Effect of prebiotics of Agave salmiana fed to healthy Wistar rats

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eEffect of prebiotics of Agave salmiana fed to healthy Wistar rats

I. Jasso-Padilla a, B. Juárez-Flores b, G. Alvarez-Fuentes b, A. De la Cruz-Martínez a, J.

González-Ramírez a, M. Moscosa-Santillán a, M. González-Chávez a, C. Oros-Ovalle c,

F. Prell d, P. Czermak d, e, F. Martinez-Gutierrez a*.

a Facultad de Ciencias Quimicas, Universidad Autonoma de San Luis Potosi, Mexico.

b Instituto de Investigacion de Zonas Deserticas, Universidad Autonoma de San Luis

Potosi, Mexico.

c Departamento de Patologia, Hospital Central, Dr. Ignacio Morones Prieto, San Luis

Potosi, Mexico.

d Justus Liebig University Giessen, Institute of Food Chemistry and Food

Biotechnology, Giessen, Germany

e University of Applied Sciences Mittelhessen, Institute of Bioprocess Engineering

and Pharmaceutical Technology, Giessen, Germany.

*Corresponding author: Fidel Martinez Gutierrez, PhD.

Laboratorio de Microbiologia. Facultad de Ciencias Quimicas, Universidad Autonoma

de San Luis Potosi. Av. Dr. Nava #6 Zona Universitaria. San Luis Potosi, SLP, Mexico

78210. Phone: +52 (444) 826 24 40 Ext 6591. E-mail: [email protected]

This article is protected by copyright. All rights reserved.

This article has been accepted for publication and undergone full peer review but has not been through the copyediting, typesetting, pagination and proofreading process, which may lead to differences between this version and the Version of Record. Please cite this article as doi: 10.1002/jsffa.7764

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eAbstract

BACKGROUND: Inulin and other fructans are synthesized and stored in mezcal agave

(Agave salmiana). Fructans provide several health benefits and have excellent

technological properties, but only few data report their physiological effect when added

in the diet. RESULTS: Here, we studied the physiological effects of fructans obtained

from A. salmiana when added in the diet of Wistar rats. Results evidence favorable

changes on Wistar rats when the fructans was added to their diet, including the decrease

of the pH in the faeces and the increase of the number of lactic acid bacteria (CFU g-1)

(Lactobacillus spp. and Bifidobacterium spp.), even these changes were enhanced with

the synbiotic diet (fructans plus B. animalis spp. lactis). Synbiotic diet, developed

changes in the reduction of cholesterol and triglycerides concentrations in serum, with

statistical differences (p<0.05). Histological analysis of colon sections showed that

synbiotic diet promoted colon cells growth suggesting that fructans from A. salmiana

confer beneficial health effects through gut microbiota modulation. CONCLUSION:

Our data underline the advantage of targeting the gut microbiota by colonic nutrients

like specific structure of fructans from A. salmiana, with their beneficial effects. More

studies are necessary to define the role of fructans to develop more solid therapeutic

solutions in humans.

Keywords: fructans, lactic acid bacteria, cholesterol, triglycerides, colon cells.


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eIntroduction

Mesoamerica supports a rich native flora that comprises many arid and semi-arid

species useful to humans for food, medicine, fibre and structural materials. This area

has been an important centre of plant domestication, ranging from prehistoric times

when crops such as maize and Agave were first domesticated, to the present when many

native plants are in various stages of domestication within traditional agricultural

systems.1 Agave species have been highly appreciated in Mexico since the pre-Hispanic

era. They have long been used by Mexican people as a source of food, forage, fibers,

poultices for wounds.2 Actually Agave species are exploited for the production of

alcoholic nondistilled and distilled beverages with national and international

recognition,3 however, everyday more plants are dedicated to high fructose syrup and

agave fructans production as ingredients for healthier food and feed.4 Agavaceae is

considerate between the seven families with major crassulacean acid metabolism

(CAM),5 some cultivated CAM plants (e.g., Agave mapisaga and Agave salmiana) may

achieve an average ground productivity of 4 kg dry mass m-2yr-1.5 Previous studies

about A. salmiana characterization showed that the qualitative and quantitative

composition of nonstructural carbohydrates depended on plant organ and ripeness.6

Recently a study at Charcas, San Luis Potosi, Mexico showed the yield of fructans from

A. salmiana of fresh stems was 210 g kg-1 and almost 800 g kg-1 from dry stems.7

Beyond the standard use of Agave plants, some evidence show other ways for their

applications, since Agave plants can grow in marginal arid and semiarid lands, where

their special ecological and physiological adaptations to environmental conditions give

them the potential to produce substantial biomass,8 which represents an opportunity in

order to stimulate sustainable economic growth in developing countries.9 Diverse

fructans are considered prebiotics, because they are not digestible through intestinal

tract enzymes and whole molecules pass into the colon, where these polymers become

fermented through beneficial intestinal microbiota, also known as probiotics, within

which are lactic acid bacteria (LAB), the composition of the gut microbiome could

modulate the induction of regulatory versus effector immune responses and improve

health outcomes.10 The health benefits of fructans from Agave species have been

recognized, in vitro study, extracts from A. salmiana showed enhanced bacterial growth

and immune system activators,11 in the same hand, in vivo study with obese rats fed

with Agave angustifolia ssp. and different sources of fructans showed influence in the

body weight and blood metabolites concentrations,12 both results can be related to an


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eimprovement of gut bacterial functions implicated in the development of the systemic

immunity and have an important influence on the host nutrition and metabolism.13 Even

more enteroendocrine cells of the gut secrete a variety of metabolically related low

molecular compounds such as short-chain fatty acids (SCFA),14 furthermore, gut

bacterial functions can influence whole-body metabolism by affecting the energy

balance,15 gut permeability,16 serum lipopolysaccharides and metabolic inflammation17

that are associated with obesity and associated disorders because their capacity of

decreased low-density lipoprotein cholesterol.18

Few reports of the physiological effect of fructans obtained from A. salmiana added in

the diet have been presented, the goal of this research was studied the combination of

the A. salmiana compounds like prebiotic looking for the specific stimulation of the

probiotic strain, as well as, their compounds combination with probiotic to formed a

synbiotic19 were assessed and their effects on animal model using different diets, on the

basis of (i) substrate composition, (ii) pH shifts, (iii) evolution of selected LAB

populations (iv) concentration of biochemical parameters and fatty acids and (v) colon

histological evaluation.

Experimental

Plant Material and Dehydration

The harvest of the agave pineapple of A. salmiana was conducted in the area of Laguna

Seca mezcal factory, which is located in the town of Charcas, San Luis Potosi, Mexico.

The pineapple maguey of A. salmiana had a state of maturity in which the maguey

contained a high concentration of fructans, this occurs immediately before the plant

emits the scape, which in turn stem emerges from the center of the maguey, presenting

only flowers at the apex.20 The stem of the pineapple head was removed, then the stem

was cut into 1.5 cm thick cubes and placed in the international fruit centrifugal extractor


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eof juice (Model EXS Series 6/18/02 Volt.). Samples of dried extract of stem of A.

salmiana were obtained by lyophilization (Free Dry System, model FreeZone 6. Brand:

Labconco Corporation, Kansas City, Missouri).

Identification of FOS

Carbohydrate analysis was performed with high performance anion-exchange

chromatography with pulsed amperometric detection (HPAEC-PAD) on a Dionex ICS-

3000 (Thermo Scientific, Waltham, Massachusetts, USA), equipped with a pulsed

electrochemical detector consisting of an amperometric flow-through cell, a gold

working electrode and a silver-silver chloride reference electrode. Elution of

carbohydrates was performed at 30° C on a Dionex Carbopac PA200 column (Thermo

Scientific, Waltham, Massachusetts, USA), at a flow rate of 0.3 mL min-1. Elution

program and solvent composition for both methods are summarized in Table 1 (adapted

from.21 Gradient elution was optimized by employing 160 mM NaOH (eluent A), 160

mM NaOH with 1 M sodium acetate (eluent B), and 1 M NaOH (eluent C). Data

processing was carried out with Chromeleon 6.80 software (Thermo Scientific,

Waltham, Massachusetts, USA).

As standards, glucose, fructose, sucrose, kestose, nystose and fructofuranosylnystose

were used in a concentration of 200 µM each. A sample of A. salmiana was dissolved

with distilled water (100 mg 10 mL-1) and filtered (membrane with 0.45 μm) before the

solution was injected.


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e

Table 1. Description of the optimized gradient.

Elution time

(min)

Phase A (%)

(160 mM NaOH)

Phase B (%)

(160 mM NaOH + 1

M sodium acetate)

Phase C (%)

(1M NaOH)

0.0 100 ND ND

0.0 100 ND ND

3.5 99.2 0.8 ND

33.5 81.1 18.9 ND

38.5 ND 100 ND

38.6 ND ND 100

45.5 ND ND 100

50.0 100 ND ND

60.0 100 ND ND


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eMicroorganism and encapsulation

Lyophilized reference strain of lactic acid bacteria (LAB), Bifidobacterium animalis

ssp. lactis (SACCO BLC1, RAFF S.A. de C.V. Zapopan, Jal, Mexico), was used. B.

animalis spp. lactis required two sequential reactivations of 24 h in of Man-Rogosa-

Sharpe broth (MRSB; Becton-Dickinson Difco, San Jose, CA, USA) supplemented with

0.5 g L-1 l-cysteine (Analytica, Monterey, NL, Mexico) under the same conditions. To

characterize the growth phases of a 24 h growth, curve was performed in a 1 L flask of

MRSB, and samples were collected at different time periods (0.5–1 h intervals).

Microbial growth was determined through plate counting using the Miles and Misra

method.22 The probiotic biomass was collected by centrifugation of broth at 4000 rpm

for 15 min at 4°C, the biomass was washed twice with phosphate-buffered saline (PBS),

under the same centrifugation conditions. Bacteria were encapsulated in commercial

low-fat milk (5 g kg-1) with 100 g kg-1 of solid content. The dilution proportion was 6 g

of wet probiotic biomass in 100 mL of milk. The solution was fed in a spray dryer (Mini

Spray Dryer B-290, Büchi, Switzerland) at 160°C, 100% aspiration and 25% pump

capacity (10 mL min-1). The spray drying yield was calculated from the total solids

content of the solution (milk-bacteria) fed to the dryer and the weight of the powder

collected at the end of each run drying. Viability tests were performed in MRS broth,

encapsulating solution and powders obtained by spray drying (microcapsules). The

percentage of survival of bacteria was obtained using the encapsulating solution and

powders viability, for each run drying. Such that the percentage of survival was

obtained from the following equations:

CFU contained in the encapsulating solution (CFU solution) = CFU mL-1 x 100 mL

Dried powders (CFU microcapsules) = CFU g-1 x g (dry powder)

% Survival = CFU (microcapsules) CFU(solution)-1 x 100


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eAnimals, diets and sampling

The animals used in this study were handled in accordance with the ethics

recommendations of the Mexican Official Standard: Technical Specifications for the

Breeding, Care and use of Laboratory Animals.23 The local University Ethics

Committee for Animal Research approved the experimental protocol used, with the

register number: CONBIOÉTICA24CEI00820131212. Four-week-old male Wistar rats

(weighing 140 to 160 g) were housed in polypropylene cages, covered with metallic

grids in a room maintained at 22 ± 2 °C, 55 ± 10% humidity and under a 12 h light–dark

cycle. After a 2-week acclimation period to the housing environment, the animals were

randomly distributed into three groups (5 rats each): Commercial group, was fed

commercial diet (Rodent Laboratory Chow 5001, Agribrands Purina, Mexico),

Commercial + FOS (Com + FOS) group received commercial diet added with dried

extract of A. salmiana at 20 g kg-1 and Commercial + Synbiotic (Com + SYN) group

received a commercial diet added with synbiotic formulation (dried extract of A.

salmiana at 20 g kg-1 plus 15 g kg-1 of powder of encapsulated Bifidobacterium

animalis spp. lactis at theoretical concentration of 2.5 1010 CFU g-1 diet), for 12 weeks.

The sampling of stool took place every 2 weeks, the recovery was held by manipulating

each animal model, which favored the defecation; one gram of faeces was collected in

an empty sterile tube. A volume of 800 μL of total blood was collected by venipuncture

of their tails. The blood samples were collected into test tubes and after clotting the

blood at room temperature, it was centrifuged (1500g, 4 °C, 10 min), then the serum

was collected and stored at -80 °C for further use. Both samples were carried out with

previous fast of 12 h. At the end of the 12th week, animals were anesthetized with

sodium pentobarbital (45 mg kg-1 body weight), the animals were killed by blood

extraction via carotid puncture. The colon was removed, opened longitudinally and


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egently rinsed with saline solution to remove residual bowel contents, finally was placed

on a previously labeled container with 10 mL of 50 mL L-1 formaldehyde for its

preserve.

Determination of pH and bacteria count on faeces

One gram of faeces was transferred into a sterile tube and mixed with 9 mL of sterile

saline solution, the pH was measured using a microelectrode (Hanna Instruments,

USA). For the bacteriological analysis, 9 mL of sterile saline solution containing 10 mL

L-1 of hydrochloride L-cysteine (Analytica, Monterey, NL, Mexico) were used and then

serially diluted (from 10-1 to 10-11). Lactobacillus spp. and Bifidobacterium spp., were

quantified using Man-Rogosa-Sharpe Agar supplemented with 0.5 g L-1 l-cysteine

(Analytica, Monterey, NL, Mexico). The Petri dishes were incubated at 37 °C under

anaerobic conditions using Gas-pack anaerobic jars. Microbial growth was determined

as CFU g-1 of LAB in faeces after 72 h incubation.

Biochemical parameters and fatty acids determination in serum

Glucose concentration was measured in the rat serum using an Autoanalyzer (RA-500,

Bayer, Spain). Diagnostic kits (Spinreact, Girona, Spain) were used for the

determination of triglycerides (TGL) and total cholesterol (TC) with a Beckman

Spectrophotometric equipment according to manufacturer’s specifications (Spinreact,

Girona, Spain).

Gas chromatography-mass spectrometry (GC-MS) analysis was carried out on a

chromatograph Agilent Technology model 6890N with a mass spectrometer model

5973 and under the following conditions: DB-5HT low bleed capillary column. The

injector temperature was kept isothermal at 250 °C; initial split conditions on: 0.01 min


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eoff and 5 min on with a split ratio 1:50; the oven was set at 100 °C for 2 min, and then

ramped at 15 °C till 165 °C for 3 min and again ramped at 15 °C min-1 till 180 °C; mass

detector in 71 eV (the m z-1 range was 33 to 800). Relative GC retention times were

obtained by comparison of authentic standard using pure standard mixtures (Aldrich,

USA).

Histological analysis

Tissue fragments were imbibed in paraffin and stained with haematoxylin and eosin

(H&E) for histological examination of the colon sections.

Experimental Design and Statistical Analyses

The experiment was conducted according to a fully randomized experimental design,

with a 2 x 3 factorial arrangement of treatments and five replicates per treatment.

Factors and levels were: (a) animal condition (healthy), and (b) diets (commercial,

commercial + fructans and commercial + Synbiotic) with five replicates. The

experimental period lasted 12 weeks, diets were provided once daily, at the beginning of

the darkness period. With the results of normality tests, ANOVA and multiple

comparisons of means (SAS Institute, Cary, North Carolina SAS, version 8.0) were

made. In the evaluation of final concentration of FFA, was performed using an analysis

of covariance, where concomitant variable was the basal concentration.

Results

Fructans in A. salmiana juice


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eThe early mature stem, without hydrolysis, of A. salmiana was used in this study.

Available standards were used to identify peaks corresponding to glucose, fructose,

sucrose, kestose, nystose, and fructofuranosylnystose on the basis of their retention

times, HPAEC chromatograms of the mixed standards (Figure 1A) and A. salmiana

(Figure 1B) are presented. The results obtained showed the presence of

monosaccharides glucose and fructose, as well as, sucrose, kestose (GF2), nystose

(GF3) and fructofuranosylnystose (GF4) were also identified, when their retention

times, HPAEC-PAD chromatograms of the mixed standards (Figure 1A) were

compared with the sample of A. salmiana (Figure 1B). Interesting peaks were detected

at the following retention times: 12.31, 14.69 and 16.57 minutes and different signals

were detected after 18 minutes of runtime.

Microorganism and encapsulation

The viability of B. animalis ssp. lactis and process survival percentage concerning 5

production batches (Table 2) showed that spray drying yield varies from 43 to 63% with

a mean value around 54.4%, those results are consistent with previous result reported in

literature, while the mean process survival of microencapsulated bacteria was about

3.5%. Furthermore, mean viability obtained in the final powders was 1.9 x 109 CFU g-1.

Since the FAO/WHO have recommended both principles, concentration up to 106 CFU

g-1 to be considered as a probiotic and the stability of them, an assessment of

microcapsules viability in storage for 15 days was carried out (Supplementary

information).


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e

Table 2. Microencapsulation results for Bifidobacterium animalis ssp. lactis bacteria by

spray drying.

Effects of the diets on pH and LAB population in faeces

The pH decreased continuously along the fermentation time in both groups where

powder from A. salmiana was added to the diets of the animals. The lowest pH values

were observed in the group fed with synbiotic food, statistical significance was

observed after the third sample (Figure 2). No changes in pH were observed in the

group with commercial food.

In this research, continuous increases in the number of Lactobacillus spp. and

Bifidobacterium spp. (CFU g-1) of both groups, where powder from A. salmiana was

added to the diets of the animals were archived. It is noteworthy that approximately 4-

Batch

Viability of Bifidobacterium animalis ssp. lactis Process

survival

(%)

Drying

yield (%) MRS Broth

CFU mL-1

Solution

CFU mL-1

Powders

CFU g-1

1 2.6 ± 0.37x108 1.2 ± 0.18x109 3.8 ± 0.66x108 1.99 ± 0.38 47.51

2 2.9 ± 0.51x108 2.9 ± 0.34x109 1.9 ± 0.21x109 3.46 ± 0.61 43.38

3 5.5 ± 0.76x108 4.6 ± 0.49x109 3.4 ± 0.31x109 4.39 ± 0.71 63.12

4 4.7 ± 0.28x108 4.0 ± 0.42x109 2.0 ± 0.08x109 3.19 ± 0.63 60.48

5 8.8 ± 0.14x108 2.7 ± 0.29x109 1.9 ± 0.34x109 4.62 ± 0.98 57.83


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elog increase was observed in the group with synbiotic food, statistical differences

(p<0.05) was observed after of the sixth sample when were compared with the group

where no powder from A. salmiana was added (Figure 3).

Effects on Biochemical and fatty acids determinations in serum

Here we evaluated the biochemical parameters of healthy Wistar rats, the statistical

analysis showed differences (p<0.05) when the rats were fed with synbiotic diet (Table

3). In this study, the concentration of glucose, cholesterol and triglycerides showed no

statistical differences (p>0.05) when the basal and final concentration were compared

with the commercial and fructans diets, however, when the rats were treated with

synbiotic diet, statistical differences (p<0.05) was observed in the reduction of

cholesterol and triglycerides serum concentrations.


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eTable 3. Effects of diets on biochemical parameters in serum of different Wistar rats

groups1.

1 Values are average + Standard error of the mean (SEM)

2Commercial= basal diet; Com + FOS= 20 g Kg-1 dried extract of A. salmiana plus the basal commercial

diet; Com + SYN= 15 g Kg-1 Bifidobacterium animalis ssp. lactis at 1.9 x 109 CFU g-1 plus 20 g Kg-1 dried

extract of A. salmiana plus basal commercial diet. All the diets were for 12 weeks. Data with different

superscript letters by row are statistically different (p < 0.05).

Group - diets2

Biochemical

data

Commercial

(mg dL-1)

Com + FOS

(mg dL-1)

Com + SYN

(mg dL-1)

BASAL FINAL SEM BASAL FINAL SEM BASAL FINAL SEM

Glucose 74.6 a 77.2 a 18.3 79.4 a 68.4 a 29.6 65.4 a 97.2 a 21.2

Cholesterol 68.4 a 46.0 a 7.7 53.4 a 53.8 a 8.7 74.2 b 47.0 b 10.9

Triglycerides 129.0

a

52.8 a 33.3 66.8 a 58.0 a 24.6 74.0 b 42.4 b 25.4


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eIn this research, the influences of the dietary on the concentration of individual FFA

contents in rat serum are shown in Table 4. It was observed that the concentrations of

palmitic acid, stearic acid, oleic acid and linolenic acid were reduced when the rats were

fed with FOS and synbiotic diets in compassion with the commercial diet. However, the

results showed no statistical differences (p>0.05) between rats fed with FOS and

synbiotic supplemented diet and those on the control diet.


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eTable 4. Effect of diets on fatty acids in serum of different Wistar rats groups1 obtained

by Gas chromatography-mass spectrometry.

Treatment Concentration

(ppm)

C16

Palmitic acid

C18

Stearic acid

C18-1

Oleic acid

C18-2

Linolenic

acid

Commercial 80.4 + 13.2 39.6 + 3.9 63.7 + 12.5 47.4 + 5.8

Com + FOS 79.1 + 13.2 32.9 + 4.1 58.9 + 13.2 34.5 + 5.7

Com + SYN 66.7 + 13.6 34.5 + 4.0 41.2 + 12.9 41.1 + 5.9

1 Values show the mean ± SD of 3 independent experiments.


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eHistological

The histological analysis of colon sections of the group fed with commercial food

(Figure 4A) showed the colon cell with their nucleus, which were increase in their

number in both groups fed with fructans from A. salmiana (Figure 4B and 4C). The

group fed with synbiotic diet showed statistical differences (p<0.05) in comparison with

the group fed with commercial diet. An increase of the number of colon cell in the

group fed with synbiotic diet was observed (Figure 4D).

Discussion

Our present study shows for the first time the effect of fructans from Agave salmiana

like a prominent prebiotic in healthy animal model. Therefore, according to the results

presented, it can be inferred that A. salmiana showed the presence of longer molecules.

The clear separation of all the molecules contained in the juice of A. salmiana was not

possible, because of the complex structure of Agave branched fructans. The same result

was obtained in the comparison of the distribution of fructans from chicory inulin

versus A. tequilana by HPAEC-PAD.24 Thus, in this study HPAEC-PAD was more

useful in the analysis of low molecular weight carbohydrate.

The viability of B. animalis ssp. lactis was 54.4%, this value is low compared with

some values reported in literature, which often exceed 60%.25 The low survival value is

probably due to the carrier agent used (commercial milk).

This study confirms and extends previous reports suggesting a beneficial effect of

prebiotics in the diet, the effects of acidify of the bowel, which is considered as a major

beneficial feature,26 it support the theory of the capacity of suppresses the growth of


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epathogens.27 The delicate balance between pro- and anti-inflammatory mechanisms,

essential for gut immune homeostasis, is affected by the composition of the commensal

microbial community.28 It has been discussed that some beneficial effects of probiotics

like Bifidobacterium ssp. result from the stabilization of the intestinal barrier and from

changes in the composition of the intestinal microbiota.29 Results obtained in a mouse

model showed that Bifidobacterium ssp. lowered gut endotoxin concentration and

enhanced mucosal immunity.30 Even more, one study on placebo-controlled cross-over

human intervention trial, used prebiotic arabinoxylan oligosaccharides, showed faecal

bifidobacteria levels were significantly increased and increased faecal SCFA

concentrations and lowered faecal pH, indicating increased colonic fermentation of the

prebiotic, however also caused a mild increase in flatulence occurrence frequency.31 The

usual target genera for prebiotics are the genera Lactobacillus and Bifidobacterium. As

knowledge of gut microbiota diversity has expanded, there may be other target genera

for prebiotic approaches such as Roseburia, Eubacterium and Faecalibacterium. In

some cases, these may produce desirable metabolites that Lactobacillus and

Bifidobacterium cannot.32

Childhood obesity not only causes long-term health problems that become obvious in

adulthood like cardiovascular diseases and cancer, but also short-term secondary

complications, including dyslipidemia, insulin resistance and non-alcoholic fatty liver

disease (NAFLD), as well as an advance to non-alcoholic steatohepatitis (NASH).33

Interestingly a genetic intervention in a rat model has demonstrated that the gut

microbiota is involved in obesity and metabolic disorders.15 Even more, In a mouse

model of non-alcoholic fatty liver disease (NAFLD), treatment with VSL#3, a mixture

of eight probiotic strains improved liver histology, reduced hepatic total fatty acid


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econtent, and decreased serum alanine aminotransferase (ALT) levels.34 Most recently, a

study performed a double-blind RCT of VSL#3 vs. placebo in obese children with

biopsy-proven NAFLD and found that a 4-month supplement of VSL#3 significantly

improved fatty liver and significantly reduced body mass index.35

Emerging data to indicate the metabolites of free fatty acids (FFAs) cause hepatic

lipotoxicity, which contributes to the pathogenesis of NASH.36 Specific FFA, such as

palmitate, has been shown to trigger endoplasmic reticulum stress that may contribute to

promoting β-cells apoptosis.37 In this research, the influences of the dietary on the

concentration of individual FFA contents in rat serum are shown in Table 4. It was

observed that the concentrations of palmitic acid, stearic acid, oleic acid and linolenic

acid were reduced when the rats fed with FOS and synbiotic diets in compassion with

the commercial diet. However, the results showed no statistical differences (p>0.05)

between rats fed with FOS and synbiotic supplemented diet and those on the control

diet. Since the mechanism of this action is no clear, some authors suggest that prebiotics

modulate hepatic steatosis by changing gene expression in the liver, a phenomenon that

could implicate micro-RNA and gut-derived hormones.38

The increase of the number of colon cell, in the group fed with synbiotic diet, suggest

that probiotics communicate with epithelial cells and different sets of cells implicated in

both innate and acquired immune response via pattern-recognition receptors. They can

enhance gut barrier function and reduce intestinal permeability for intestinal

microorganisms and other antigens.39


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eConclusions

Here is presented information about the structural heterogeneity of fructans from A.

salmiana, which may be useful in the knowledge of the possibility to use it like specific

prebiotic, as well as their contribution to maintain persistence of representative

probiotic strains in vivo. Our results showed the physiological effect of fructans

obtained from A. salmiana when added in the diet of Wistar rats, which evidences

favorable changes with the FOS diet, even these changes are enhanced with the

synbiotic diet. The variation of the gut microbiota composition that might be caused by

microbiota-targeted therapies might also have different results in different individuals.

More detailed studies, including clinical trials are warranted to confirm the role of FOS

to develop more solid therapeutic solutions in the human.

Funding

The funding for these studies were provided by: Fundación Produce de San Luis A.C.

and scholarships from Sistema Nacional de Investigadores (SNI 48199) and Consejo

Nacional de Ciencia y Tecnología (CONACyT): 233241.

Acknowledgements

We thank Evelyn Regalado Renteria for technical assistance with the animal model.

Conflict of Interest. All authors report no conflict of interest.

Ethical approval: All procedures performed in studies involving animals were in

accordance with the ethics recommendations and approved by the Local University

Ethics Committee.


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Effect of prebiotics of Agave salmiana fed to healthy Wistar rats

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