t denotes p < 0.05, comparison vs. all groups (two-way repeated measures ANOVA and Student—Newman—Keuls tests). n = 6 mice per group.
Diabetology 06 00056 g004
Diabetology 06 00056 g005
Figure 5. Glucose levels recorded in mice under different treatments for 36 days (6 weeks). Data is expressed as mean ± standard error of the mean (SEM). * Denotes p < 0.05; comparison of diabetic + J. secunda vs. diabetic. t denotes p < 0.05; comparison of Diabetic + J. secunda + acarbose vs. diabetic and diabetic + J. secunda. (two-way repeated measures ANOVA and Student—Newman—Keuls tests). n = 6 mice per group.
Figure 5. Glucose levels recorded in mice under different treatments for 36 days (6 weeks). Data is expressed as mean ± standard error of the mean (SEM). * Denotes p < 0.05; comparison of diabetic + J. secunda vs. diabetic. t denotes p < 0.05; comparison of Diabetic + J. secunda + acarbose vs. diabetic and diabetic + J. secunda. (two-way repeated measures ANOVA and Student—Newman—Keuls tests). n = 6 mice per group.
Diabetology 06 00056 g005
Diabetology 06 00056 g006
Figure 6. Triglyceride (TG) levels recorded in mice under different treatments for 36 days (6 weeks). Data is expressed as mean ± standard error of the mean (SEM). * denotes p < 0.05; comparison of diabetic + J. secunda vs. control. t denotes p < 0.05; comparison of diabetic vs. all treatments from week 2 of administration (two-way repeated measures ANOVA and Student—Newman—Keuls tests). n = 6 mice per group.
Figure 6. Triglyceride (TG) levels recorded in mice under different treatments for 36 days (6 weeks). Data is expressed as mean ± standard error of the mean (SEM). * denotes p < 0.05; comparison of diabetic + J. secunda vs. control. t denotes p < 0.05; comparison of diabetic vs. all treatments from week 2 of administration (two-way repeated measures ANOVA and Student—Newman—Keuls tests). n = 6 mice per group.
Diabetology 06 00056 g006
Table 1. Secondary metabolites detected by qualitative phytochemistry in the fresh leaves of Justicia secunda Vahl.
Table 1. Secondary metabolites detected by qualitative phytochemistry in the fresh leaves of Justicia secunda Vahl.
Secondary
Metabolite		ReactionAcid ExtractEthanolic ExtractAqueous Extract
AlkaloidsDragendorff+
Mayer+
FlavonoidsShinoda (flavones) +
10% NaOH (flavonols)+
SterolsLiebermann Buchard +
TanninsGelatin reagent +
1% FeCl3 (phenolic compounds)+
+ Presence.
Table 2. Number of erythrocytes with micronuclei in mice under different treatments (mean ± SEM).
Table 2. Number of erythrocytes with micronuclei in mice under different treatments (mean ± SEM).
DaysControlAnthracene (10 mg/kg)Anthracene +
J. secunda (500 mg/kg)
01.75 ± 0.14.7± 0.74.5 ± 0.9
12.0 ± 0.212.2 ± 1.512.7 ± 2.7
22.25 ± 0.269.0 ± 1.010.0 ± 1.3
32.75 ± 0.112.0 ± 1.99.0 ± 2.6
42.5 ± 0.1110.7 ± 1.911.8 ± 1.6
52.25 ± 0.121.0 ± 2.715.5 ± 2.3
62.4 ± 0.3219.2 ± 2.914.0 ± 3.4
Table 3. Body weight of groups of mice under different treatments over 6 weeks (mean ± SEM).
Table 3. Body weight of groups of mice under different treatments over 6 weeks (mean ± SEM).
WeeksControlDiabetic (STZ, 120 mg/kg)Diabetic +
J. secunda (500 mg/kg)		Diabetic + J. secunda
+ Acarbose (300 mg/kg)
030.3 ± 1.930.2 ± 0.831.1 ± 1.6529.8 ± 0.9
130.3 ± 2.126.2 ± 1.131.1 ± 1.6329.4 ± 1.4
234.21 ± 1.525.5 ± 1.1129.1 ± 1.9828.8 ± 1.5
333.5 ± 1.522.8 ± 0.829.0 ± 1.629.0 ± 1.5
433.7 ± 1.325.7 ± 1.529.3 ± 1.6629.9 ± 1.3
536.3 ± 1.324.8 ± 2.027.8 ± 1.9230.0 ± 1.3
636.0 ± 1.219.3 ± 1.5 29.7 ± 2.2128.1 ± 1.3
Table 4. Blood glucose levels of mice under different treatments over 6 weeks (mean ± SEM).
Table 4. Blood glucose levels of mice under different treatments over 6 weeks (mean ± SEM).
WeeksControlDiabetic (STZ, 120 mg/kg)Diabetic +
J. secunda (500 mg/kg)		Diabetic + J. secunda
+ Acarbose (300 mg/kg)
0108.5 ± 8.678.5 ± 6.1112.1 ± 9.383.9 ± 8.1
1105.2 ± 13.8420.81 ± 30.2317.9 ± 43.8352.3 ± 39
2122 ± 3.4308.2 ± 47.8226.3 ± 61.2304.3 ± 60.5
3105.7 ± 9.4377 ± 21.5128.9 ± 25.1364 ± 38.7
470.7 ± 4.5463.5 ± 18.8166.9 ± 35.2360.1 ± 35.8
5109.3 ± 5.4437.7 ± 17.5121.4 ± 16.3339.5 ± 35.6
6101 ± 5.7494.7 ± 5.3 233.6 ± 37.8292.3 ± 56.1
Table 5. Blood triglyceride levels of mice under different treatments over 6 weeks (mean ± SEM).
Table 5. Blood triglyceride levels of mice under different treatments over 6 weeks (mean ± SEM).
WeeksControlDiabetic (STZ, 120 mg/kg)Diabetic +
J. secunda (500 mg/kg)		Diabetic + J. secunda
+ Acarbose (300 mg/kg)
0133.2 ± 9.6120 ± 6.1108.6 ± 9.9118 ± 10.2
1109.3 ± 11.9180.2 ± 30.2107.6 ± 15.3161 ± 6.2
2132 ± 9.7201.8 ± 30.883.6 ± 12.3166.4 ± 6.5
3135.7 ± 8.4196 ± 27.888.5 ± 12.5146.9 ± 10.3
4114.3 ± 9.8209.2 ± 30.688.9 ± 6.9164 ± 8.2
583.3 ± 17.9200.5 ± 25.786.1 ± 11.1162.4 ± 11.3
6126.2 ± 18.6212.4 ± 12.1 95.6 ± 10.8149.1 ± 12.4
Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content.

Share and Cite

MDPI and ACS Style

Fregoso-Aguilar, T.; Escamilla-Ramírez, P.X.; Nicolás-Álvarez, D.E.; Mendoza-Pérez, J.A.; Madrigal-Santillán, E.O.; Tirado-Lule, J.M.; Rodríguez-Negrete, E.V.; Madrigal-Bujaidar, E.; Álvarez-González, I.; Ibáñez-Cervantes, G.; et al. Effect of Chronic Administration of Justicia secunda Vahl in Mice Diabetized with Streptozotocin. Diabetology 2025, 6, 56. https://doi.org/10.3390/diabetology6060056

AMA Style

Fregoso-Aguilar T, Escamilla-Ramírez PX, Nicolás-Álvarez DE, Mendoza-Pérez JA, Madrigal-Santillán EO, Tirado-Lule JM, Rodríguez-Negrete EV, Madrigal-Bujaidar E, Álvarez-González I, Ibáñez-Cervantes G, et al. Effect of Chronic Administration of Justicia secunda Vahl in Mice Diabetized with Streptozotocin. Diabetology. 2025; 6(6):56. https://doi.org/10.3390/diabetology6060056

Chicago/Turabian Style

Fregoso-Aguilar, Tomás, Perla Xóchitl Escamilla-Ramírez, Dulce E. Nicolás-Álvarez, Jorge Alberto Mendoza-Pérez, Eduardo Osiris Madrigal-Santillán, Judith Margarita Tirado-Lule, Elda Victoria Rodríguez-Negrete, Eduardo Madrigal-Bujaidar, Isela Álvarez-González, Gabriela Ibáñez-Cervantes, and et al. 2025. "Effect of Chronic Administration of Justicia secunda Vahl in Mice Diabetized with Streptozotocin" Diabetology 6, no. 6: 56. https://doi.org/10.3390/diabetology6060056

APA Style

Fregoso-Aguilar, T., Escamilla-Ramírez, P. X., Nicolás-Álvarez, D. E., Mendoza-Pérez, J. A., Madrigal-Santillán, E. O., Tirado-Lule, J. M., Rodríguez-Negrete, E. V., Madrigal-Bujaidar, E., Álvarez-González, I., Ibáñez-Cervantes, G., Morales-González, Á., & Morales-Gonzalez, J. A. (2025). Effect of Chronic Administration of Justicia secunda Vahl in Mice Diabetized with Streptozotocin. Diabetology, 6(6), 56. https://doi.org/10.3390/diabetology6060056

Article Metrics

Back to TopTop