TECNOCIENCIA CHIHUAHUA, Vol. XVII (2) e 1221 (2023)

https://vocero.uach.mx/index.php/tecnociencia

ISSN-e: 2683-3360

Artículo Científico

Inhibitory effects of Buddleja scordioides (salvilla)

leaves on digestive enzymes and carbohydrate

absorption in vivo

Efectos inhibidores de las hojas de Buddleja scordioides (salvilla) sobre

las enzimas digestivas y la absorción de carbohidratos in vivo

*Correspondencia: nrocha@itdurango.edu.mx (Nuria E. Rocha-Guzmán), clau140382@hotmail.com (Claudia I. Gamboa-

Gómez2)

DOI: https://doi.org/10.54167/tch.v17i2.1221

Recibido:: 31 de mayo de 2023; Aceptado: 15 de agosto de 2023

Publicado por la Universidad Autónoma de Chihuahua, a través de la Dirección de Investigación y Posgrado

Abstract

The effects of Buddleja scordioides (BsLI) leaf infusions on digestive enzymes and carbohydrate

absorption were evaluated. The BsLI yield was 21.64 %. In addition, a chemical characterization was

carried out identifying hydroxybenzoic acids, hydroxycinnamic acids, flavonols, flavanones and

flavones. In vitro studies were performed to determine the inhibitory action of BsLI on lipase, α-

amylase, and α-glucosidase. Then, in rats, oral starch tolerance tests (OSTT) were carried out using

BsLI at a dose of 9.5 mg/kg body weight. Results showed moderate inhibition of lipase and α-

glucosidase, but greater inhibition of α-amylase compared to positive controls. During the OSTT

trial, the group receiving BsLI showed a significant reduction in glucose levels compared to the

negative control group. Bioactive compounds, such as naringenin, luteolin, quercetin, and coumaric

acid, were identified after BsLI administration. Furthermore, the consumption of BsLI was safe and

showed antioxidant activity like Trolox. In conclusion, BsLI may have an enhanced effect on glucose

metabolism by inhibiting carbohydrate absorption.

Laura J. Barragan-Zuñiga2, Luis E. Simental-Mendía2, Mayra Denise Herrera3, Rubén F.

González-Laredo1, J. Alberto Gallegos-Infante1, José Salas-Pacheco4, Martha R. Moreno-

Jiménez1, Nuria E. Rocha-Guzmán1*, Claudia I. Gamboa-Gómez2 *

1 Instituto Tecnológico de Durango. Blvd. Felipe Pescador 1830, Nueva Vizcaya, 34080 Durango, Dgo.

2 Unidad de Investigación Biomédica del Instituto Mexicano del Seguro Social. Predio Canoas, 34077

Durango, Durango.

3 Instituto Nacional de Investigaciones Forestales Agrícolas y Pecuarias. Campo Experimental Zacatecas

Kilómetro 24.5, Zacatecas - Fresnillo, Zacatecas.

4 Instituto de Investigación Científica, Universidad Juárez del Estado de Durango. Universidad y Fanny

Anitua SN, Zona Centro, 34000 Durango, Durango.

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TECNOCIENCIA CHIHUAHUA, Vol. XVII (2) e 1221 (2023)

Keywords: medicinal plant, carbohydrate absorption, antioxidant, Buddleja scordioides

Resumen

Se evaluaron los efectos de las infusiones de hojas de Buddleja scordioides (BsLI) sobre enzimas

digestivas y la absorción de carbohidratos. El rendimiento de BsLI fue del 21.64 %. Además, se

caracterizaron componentes químicos: ácidos hidroxibenzoicos, ácidos hidroxicinámicos,

flavonoles, flavanonas y flavonas. En estudios in vitro, se examinó cómo BsLI inhibe la lipasa, α-

amilasa y α-glucosidasa. Luego, en ratas, se probó su efecto en la tolerancia oral al almidón (OSTT)

a dosis de 9.5 mg/kg de peso corporal. Resultados indicaron moderada inhibición de lipasa y α-

glucosidasa, y mayor inhibición de α-amilasa comparado con controles. Durante la OSTT, el grupo

con BsLI tuvo menor glucosa que el control negativo. Tras administrar BsLI, se detectaron

compuestos bioactivos: naringenina, luteolina, quercetina y ácido cumárico. Además, BsLI fue

seguro, con actividad antioxidante similar al Trolox. En conclusión, BsLI puede tener un efecto

beneficioso sobre el metabolismo de la glucosa al inhibir la absorción de carbohidratos.

Palabras clave: planta medicinal, absorción de carbohidratos, antioxidante, Buddleja scordioides

1. Introduction

The western lifestyle is characterized by a lack of physical activity and hypercaloric diets. These

are the main contributing factors to the development of metabolic diseases such as obesity and type

2 diabetes (Kopp, 2019). A therapeutic target of these diseases is the inhibition of digestive enzymes

such as pancreatic lipase, α-amylase, and α-glucosidase (Patil et al., 2015; Irondi et al., 2018).

These digestive enzymes break down dietary lipids and carbohydrates to produce absorbable

molecules such as free fatty acids and monosaccharides. Thus, the inhibition of these enzymes

reduces the amounts of calories absorbed into the body and postprandial glucose (Awosika et al.,

2019). Pharmacological drugs such as acarbose and orlistat inhibit digestive enzymes involved in

glucose and lipid metabolism. However, they often exhibit side effects ranging from diarrhea to

hepatotoxicity, which limit their use in the clinical setting (Lunagariya et al., 2014). In this regard,

previous studies reported that different bioactive compounds (e.g. polyphenols) present in medicinal

plants inhibit one or more digestive enzymes with lesser adverse effects than the drugs

(Ardeshirlarijani et al., 2019; Gutiérrez-Grijalva et al., 2019).

Several species of Buddleja spp. are recognized to treat ills related to inflammatory processes (Estrada-

Zúñiga et al., 2019). Additionally, Hwang et al., (2009) found that B. officinalis improves

hyperglycemia, endothelium-dependent vascular relaxation, and inflammation in a diabetic

atherosclerotic mouse model. Previous experimental studies have revealed the diverse biological

activities of Buddleja scordioides, commonly known as salvilla. In a study published in 2002,

VanderJagt et al. demonstrated the anti-inflammatory potential of salvilla. Their findings indicated

that the plant extract could effectively inhibit the production of inflammatory cytokines in rats,

suggesting its potential for reducing inflammation in conditions like arthritis, asthma, and

inflammatory bowel disease. Rocha-Guzmán et al. conducted research in 2018 to explore the

gastroprotective effects of salvilla. Their study revealed that the extract provided protective benefits

to the stomach lining. Furthermore, a study by Villegas-Novoa et al. in 2020 reported the antioxidant

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effects of Buddleja scordioides extract in rats. The research demonstrated that the plant's extract

exhibited antioxidant properties, protecting cells from damage caused by free radicals. These

observed benefits have been associated with the bioactive compounds found of this herbal product,

which include iridoids, phenylpropanoids, sesquiterpenes, saponins, verbascosides, and flavonoids.

These compounds have shown a variety of beneficial effects, including antioxidant and anti-

inflammatory properties, as reported by Santos-Cruz et al. (2012) and Gutiérrez-Rebolledo et al.

(2019).

However, to the best of our knowledge, there is no evidence investigating the impact of B. scordioides

on digestive enzymes and carbohydrate absorption in vivo. Therefore, the aim of this study was to

evaluate the in vitro inhibitory effects of BsLI on digestive enzymes and carbohydrate absorption in

vivo. In addition, the oral BsLI-bioactive-compounds absorption (OBCA), antioxidant activity, and

safety of this medicinal plant were investigated.

2. Materials and methods

2.1 Materials

Samples of Buddleja scordioides Kunth (salvilla) were collected at Ignacio Ramirez road in

Durango, México (coordinates 24°30´17.41” N, length 104°4´51.23” W, elevation 2030 m average).

The botanist Dr. Socorro González-Elizondo taxonomically identified leaves and the voucher

specimens (47538) were deposited at the Herbarium of the Centro Interdisciplinario de Investigación

para el Desarrollo Integral Regional del Instituto Politécnico Nacional, Unidad Durango, México.

The leaves were air dried in the shade at 25 C followed by milling to a particle size of 0.7-1.0 mm.

2.2 Preparation of Buddleja scordioides leaves infusions (BsLI)

The infusion concentration was commonly used by the general population (1 % m/v). The dried

leaves sample (2 g) was added to 200 mL boiling water and kept stirring for 10 min. Infusions were

obtained by subsequent filtration and lyophilized (FreeZone 18 Liter Console Freeze Dry System,

Kansas USA). Samples were stored in amber vessels until use.

2.3 Yields of extractable solids

Infusion yield was determined as follows:

Yield (%)=[(Lyophilized infusion (g))/(Dried plant material (g)) ]100 Eq (1)

Results are reported as means of two independent infusion preparation.

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2.4 Chemical characterization of BsL

Sample analysis was carried out with an Acquity UPLC system (Waters Corp., Milford)

coupled with a tandem Xevo TQ-S triple quadrupole mass spectrometer (Waters Corp., Wexford).

The LC system consisted of a sample manager (5 C) and a quaternary solvent manager. The column

used to determine the phenolic profile was an Acquity UPLC BEH C8, 1.7 µm, 2.1 mm x 100 mm

(Waters Corp., Wexford), operated at 30 C. The mobile phase included two solvents: acidified Milli

Q water with 7.5 mM formic acid (Solvent A) and acetonitrile LC-MS (Solvent B): Initial – 5 % B, 0.8

min isocratic to 5 % B, 1.2 min gradient to 10 % B; 1.9 min isocratic 10 % B; 2.4 min gradient to 15 %

B; 3.7 min isocratic 15 % B; 4.0 min gradient to 21 % B; 5.2 min isocratic 21 % B; 5.7 min gradient to

reach 27 % B; 8.0 min gradient to reach 50 % B; 9.0 (linear gradient) for column washing 100 % B;

subsequent at 11.5 min linear gradient 5 % B since 13.5 min for column stabilization at a flow rate of

250 µL/min. Electron spray ionization (ESI) in negative mode was as follows: capillary voltage 2.5

kV, desolvation temperature 300 C, source temperature 150 C, desolvation gas flow 500 L/h, and

cone gas flow 150 L/h, collision gas flow was 0.14 mL/min, MS mode collision energy 5.0 and MS/MS

mode collision energy 20.0. For identification and quantification of the phenolic profile, a mixture of

standards (20 ng/µL) was used for monitoring retention times, m/z values, and MS/MS transitions.

Samples and standards were monitoring at multiple modes. The UPLC and Tandem Xevo TQ-S

triple quadrupole mass spectrometer control and data processing was using MassLynx v. 4.1

Software (Waters Corp.).

2.5 Inhibitory effects of BsLI on digestive enzymes

2.5.1 Pancreatic lipase inhibition assay

The assay was performed following the methodology reported by McDougall et al. (2009),

with some modifications. In brief, 150 μL of a solution (10 mg/mL) of porcine pancreatic lipase type

II (Sigma- Aldrich, Cat. No. L3126) was mixed with BsLI (at 10, 50, 100, 200, 400, 600, 800, and 1000

µg/µL), 400 μL of Tris buffer (100 mM, pH 8.2), 400 µL of p-nitrophenyl laurate (SIGMA Co., St.

Louis, USA), and substrate solution (0.08 % w/v dissolved in 5 mM sodium acetate at pH 5.0

containing 1 % Triton X-100). Samples were incubated at 37 °C for 2 h and centrifuged at 16,000 RPM,

for 3 min. Finally, the supernatant was read at 400 nm using a microplate reader (MultiScan Go,

Thermo Scientific, USA). The positive control was Orlistat (Redustat® Laboratorios Liomont S.A. DE

C.V.).

The lipase inhibitory activity was expressed as a percentage of inhibition:

Inhibition %=100⦋S_0-S_1)/S_(0)⦌ Eq (2)

Where S_0 is the absorbance of the blank and S_1 is the absorbance of BsLI.

Additionally, BsLI concentration that provided 50 % inhibition (IC50) was calculated by plotting the

inhibition percentage versus the log concentration curve (Coruh et al., 2007).

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2.5.2  -Amylase inhibition assay

The assay was performed following the methodology reported by Tamil et al. (2010) with

several modifications. Substrate solution was performed using starch (4 %), CaCl2 0.01 M, and

distilled water (5 mL) for 5 min at 96 °C. The BsLI (at 10, 50, 100, 200, 400, 600, 800, and 1000 µg/µL)

were incubated for 1 h at 37 °C with 100 µL of porcine pancreas α-amylase solution (2.5 mU/µL in

sodium phosphate buffer (0.02 M, pH 6.9) and sodium chloride (6 mM)) (SIGMA Co., St. Louis, USA).

Afterward, to stop the reaction, NaOH (4 mL, 0.1 M) was added, and a centrifugation step was

performed (700 xg for 5 min). Glucose concentration in the supernatant was determined using an

enzymatic kit (Biosystems Instruments Reagents, Barcelona, Spain). Acarbose (Laboratorios

Alpharma, CDMX, México) was used as a positive control. Results were expressed as inhibition

percentage (equation 2) and IC50.

2.5.3  -Glycosidase inhibition assessment

The assay was performed following the methodology reported by Apostolidis et al. (2007).

Sample of BsLI (25 μL at 20, 40, 60, 80, and 100 µg/µL), 100 µL of glucosidase enzyme (SIGMA Co.,

St. Louis, USA) (0.19 mU/µL), and 50 μL of phosphate buffer (0.1 M, pH 6.9) were mixed and

incubated for 10 min at 37 °C. Then, 25 µL of p-nitrophenyl-α-D-glucopyranoside solution (5 mM/L

prepared in a 0.1 M/L of citrate–phosphate buffer, pH 7) (SIGMA Co., St. Louis, USA) was added

and incubated 30 min at 37 °C. The reaction was stopped by adding 1 mL of a 0.05 mol/L NaOH

solution. Samples were read at 410 nm with the Synergy HT Microplate Reader (Biotek Instruments,

Winooski, Vermont, USA.). Acarbose (Laboratorios Alpharma, CDMX, México) was used as a

positive control. Results were expressed as inhibition percentage (equation 2) and IC50.

2.6 Antioxidant activity of BsLI in vitro.

2.6.1 Determination of radical scavenging activity using the ABTS assay

The ABTS assay was performed following the methodology reported by Re et al. (1999). In

brief, potassium persulfate (2.5 mM) was mixed and incubated in dark for 16 h at 25±1 °C with ABTS

(7 mM) to produce the radical cations. The BsLI (2.5 µL at 10, 50, 150, 200, 250, 300, 350, and 400

μg/mL) were added to the ABTS solution (200 µL). Trolox was the positive control. Afterward, the

absorbance was recorded, and the results were expressed as the inhibition percentage (equation 2)

and as IC50.

2.6.2 Radical scavenging assay: Diphenylpicrylhydrazyl (DPPH)

The DPPH assay was performed following the methodology reported by Brand-Williams et

al. (1995). One mL of the stable free radical DPPH solution (20 mg/L) was mixed with 200 µL of BsLI

(at concentration of 10, 50, 100, 200, 400, 500, 600, 700 and 800 μg/mL). Samples were incubated for

30 min in dark at room temperature (26 C).The absorbance was registred at 515 nm. Trolox was the

positive control. The results were expressed as the inhibition percentage (equation 2) and as IC50.

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2.7 Effect of BsLI on carbohydrate absorption in vivo.

2.7.1 Animals/ethics approval

Animal experiments were carried out in accordance with the Mexican guidelines

(NOM-062-ZOO-1999) and the National Institutes of Health (2002) recommendations for research

with animals.

Animals used in the experiments were male Wistar rats (ten weeks old, n = 8, 180 ± 20 g of body

weight) and male CD-1 mice (eight weeks old, n=28, 25 ± 5 g of body weight), as well as non-pregnant

female CD-1 mice (eight weeks old, n=6, 25 ± 5 g of body weight). All animals were obtained from

the same source, Universidad Nacional Autónoma de México (UNAM), campus Juriquilla,

Querétaro, México.

They were housed in a controlled environment with a 12–12 h light–dark cycle and maintained at a

temperature of 27 ± 1 C. Water and food (Rodent Lab Chow 5001, Purina®, Québec, Canada) were

provided ad libitum for all the animals during the experiments.

2.7.2 Oral starch tolerance test (OSTT)

Effects of BsLI on carbohydrate absorption were assessed with an oral starch tolerance test

(OSTT). Starch was the negative control, whereas the positive control was acarbose.

In fasting conditions, male Wistar rats (ten weeks old, n = 8, 180 ± 20 g of body weight) were given a

starch load at a dose of 3 g/kg of body weight (equivalent to 519.26 mg contained in an infusion cup

of 240 mL consumed by an adult of 70 kg by a meal). Additionally, 5.12 mg/Kg of body weight of

acarbose was used as a positive control (equivalent to 50 mg by a meal in humans). Blood glucose

levels were determined in blood samples collected from the tail vein using a glucometer (Stat Strip®

Glucose, Nova Biomedical, Waltham, MA, US) at 0, 30, 60, and 120 min. The variations in serum

glucose onset time, peak, and AUC were considered to the determination of the relative rate of

carbohydrate digestion and absorption.

The human dose extrapolation (HED) to the animal was estimated according to the following

formula:

HED = animal dose (mg/kg) [animal weight (kg/human weight in kg)] 0.33

Reagan-Shaw et al. (2008)

Eq (3)

2.8 Oral absorption assay of BsLI bioactive compounds (OABC)

The oral absorption assay was conducted on male CD-1 mice (eight weeks old, n=28, 25 ± 5

g of body weight). The mice were placed in metabolic cages and fasted for 12 hours before the

experiment. Treatments (n=4 per time point) were administered through gavage at a dose of 2500

mg/kg of body weight to the intervention groups at various time intervals (0, 0.5, 2, 4, 8, 12, and 24

hours). The negative control group received water as the vehicle. After each treatment, the mice were

anesthetized and euthanized by cardiac puncture. Serum was separated from the blood and

immediately frozen at -80 C for later analysis.

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The analysis of the serum samples involved adding 100 µL of serum to a mixture containing 37 μL

of water with 200 µg/µL of ascorbic acid and 1 µg/µL of EDTA. The mixture was then centrifuged at

14000 g for 10 minutes, and 250 μL of ethyl acetate was added for liquid-liquid micro-extraction. The

suspension was mixed using a vortex for 1 minute. The collected volumes were dried using a

vacuum centrifugal console, reconstituted in methanol (200 µL), and filtered through 0.45 µm

syringe filters. The identification and quantification of BsLI bioactive compounds present in mouse

serum were performed following the methodology previously described in section 2.4 on "the

chemical characterization of BsLI".

2.9 Toxicity assessment of BsLI

2.9.1 Acute toxicity

The acute toxicity assay was carried out on non-pregnant female CD-1 mice (eight weeks

old, n=6, 25 ± 5 g of body weight) (Universidad Nacional Autónoma de México, campus Juriquilla,

Querétaro, México). The guidelines of the Organization for Economic Cooperation and Development

for testing of chemicals 420 (OECD, 2001) were followed. The Mice were divided into two groups of

three animals each. A single dose of BsLI (5000 mg/kg of body weight) was administered via gavage

(in distilled water). Mice were observed for signs of possible toxicity (convulsions, tremors, lethargy,

salivation, diarrhea, sleepiness, and coma) every hour for the first four hours, and then the animals

were fed. Thereafter, mice were monitored for any signs of toxicity (changes in the eyes, skin, fur,

mucous membranes, and the respiratory, circulatory, and autonomic nervous and central nervous

systems) and mortality for 14 days. Measurements of body weight and food and water intake were

performed daily. On the last day, animals were sacrificed under deep ether anesthesia and the

median lethal dose (LD50) values were estimated. Vital organs such as the heart, kidney, liver,

spleen, and lung were isolated and weighted to assess histopathological changes. The organs were

fixed in 10 % buffered neutral formalin, embedded in paraffin wax, cut (5 µm) on glass slides, and

stained with hematoxylin and eosin. The slides were examined under a light microscope.

2.9.2 Sub-chronic toxicity

For the sub-chronic toxicity assessment, both female and male mice aged eight weeks and

weighing 25 ± 5 g of body weight (UNAM, Campus Juriquilla, Querétaro, México) were used.

The assay was performed according to the methodology described by the Organization for Economic

Co-operation and Development-407 (OECD, 2008). Animals were divided into four groups of 10 each

(five males and five females). The control group received distilled water (vehicle), while the three

intervention groups received BsLI by gavage at doses of 250, 750, and 2500 mg/kg of body weight,

respectively, for 28 days. Mice were observed for signs of abnormalities during the treatment period.

Measurements of body weight and food and water intake were performed daily.

Before sacrifice, urine was collected and analyzed using urinalysis strips (Bio-Uridiag-A10) for the

measurement of leucocytes, nitrite, urobilinogen, protein, blood, specific gravity, ketone, bilirubin,

and glucose. At the end of the treatment period, mice were sacrificed, and blood samples were

obtained via cardiac puncture into non-heparinized and ethylenediaminetetraacetic acid (EDTA)

containing tubes for biochemical and hematological analyses.

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Heart, kidney, liver, spleen, and lung were isolated to assess histopathological changes. These organs

were excised, weighed (Scout PRO Ohaus, Mississauga, Ontario, Canada), examined

macroscopically, and fixed in 10 % buffered neutral formalin. Fixed organs were processed for

paraffin embedding and cuts of 5 mm thick were obtained by microtome and then processed using

an alcohol xylene series for being later stained with hematoxylin and eosin.

Hematological parameters such as red blood cell (RBC), mean corpuscular volume (MCV), mean

corpuscular hemoglobin (MCH), mean corpuscular hemoglobin concentration (MCHC), and white

blood cell (WBC) were analyzed using an automatic hematology analyzer (Hemat Technology,

Newton, Massachusetts, USA).

Serum biochemical parameters such as glucose, total cholesterol, triglycerides, high-density

lipoprotein cholesterol (HDL-c), alanine aminotransferase (ALT), aspartate aminotransferase (AST),

albumin, total protein, and total bilirubin were analyzed using Biosystem commercial test kits with

an automated A15 spectrophotometer (Biosystems Instruments Reagents, Barcelona, Spain). The

concentrations of low-density lipoprotein cholesterol (LDL-c) were estimated using the formula by

Friedewald et al., (1972):

LDL-c (mg/dL) = [total cholesterol (TC) (mg/dL) – HDL (mg/dL) - (triglycerides

(TG) (mg/dL)/5)] Eq (4)

2.10 Statistical analysis

Data were expressed as mean values ± standard error (SE). Statistical significance was

determined by one-way variance analysis (ANOVA) (p < 0.05) followed by Tukey´s test, Statistical

analysis was conducted using the Sigma Plot software version 13.0 (Systat Software, Inc., San Jose,

CA, USA).

3. Results and discussion

3.1 Yield and chemical characterization

The yield of BsLI was 21.64 %. Regarding chemical characterization results, hydroxybenzoic

acids, hydroxycinnamic acids, flavonols, flavanones, and flavones were identified and quantified.

The protocatechuic acid, salicylic acid, vanillic acid, and quinic acid were the phenolic acids with the

highest concentration; whereas, quercetin, quercetin 3-O-glucoside, and luteolin were the main

flavonol compounds detected (Table 1).

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Table 1. Chemical characterization of Buddleja scordioides leaves infusion (BsLI)(1 %).

Tabla 1. Caracterización química de la infusión de hojas de Buddleja scordioides (BsLI)(1 %).

No.

Compound

(min)

[M-H]-

m/z

Transitions

Content

[ng/mg of crude

extract]

Hydroxybenzoic acids

Gallic acid

1.64

169

125, 79

8.529  0.73

Protocatechuic acid

3.13

153

109, 91

258.914  9.73

2,5-dihydroxybenzoic acid

4.21

153

109, 81

183.945  0.27

4-hydroxybenzoic acid

4.41

137

93, 65

106.790  2.60

Vanillic acid

4.85

167

152, 123

138.336  1.13

Tri-hydroxybenzaldehyde

6.21

153

16.584  1.33

Salicylic acid

7.70

137

93, 65

401.480  43.62

Hydroxycinnamic acids

Quinic acid

1.01

191

93, 85

210.486  24.05

Chlorogenic acid

4.32

353

191, 85

27.11  2.95

10.

4-O-caffeoylquinic acid

4.51

353

353, 179

13.717  0.48

11.

Caffeic acid

4.85

179

135

93.348  6.94

12.

Coumaric acid

6.19

163

119, 98

47.224  10.48

13.

Ferulic acid

6.54

193

178, 134

51.178  1.08

14.

4,5-dicaffeoylquinic acid

7.14

515

353, 179

3.446  0.47

15.

Rosmarinic acid

7.73

359

197, 161

36.189  0.02

Flavonols

16.

Rutin

6.25

609

300, 271

69.162  1.15

17.

Quercetin 3-O-glucoside

6.48

463

300, 271

433.894  1.48

18.

Quercetin 3-O-ß-glucuronide

6.54

477

301, 151

164.200  9.09

19.

Quercetin

8.41

301

179, 151

466.554  2.06

Flavanones

20.

Neohesperidin

7.77

609

301, 164

48.328  1.00

21.

Eriodictyol

8.42

287

151, 135

5.225  0.90

Flavones

22.

Luteolin

8.40

285

151, 133

231.853  4.55

23.

Apigenin

8.80

269

148, 117

13.022  1.36

24.

9.80

283

268, 211

21.021  1.30

Values are means of duplicate determinations ± standard error.

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3.2 Inhibition of digestive enzymes

3.2.1 Lipase inhibition

Results of lipase assessment are depicted in Fig. 1. The BsLI showed a lower percentage of

lipase inhibition (~44%) than orlistat (~92 %). Additionally, the IC50 values were significantly higher

as compared with the positive control (16-fold) (Table 2).

Figure 1. Lipase inhibition of Buddleja scordioides leaves infusion (BsLI). Orlistat was the positive control.

Different letters in the graph indicate significant differences in the percentage of lipase inhibition. Values are

means of duplicate determinations ± standard error. (p ˂ 0.05) by Tukey's test.

Figura 1. Inhibición de la lipasa de la infusión de hojas de Buddleja scordioides (BsLI). Orlistat fue el control

positivo. Las letras diferentes en el gráfico indican diferencias significativas en el porcentaje de inhibición de la

lipasa. Los valores son medias de determinaciones duplicadas ± error estándar. (p ˂ 0.05) mediante la prueba

de Tukey.

3.2.2 -amylase inhibition

The amylase results are shown in Fig. 2. The BsLI had a higher percentage of α-amylase

inhibition than acarbose, achieving a ~96 % while acarbose had a ~78 % inhibition. On other hand,

no differences were observed for the IC50 values between BsLI and positive control (acarbose) (Table

2).

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Table 2. Inhibitory digestive enzymes and antioxidant assessment of Buddleja scordioides leaves infusion (BsLI)

IC50 values.

Tabla 2. Inhibición de enzimas digestivas y evaluación antioxidante de la infusión de hojas de Buddleja

scordioides (BsLI) Valores IC50.

Figure 2. α-amylase inhibition (1) and α-glucosidase inhibition (2) of Buddleja scordioides leaves infusion (BsLI).

Acarbose was the positive control. Values are means of duplicate determinations ± standard error. a,b Different

letters indicate statistical differences between BsLI and positive control (p ˂ 0.05) by Tukey's test.

Figura 2. Inhibición de α-amilasa (1) e inhibición de α-glucosidasa (2) de la infusión de hojas de Buddleja

scordioides (BsLI). La acarbosa fue el control positivo. Los valores son medias de determinaciones duplicadas ±

error estándar. a,b Las letras distintas indican diferencias estadísticas entre BsLI y el control positivo (p ˂ 0.05)

mediante la prueba de Tukey.

Orlistat

(µg/µL)

Acarbose

(µg/µL)

Trolox

(µg/µL)

BsLI

(µg/µL)

Lipase

0.34 ± 0.01a

---

5.50 ± 0.01b

-amylase

---

1.69 ± 0.81a

---

1.83 ± 0.01a

-glucosidase

---

1.64 ± 0.04a

---

7.60 ± 0.10b

ABTS

---

1.93 ± 0.01a

1.48 ± 0.01a

DPPH

---

2.11 ± 0.01a

2.05 ± 0.01a

Results are expressed as inhibitory median concentration (IC50) values. Values are means of

duplicate determinations ± standard error. a-c Different letters indicate statistical differences

(p ˂ 0.05) by Tukey's test.

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3.2.3 α -glucosidase inhibition

Fig. 2 shows the results of α-glucosidase inhibition. The BsLI had a lower percentage of α-

glucosidase inhibition (~19 %) than acarbose (~94 %). Regarding IC50 values, BsLI was 4-fold higher

than the positive control (Table 2).

3.4 Oral starch tolerance test (OSTT)

Variations in serum-TG onset time and area-under-time-curve (AUC) are depicted in Fig. 3.

Both groups treated with acarbose and BsLI showed significantly reduced glucose levels at 60 (above

12 %) and 120 min (above 14 %) in comparison with the control group.

Figure 3. Oral starch tolerance test (OSTT) (1) and Area-under-time-curve (AUC) (2) in rats treated with Buddleja

scordioides leaves infusion (BsLI). Acarbose was used as apositive control. Values are means of duplicate

determinations ± standard error. (*) indicates statistical differences between study groups, whereas for AUC a,b

different letters indicate statistical differences between groups (p ˂ 0.05) by Tukey's test.

Figura 3. Prueba de tolerancia oral al almidón (OSTT) Prueba oral de tolerancia al almidón (OSTT) (1) y área

bajo la curva de tiempo (AUC) (2) en ratas tratadas con infusión de hojas de Buddleja scordioides (BsLI). La

acarbosa se utilizó como control positivo. Los valores son medias de determinaciones duplicadas ± error

estándar. (*) indica diferencias estadísticas entre los grupos de estudio, mientras que para AUC a,b letras

diferentes indican diferencias estadísticas entre los grupos (p ˂ 0,05) por la prueba de Tukey.

3.5 Oral BsLI-bioactive-compounds absorption evaluation (OBAC)

Polyphenols detected in the serum of mice are shown in Fig. 4. Bioactive compounds such

as 4-hydroxybenzoic acid, naringenin, eriodictyol, luteolin, quercetin, and coumaric acid were

identified in serum mice at different times after BsLI administration. The coumaric acid (13.84 ng/µL)

and luteolin (16.01 ng/µL) exhibited the highest concentration; nevertheless, these levels declined

rapidly.

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Figure 4. Inhibition percentage of Buddleja scordioides leaves infusion (BsLI) on ABTS (1) and DPPH (2)

assessment. Trolox was the positive control. Values are means of duplicate determinations ± standard error. a,b

Different letters indicate statistical differences between BsLI and positive control (p ˂ 0.05) by Tukey's test.

Figura 4. Porcentaje de inhibición de la infusión de hojas de Buddleja scordioides (BsLI) en la evaluación ABTS (1)

y DPPH (2). Trolox fue el control positivo. Los valores son medias de determinaciones duplicadas ± error

estándar. a,b Las letras distintas indican diferencias estadísticas entre BsLI y el control positivo (p ˂ 0.05)

mediante la prueba de Tukey.

3.6 Antioxidant activity of BsLI in vitro.

3.6.1 ABTS and DPPH

Antioxidant evaluation of the BsLI by ABTS and DPPH assays is shown in Fig. 5. On one

hand, ABTS results, showed that the BsLI had inhibition percentage like Trolox. On other hand, BsLI

exhibited a lower inhibition percentage (~70 %) of DPPH radical compared with Trolox (~91 %).

There were no significant differences for IC50 between the treatments for both, ABTS and DPPH

assessment (Table 2).

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Figure 5. Means of total polyphenols of Buddleja scordioides leaves infusion (BsLI) in mice serum: (1)

hydroxybenzoic acids, (2) hydroxycinnamic acids, (3) flavones y flavonols, and (4) flavanones. Values are

expressed as mean ± standard error.

Figura 5. Medias de polifenoles totales de la infusión de hojas de Buddleja scordioides (BsLI) en suero de

ratones: (1) ácidos hidroxibenzoicos, (2) ácidos hidroxicinámicos, (3) flavonas y flavonoles, y (4) flavanonas. Los

valores se expresan como media ± error estándar.

3.7 Acute and sub-chronic toxicity evaluation

A single oral dose of infusion of BsLI (5000 mg/kg of body weight) produced no signs of

toxicity (tremors, convulsions, salivation, diarrhea, lethargy, sleepiness, or coma) or mortality after

14 days. No significant difference in body weight gain was observed. No gross pathological

abnormalities were observed in both groups. Furthermore, histological analysis of isolated organs

showed no abnormal changes (data not shown). Thus, the LD50 value was found to be greater than

5000 mg/kg.

Regarding sub-chronic toxicity evaluation, there were no significant differences between the study

groups for body weight, water, and food consumption (data not shown).

Macroscopic analysis of liver, lung, spleen, kidney, and heart exhibited no significant changes in

color and texture compared with the control group. Additionally, intervention with BsLI samples at

different doses did not affect the weight of organs.

Sub-chronic oral administration BsLI samples at different doses had no significant differences in the

urinary biochemical parameters compared with the control group (data not shown). Moreover,

treatment with BsLI showed no significant changes in serum concentrations of glucose, liver

enzymes, total bilirubin, total protein content, albumin, total cholesterol, HDL-c, LDL-c, and

triglycerides in comparison with the controls (Table 3).

Sub-chronic oral administration BsLI samples at different doses exhibited no significant changes on

hematological profile compared with the control group (Table 4).

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Histological analysis of heart, kidney, liver, spleen, and lung of intervention groups treated with

different doses of BsLI (250, 750 y 2500 mg/kg of body weight) revealed no abnormal changes

compared with the control group (data not shown).

Our results showed that BsLI decreases carbohydrate absorption in vivo, which could be explained

by the inhibition of digestive enzymes involved in glucose metabolism. Also, we found an

antioxidant activity in these herbal infusions.

Although the BsLI exhibited a lower percentage of lipase inhibition than orlistat, this effect could

reduce the absorption of lipids in vivo. However further studies to examine the impact of BsLI on

lipid metabolism are required.

Regarding carbohydrate digestive enzymes, BsLI showed higher inhibition of α-amylase than

acarbose. This effect could be attributed to bioactive compounds present in BsLI such as polyphenols.

It has been reported that the hydroxyl groups of these compounds interact with the enzyme active

site, allowing the formation of ligands with the catalytic residues of the binding site and,

consequently the generation of a conjugated system, resulting in the inactivation of the enzyme

(Narita and Inouye, 2009).

On other hand, BsLI had a minimal inhibitory effect on α-glucosidase. This lack of effect could be

because we employed p-nitrophenyl glucoside as substrate and α-glucosidase from a yeast,

Saccharomyces cerevisiae. This test only indicates a general glucosidase activity (Williamson, 2013);

therefore, further investigation to determine the effect of BsLI on α-glucosidase inhibition using

other substrates such as sucrose and enzyme from other sources (e.g. porcine small intestine) is

mandatory.

Our results showed that the BsLI decreases the absorption of starch similarly to acarbose. We

attribute this effect to the inhibitory activity of BsLI on α-amylase. Given that the hydrolysis of the

polysaccharide to produce limited dextrins, maltose, and maltotriose is catalyzed by α-amylase,

inhibition of both salivary and pancreatic α-amylase decreases meal-derived carbohydrate

absorption (Robyt et al., 2008).

In the present study, we detected phenolic compounds (quercetin, rutin, luteolin, and phenolic acids)

with well-established biological activity (Deng et al., 2020; Sok et al., 2021), which could be

responsible for the beneficial effects of BsLI. However, it is important to consider that the biological

action of polyphenols depends on their absorption and interaction with target tissues (Silberberg et

al., 2006). In this regard, we analyzed the absorption of bioactive compounds of BsLI in serum mice.

The main metabolites identified in BsiLI were phenolic acids and flavonoids. Additionally, the

plasma levels of these compounds suggest relatively rapid absorption, which is partly consistent

with previous studies (Liu et al., 2002).

Also, it has been reported that the bioactive compounds contained in this herbal infusion have

antioxidant effects (Moretti et al., 2012; Tian et al., 2021). In this line, the results of our study

demonstrated that BsLI exhibits antioxidant properties by scavenging free radicals (ABTS and

DPPH). It has been suggested that an ideal antidiabetic agent should have antioxidant activity (Mai

et al., 2007); thus, in addition to the hypoglycemic action, the BsLI could emerge as a therapeutic

alternative for diabetes management.

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The toxicological activity of medicinal plants is an important aspect that must be assessed before

being recommended as alternative therapies (Nasri, 2013). In this context, we evaluate the acute and

chronic toxicological effects of BsLI and our results revealed no signs of toxicity, macroscopic

abnormalities, or mortality, suggesting non-toxic effects of these herbal infusions. Usually, the

consumption of infusions in Mexican communities is one cup per day at 1 % (m/v) concentration

(equivalent to 33 mg/kg of body weight). Interestingly, although we used higher doses (7.5 [250

mg/kg], 22.7 [750 mg/kg] and 75 [2500 mg/kg] times), none of these caused signs of toxicity, damage

symptoms or mortality. Additionally, all hematological and biochemical parameters remained

within the reference range for CD1 mice (Titlow, 2013) and no significant differences were observed

between the study groups. Particularly, it is noteworthy that after treatment with BsLI we found no

significant differences in transaminase levels or abnormal changes in liver tissue by histological

examination, suggesting that this natural agent does not induce hepatocellular damage. Also,

macroscopic and microscopic evaluation of the organs from mice receiving oral administration of

BsLI for 28 days at different doses (250, 750, and 2500 mg/kg) revealed normal architecture indicating

no detrimental changes or morphological disorders in the sub-chronic toxicity assessment. Hence,

the no-observed-adverse-effect level (NOAEL) may be considered for this herbal product. The BsLI

could be considered a promising and attractive strategyin the treatment of chronic diseases such as

obesity and type II diabetes; however, future clinical studies investigating the effects of these herbal

infusions are needed to confirm our findings.

Table 3. Liver function indices of CD-1 mice administrated orally with Buddleja scordioides leaves infusion (BsLI)

for 28 consecutive days.

Tabla 3. Índices de función hepática de ratones CD-1 administrados por vía oral con infusión de hojas de

Buddleja scordioides (BsLI) durante 28 días consecutivos.

Control group

250

(mg/kg)

750

(mg/kg)

2500

(mg/kg)

AST (IU/L)

193.00 ± 6.20 a

184.55 ± 11.46 a

180.78 ± 13.87 a

193.52 ± 19.53 a

ALT (IU/L)

65.47 ± 1.85 a

59.75 ± 4.17 a

49.56 ± 3.16 a

49.05 ± 5.70 a

Total Protein (g/dL)

4.99 ± 0.06 a

5.05 ± 0.06 a

5.31 ± 0.05 a

5.07 ± 0.10 a

Albumin (g/dL)

2.53 ± 0.28 a

1.61 ± 0.07 a

1.96 ± 0.07 a

1.86 ± 0.08 a

Total bilirubin

(mg/dL)

0.14 ± 0.09 a

0.12 ± 0.09 a

0.15 ± 0.35 a

0.28 ± 0.18 a

Total cholesterol

(mg/dL)

104.12 ± 1.89 a

99.14 ± 3.25 a

101.71 ± 2.87 a

98.43 ± 1.47 a

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ALT: alanine aminotransferase, AST: aspartate aminotransferase, HDL-c: high density lipoprotein, and LDL-

c: low density lipoprotein. Values are expressed as mean ± standard error. Different letters indicate

statistically significant differences between groups *p ≤ 0.05 significantly different from the control were

determined by using one way ANOVA followed by Tukey’s multiple comparison tests.

Table 4. Effect on hematological parameters of oral administration with infusions of Buddleja scordioides leaves

infusion (BsLI) for 28 consecutive days.

Tabla 4. Efecto sobre los parámetros hematológicos de la administración oral con infusión de hojas de Buddleja

scordioides (BsLI) durante 28 días consecutivos.

RBC: red blood cell, MCV: mean corpuscular volume, MCH: mean corpuscular hemoglobin, MCHC: mean

corpuscular hemoglobin concentration, WBC: white blood cell. Values are expressed as mean ± standard

HDL-c (mg/dL)

58.16 ± 1.64 a

62.18 ± 2.23 b

59.03 ± 3.29 a

58.26 ± 1.53 a

LDL-c (mg/dL)

38.85 ± 2.31 a

24.22 ± 2.20 b

31.46 ± 1.82 a

30.73 ± 2.09 a

Triglyceride (mg/dL)

59.86 ± 3.13 a

69.86 ± 2.68 a

68.50 ± 3.83 a

77.2 ± 3.23 b

Control group

250

(mg/kg)

750

(mg/kg)

2500

(mg/kg)

RBC (×106/µL)

9.02 ± 0.19 a

8.31 ± 0.26 b

9.56 ± 0.14 a

8.67 ± 0.17 b

Hemoglobin (g/dL)

17.24 ± 0.37 b

16.51 ± 3.28 a

18.45 ± 0.34 a

16.88 ± 0.29 a

Hematocrit (%)

46.52 ± 0.98 b

44.13 ± 1.26 a

49.62 ± 0.90 a

44.77 ± 0.79 a

MCV (fL)

51.56 ± 0.07 a

53.26 ± 0.29 a

51.73 ± 0.33 a

51.671 ± 0.29 a

MCH (pg)

19.10 ± 0.04 a

19.93 ± 0.08 a

19.24 ± 0.15 a

19.50 ± 0.11 a

MCHC (g/dL)

37.03 ± 0.06 a

37.44 ± 0.13 a

37.18 ± 0.08 a

37.73 ± 0.13 a

Platelets (×103/µL)

1171.66 ± 4.77 a

727.85 ± 4.91 b

1047.33 ± 3.97 a

978.28 ± 4.41 a

WBC (×103/µL)

8.26 ± 2.91 a

5.68 ± 2.71 a

6.26 ± 3.03 a

5.20 ± 2.54 a

Neutrophils (%)

11.00 ± 0.51 b

9.71 ± 0.75 b

14.11 ± 0.58 a

11.86 ± 0.57 a

Eosinophils (%)

0.55 ± 0.11 a

0.57 ± 0.16 a

0.00 ± 0.00 b

0.57 ± 0.16 a

Basophils (%)

0.00 ± 0.00 c

0.14 ± 0.05 a

0.11 ± 0.04 b

0.14 ± 0.05 a

Lymphocytes (%)

87.22 ± 0.49 a

86.71 ± 0.93 a

83.56 ± 0.59 b

85.71 ± 0.43 a

Monocytes (%)

1.22 ± 0.12 c

2.86 ± 0.30 a

2.22 ± 0.90 a

1.71 ± 0.15 a

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error. Different letters indicate statistically significant differences between groups *p ≤ 0.05 significantly

different from the control were determined by using one way ANOVA followed by Tukey’s multiple

comparison tests.

4. Conclusion

The study demonstrates that the infusion of Buddleja scordioides (BsLI) exerts inhibitory

effects on carbohydrate absorption in vivo by targeting digestive enzymes involved in glucose

metabolism. Moreover, BsLI exhibits antioxidant properties and contains bioactive compounds,

including phenolic acids and flavonoids. The infusion proves to be safe, with no indications of

toxicity or adverse effects on organ histology. These findings suggest that BsLI holds promise as a

potential therapeutic alternative for managing conditions such as obesity and type II diabetes.

However, further clinical studies are necessary to validate these results and explore the full potential

of BsLI as a treatment option.

Acknowledgments

This project received support from two grants: TecNM-5557.15-P and Conacyt-INFR, 2015-

253333. The author LJBZ is grateful for the graduate scholarship 1165903777 provided by the

National Council of Humanities, Sciences and Technologies (CONAHCYT). Additionally, we would

like to express our gratitude to Alejo Macías-Salas for his valuable technical support in the

histopathologic analysis.

Conflict of Interest statement

The authors declare no conflicts of interest.

5. References

Apostolidis, E., Kwon, Y. I. & Shetty, K. 2007. Inhibitory potential of herb, fruit, and fungal-enriched

cheese against key enzymes linked to type 2 diabetes and hypertension. Innov. Food Sci.

Emerg .Technol. 8(1): 46-54. https://doi.org/10.1016/j.ifset.2006.06.001

Ardeshirlarijani, E., Namazi, N., B Jalili, R., Saeedi, M., Imanparast, S., Adhami, H. R., Faramarzi,

M.A., Ayati, M.H., Mahdavi, M. & Larijani, B. 2019. Potential Anti-obesity effects of some

medicinal herb: In vitro α-amylase, α-glucosidase and lipase inhibitory activity. Int. J. Biol.

Biomed. 5(2): 2-8. http://ibbj.org/article-1-228-en.html

Awosika, T. O. & Aluko, R.E. 2019. Inhibition of the in vitro activities of α‐amylase, α‐glucosidase

and pancreatic lipase by yellow field pea (Pisum sativum L.) protein hydrolysates. Int. J. Food

Sci. 54(6): 2021-2034. https://doi.org/10.1111/ijfs.14087

Barragán-Zúñiga et.al

TECNOCIENCIA CHIHUAHUA, Vol. XVII (2) e 1221 (2023)

Brand-Williams, W., Cuvelier, M.E., & Berset, C. 1995. Use of a free radical method to evaluate

antioxidant activity. LWT - Food Science and Technology 28(1): 25-30.

https://doi.org/10.1016/S0023-6438(95)80008-5

Coruh, N., Celep, A.S. & Özgökçe, F. 2007. Antioxidant properties of Prangos ferulacea (L.) Lindl.,

Chaerophyllum macropodum Boiss. and Heracleum persicum Desf. from Apiaceae family used

as food in Eastern Anatolia and their inhibitory effects on glutathione-S-transferase. Food

Chem. 100(3): 1237-1242. https://doi.org/10.1016/j.foodchem.2005.12.006

Deng, N., Zheng, B., Li, T. & Liu, R.H. 2020. Assessment of the phenolic profiles, hypoglycemic

activity, and molecular mechanism of different highland barley (Hordeum vulgare L.)

varieties. Int. J. Mol. Sci. 21(4): 1175. https://doi.org/10.3390/ijms21041175

Estrada-Zúñiga, M.E., Gutiérrez-Rebolledo, G.A., Nieto-Trujillo, A., Bernabé-Antonio, A.& Sosa, F.C.

2019. Buddleja species distributed in Mexico against inflammatory diseases, their therapeutic

activities, secondary metabolites and biotechnology. Adv. Biol. Res. 5: 78-98.

https://stm1.bookpi.org/index.php/rabr-v5/article/view/375

Friedewald, W. T., Levy, R.I. & Fredrickson, D.S. 1972. Estimation of the concentration of low-density

lipoprotein cholesterol in plasma, without use of the preparative ultracentrifuge. Clin. Chem.

18(6): 499-502. https://doi.org/10.1093/clinchem/18.6.499

Gutiérrez-Grijalva, E. P., Antunes-Ricardo, M., Acosta-Estrada, B. A., Gutiérrez-Uribe, J.A. &

Heredia, J.B. 2019. Cellular antioxidant activity and in vitro inhibition of α-glucosidase, α-

amylase and pancreatic lipase of oregano polyphenols under simulated gastrointestinal

digestion. Int. Food Res. J. 116: 676-686. https://doi.org/10.1016/j.foodres.2018.08.096

Gutiérrez-Rebolledo, G.A., Estrada-Zúñiga, M. E., Garduño-Siciliano, L., García-Gutiérrez, G. E.,

Mora, C.A.R., Calderón-Amador, J. & Cruz-Sosa, F. 2019. In vivo anti-arthritic effect and

repeated dose toxicity of standardized methanolic extracts of Buddleja cordata Kunth

(Scrophulariaceae) wild plant leaves and cell culture. J. Ethnopharmacol. 240: 111875.

https://doi.org/10.1016/j.jep.2019.111875

Hwang, S.M., Lee, Y.J., Kim, E.J., Kim, H.Y., Li, X., Choi, Y.J., Cho, N.G., Lee, H.S. & Kang, D.G. 2009.

Effect of Buddleja officinalis in Diabetic Atherosclerotic Mouse Model Using High Fat Diet.

Korea J. Herbol. 24(4): 55-62. https://doi.org/10.6116/kjh.2009.24.4.055

Irondi, E.A., Angola, S.O. & Obligor, I.E. 2018. Inhibitory effects of tropical almond leaf extract on

xanthenes oxidize, pancreatic lipase, and angiotensin 1-converting enzyme, in vitro. J. Food

Biochem. 42(4): e12481. https://doi.org/10.1111/jfbc.12481

Kopp, W. 2019. How western diet and lifestyle drive the pandemic of obesity and civilization

diseases. Diabetes Metab. Syndr. Obes.: Targets Ther. 12: 2221-2236.

https://doi.org/10.2147/DMSO.S216791

Liu, Y. & Hu, M. 2002. Absorption and metabolism of flavonoids in the caco-2 cell culture model and

a perused rat intestinal model. Drug Metabolism and Disposition 30(4): 370-377.

https://doi.org/10.1124/dmd.30.4.370

Lunagariya, N.A., Patel, N.K., Jagtap, S.C. & Bhutani, K.K. 2014. Inhibitors of pancreatic lipase: state

of the art and clinical perspectives. EXCLI J. 13: 897 -921.

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4464291/

Barragán-Zúñiga et.al

TECNOCIENCIA CHIHUAHUA, Vol. XVII (2) e 1221 (2023)

Mai, T.T., Thu, N.N., Tien, P.G. & Van Chuyen, N. 2007. Alpha-glucosidase inhibitory and

antioxidant activities of Vietnamese edible plants and their relationships with polyphenol

contents. J. Nutr. Sci. Vitaminol. 53(3): 267-276. https://doi.org/10.3177/jnsv.53.267

McDougall, G.J., Kulkarni, N.N. & Stewart, D. 2009. Berry polyphenols inhibit pancreatic lipase

activity in vitro. Food Chemistry 115(1): 193-199.

https://doi.org/10.1016/j.foodchem.2008.11.093

Moretti, E., Mazzi, L., Terzuoli, G., Bonechi, C., Iacoponi, F., Martini, S., Rossi, C. & Collodel, G. 2012.

Effect of quercetin, rutin, naringenin and epicatechin on lipid peroxidation induced in

human sperm. Reprod. Toxicol. 34, 651-657. https://doi.org/10.1016/j.reprotox.2012.10.002

Narita, Y. & Inouye, K. 2009. Kinetic analysis and mechanism on the inhibition of chlorogenic acid

and its components against porcine pancreas α-amylase isozymes I and II. J. Agric. Food

Chem. 57(19): 9218-9225. https://doi.org/10.1021/jf9017383

Nasri H. & Shirzad H. 2013. Toxicity and safety of medicinal plants. J HerbMed. Plarmacol. 2(2): 21-

22.

https://www.researchgate.net/publication/285306673_Toxicity_and_safety_of_medicinal_pl

ants

National Institutes of Health (US). 2002. Office of Laboratory Animal Welfare, United States. Public

Health Service. Public Health Service policy on humane care and use of laboratory animals.

Office of Laboratory Animal Welfare, National Institutes of Health, Department of Health

and Human Services.

https://grants.nih.gov/grants/olaw/references/phspolicylabanimals.pdf

Norma Oficial Mexicana NOM-062-ZOO-1999. 1999. Especificaciones técnicas para la producción,

cuidado y uso de los animales de laboratorio.

https://www.gob.mx/cms/uploads/attachment/file/203498/NOM-062-ZOO-1999_220801.pdf

OECD. 2001. Guidelines for Acute Toxicity of Chemicals. Organization for Economic Co-operation

and Development No. 420. Published online Paris, France.

OECD. 2008. Guidelines for Repeated Dose 28-day Oral Toxicity Study in Rodents Organization for

Economic Co-operation and Development No. 407. Published online Paris, France.

Patil, P., Mandal, S., Tomar, S.K. Anand, S. 2015. Food protein-derived bioactive peptides in

management of type 2 diabetes. Eur. J. Nutr. 54, 863–880. https://doi.org/10.1007/s00394-

015-0974-2

Re, R., Pellegrini, N., Proteggente, A., Pannala, A., Yang, M., Rice-Evans, C. 1999. Antioxidant activity

applying an improved ABTS radical cation decolorization assay. Free Radic. Biol. Med. 26,

1231-1237. https://doi.org/10.1016/S0891-5849(98)00315-3

Reagan‐Shaw, S., Nihal, M., Ahmad, N. 2008. Dose translation from animal to human studies

revisited. FASEB J. 22, 659-661. https://doi.org/10.1096/fj.07-9574LSF

Robyt, J.F. 2008. in: Fraser-Ried, B. O., Tatsuta, K., Thiem, J., Cote´, G.L., (Eds.), Glycoscience,

Springer-Verlag, Berlin, Heidelberg, Germany; 1437–1472.

Rocha-Guzmán, N. E., Simental-Mendía, L. E., Barragán-Zúñiga, L. J., Ramírez-España, J. C.,

Gallegos-Infante, J. A., Lujan-Mendoza, C. I., Gamboa-Gómez, C. I. (2018). Effect of Buddleja

Barragán-Zúñiga et.al

TECNOCIENCIA CHIHUAHUA, Vol. XVII (2) e 1221 (2023)

scordioides K. leaves infusion on lipid peroxidation in mice with ultraviolet light-induced

oxidative stress. J. Med. Chem. 27, 2379-2385. https://doi.org/10.1007/s00044-018-2243-4

Santos-Cruz, L.F., Ávila-Acevedo, J.G., Ortega-Capitaine, D., Ojeda-Duplancher, J.C., Perdigón-

Moya, J.L., Hernández-Portilla, L.B., López-Dionicio, H., Drán-Díaz, A., Dueñas-Gracía, I. E.,

Castañeda-Partida, L., García-Bores, A. M., Heres-Pulido, M. E. 2012. Verbascoside is not

genotoxic in the ST and HB crosses of the Drosophila wing spot test, and its constituent,

caffeic acid, decreases the spontaneous mutation rate in the ST cross. Food Chem. Toxicol.,

50(3-4), 1082-1090. https://doi.org/10.1016/j.fct.2011.12.006

Silberberg, M., Besson, C., Manach, C., Remesy, C., Morand, C. 2006. Influence of dietary antioxidants

on polyphenol intestinal absorption and metabolism in rats. J. Agric. Food Chem. 54, 3541-

3546. https://doi.org/10.1021/jf060104e

Sok Yen, F., Shu Qin, C., Tan Shi Xuan, S., Jia Ying, P., Yi Le, H., Darmarajan, T., Gunasekaran, B.,

Salvamani, S. 2021. Hypoglycemic Effects of Plant Flavonoids: A Review. Evid. Based

Complement. Alternat Med. 2021. https://doi.org/10.1155/2021/2057333

Tamil, I.G., Dineshkumar, B., Nandhakumar, M., Senthilkumar, M., Mitra, A. 2010. In vitro study on

α-amylase inhibitory activity of an Indian medicinal plant, Phyllanthus amarus. Indian J.

Pharmacol. 42, 280. https://doi.org/10.4103/0253-7613.70107

Tian, C., Liu, X., Chang, Y., Wang, R., Lv, T., Cui, C., Liu, M. 2021. Investigation of the anti-

inflammatory and antioxidant activities of luteolin, kaempferol, apigenin and quercetin. S.

Afr. J. Bot. 137, 257-264. https://doi.org/10.1016/j.sajb.2020.10.022

Titlow WB. 2013. Characterization of Toxicological Properties of L-Lysine Polymers in CD-1 Mice. J.

Microbiol. Biotechnol. 23, 1015-1022. https://doi.org/10.4014/jmb.1302.02055

VanderJagt, T.J., Ghattas, R., VanderJagt, D.J., Crossey, M., Glew, R.H. 2002, Comparison of the total

antioxidant content of 30 widely used medicinal plants of New Mexico. Life Sci. 70, 1035-

1040. https://doi.org/10.1016/S0024-3205(01)01481-3

Villegas-Novoa, C., Moreno-Jiménez, M.R., Rocha-Guzmán, N.E. 2020. Infusión de la planta

medicinal Buddleja scordioides Kunth utilizada para tratar la inflamación intestinal.

CienciaUAT. 14, 21. https://doi.org/10.29059/cienciauat.v14i2.1287

Williamson, G. 2013. Possible effects of dietary polyphenols on sugar absorption and digestion. Mol.

Nutr. Food Res. 57, 48-57. https://doi.org/10.1002/mnfr.201200511

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