Melesio Gutiérrez-Lomelí

Biotechnological Production of Plant Secondary Metabolites

Description: Modern techniques have been developed to overcome problems associated with the extraction of natural products from plants. These techniques include production of secondary metabolites by biotechnological methods such as plant tissue culture and microbial biotransformation of natural substances. Such methods have led to an increased yield of secondary metabolite amount, the discovery of new biochemical derivatives and agricultural development. For instance, use of these techniques in agricultural area have led to some beneficial traits such as formation of new varieties of known plant species, better crop quality, higher yield, better nutritive properties, more resistant species to insects and pests. Microbial biotransformation is beneficial in plant secondary metabolite production and derivatization (chemical modification of any compound made by a microorganism) and is chiefly applicable to several varieties of terpenes and steroids. This E-book demonstrates recent developments in this field. It will be of particular interest to the professionals in pharmaceutical and biotechnological industries, as well as natural product chemists, medicinal chemists, plant biochemists, and molecular biologists.

Antimutagenicity of Methanolic Extracts from Anemopsis californica in Relation to Their Antioxidant Activity

Anemopsis californica has been used empirically to treat infectious diseases. However, there are no antimutagenic evaluation reports on this plant. The present study evaluated the antioxidant activity in relation to the mutagenic and antimutagenic activity properties of leaf (LME) and stem (SME) methanolic extracts of A. californica collected in the central Mexican state of Querétaro. Antioxidant properties and total phenols of extracts were evaluated using DPPH (1,1-diphenyl-2-picrylhydrazyl) and Folin-Ciocalteu methods, respectively. Mutagenicity was evaluated using the Ames test employing Salmonella enterica serovar Typhimurium strains (TA98, TA100, and TA102), with and without an aroclor 1254 (S9 mixture). Antimutagenesis was performed against mutations induced on the Ames test with MNNG, 2AA, or 4NQO. SME presented the highest antioxidant capacity and total phenolic content. None of the extracts exhibited mutagenicity in the Ames test. The extracts produced a significant reduction in 2AA-induced mutations in S. typhimurium TA98. In both extracts, mutagenesis induced by 4NQO or methyl-N′-nitro-N-nitrosoguanidine (MNNG) was reduced only if the exposure of strains was <10 μg/Petri dish. A. californca antioxidant properties and its capacity to reduce point mutations render it suitable to enhance medical cancer treatments. The significant effect against antimutagenic 2AA suggests that their consumption would provide protection against carcinogenic polycyclic aromatic compounds.

Storage Effect on Phenols and on the Antioxidant Activity of Extracts from Anemopsis californica and Inhibition of Elastase Enzyme

The amount of total phenols and flavonoids and the antioxidant activity of leaf, stem, and rhizome methanolic extracts from a commonly consumed Anemopsis californica under different storage conditions were investigated. Storage conditions were at 50, 25, 4, and −20°C, protected or not from light, during 180 days. The inhibition of the elastase enzyme was also evaluated. The results demonstrated that leaf, stem, and rhizome methanolic extracts of Anemopsis californica maintain approximately up to 97 and 95% stability in phenolic content and antioxidant activity, respectively, when stored during 60 days at −20°C in the dark. Additionally, these extracts, principally from leaf and rhizome, showed an elastase inhibitory effect by 75 and 71.8%, respectively. Therefore, this study provides the basis for further research on the anti-inflammatory activity. On the other hand, Anemopsis californica could comprise a good alternative of use as antioxidant in foods.

Modulation of antioxidant defense system after long term arsenic exposure in Zantedeschia aethiopica and Anemopsis californica

Zantedeschia aethiopica (calla lily) and Anemopsis californica (yerba mansa) are plant species capable of accumulating arsenic (As) and therefore proposed as phytoremediation for removal of As from drinking water. The effects of a continuous 6 month As exposure (34±11 μg/L) from local contaminated groundwater on the antioxidant response of Z. aethiopica and A. californica were evaluated in leaves and stems of the plants bimonthly in a subsurface flow constructed wetland. As increased the activities of the antioxidant enzymes ascorbate peroxidase, glutathione reductase and catalase where higher levels were observed in Z. aethiopica than A. californica. No significant differences were detected on lipid peroxidation levels or antioxidant capacity evaluated by ORAC and DPPH assays or total phenol contents in any part of the plant, although in general the leaves of both plants showed the best antioxidant defense against the metal. In conclusion, Z. aethiopica and A. californica were able to cope to As through induction of a more sensitive enzymatic antioxidant response mechanism.

Antimicrobial activity of disinfectants commonly used in the food industry in Mexico

OBJECTIVES Disinfectants are widely used in food processing environments for microorganism control; their activity can vary according the microorganism and their used in the appropriated concentrations is vital. Hence, the aim of this study was determined the effectiveness and minimum inhibitory concentration (MIC) of 15 disinfectants commonly used in the food industry in Mexico. METHODS The antimicrobial activity and the MIC were determined according to AOAC and CLSI, respectively, with approved strains. RESULTS Most disinfectants reduced 99.999% of microorganisms in suspension after 30s of contact, so reduction rate corresponded at least 5 Log10. Only for Pseudomonas aeruginosa ATCC 15442, did all disinfectants have 99.999% effectiveness. For the MIC, only the third generation quaternary ammonium compounds (QACs) in acid medium did not have values within the range in which is used in the food industry for Staphylococcus aureus ATCC 25923. In addition, for all disinfectants the MIC at 5min was two to four times greater than the concentration with the same effect at 10min; moreover, in most cases there was no difference in the MIC at 10 and 15min (p>0.05). CONCLUSIONS At recommended concentrations, disinfectants had bactericidal activity for at least three of the six microorganisms evaluated. However, the MIC was affected by the exposure time: it was bigger at 5min than at 10min; moreover, in the majority of the cases, it was equal at 10 and 15min; with the results we could have a better understanding of disinfectants use in food processing environments.

Biofilm formation by Staphylococcus aureus and Salmonella spp. under mono and dual-species conditions and their sensitivity to cetrimonium bromide, peracetic acid and sodium hypochlorite

The aim of this study was evaluated the biofilm formation by Staphylococcus aureus 4E and Salmonella spp. under mono and dual-species biofilms, onto stainless steel 316 (SS) and polypropylene B (PP), and their sensitivity to cetrimonium bromide, peracetic acid and sodium hypochlorite. The biofilms were developed by immersion of the surfaces in TSB by 10 d at 37 °C. The results showed that in monospecies biofilms the type of surface not affected the cellular density (p > 0.05). However, in dual-species biofilms on PP the adhesion of Salmonella spp. was favored, 7.61 ± 0.13 Log10 CFU/cm2, compared with monospecies biofilms onto the same surface, 5.91 ± 0.44 Log10 CFU/cm2 (p < 0.05). The mono and dual-species biofilms were subjected to disinfection treatments; and the most effective disinfectant was peracetic acid (3500 ppm), reducing by more than 5 Log10 CFU/cm2, while the least effective was cetrimonium bromide. In addition, S. aureus 4E and Salmonella spp. were more resistant to the disinfectants in mono than in dual-species biofilms (p < 0.05). Therefore, the interspecies interactions between S. aureus 4E and Salmonella spp. had a negative effect on the antimicrobial resistance of each microorganism, compared with the monospecies biofilms.

Efficient protocols for in vitro axillary bud proliferation and somatic embryogenesis of the medicinal plant Anemopsis californica

Anemopsis californica, commonly known as yerba mansa, is an important medicinal plant that has traditionally been used to treat a variety of digestive, respiratory and skin ailments, and has been reported to possess anticancer properties. The increase in demand for this medicinal plant puts wild populations at risk due to over-harvesting; therefore, the development of alternatives such as in vitro propagation may help reduce this pressure on natural plant resources. The aim of this work was to develop protocols for the in vitro propagation of A. californica through axillary bud proliferation and somatic embryogenesis. Shoot bud multiplication was achieved using nodal explants from axenic seedlings, reaching the highest multiplication rate (on average 15 shoots per explant) when the culture medium was supplemented with 4 mg/L kinetin. Propagated shoots produced roots in medium containing 1 mg/L indole-3-butyric acid. Best induction of somatic embryogenesis (on average 19.75 embryos per explant) was obtained from leaf explants cultured on medium supplemented with 4 mg/L naphthalene acetic acid. These protocols may be used either for the commercial mass propagation of the species, studies on the production of secondary metabolites or as a basis for the development of genetic improvement strategies.

Resistance of pathogenic and spoilage microorganisms to disinfectants in the presence of organic matter and their residual effect on stainless steel and polypropylene

OBJECTIVES The effectiveness of disinfectants can vary according to the microorganism, type of residues and surface. The aim of this study was to determine the effectiveness of four disinfectants in the presence of organic matter and their residual effect on stainless steel grade 304 (SS) and polypropylene B (PP-B). METHODS The effectiveness of the disinfectants in the presence of meat extract, egg yolk and whole milk was determined according to AOAC and UNE-EN 1040:2015, and the residual effect was determined according to UNE-EN 13697:2015 using approved strains. RESULTS The effectiveness of the disinfectants was affected to different degrees depending on the organic matter present. SANICIP Q5 [400μg/mL; fifth-generation quaternary ammonium compound (QAC)] was most effective in the presence of 10% meat extract, whilst SANICIP PAA (200μg/mL; peracetic acid) showed better activity in the presence of 10% egg yolk and whole milk. In the evaluation of residual effect on SS and PP-B, the QAC had a better effect, reducing Listeria monocytogenes ATCC 19111 by 6 Log10 CFU/mL at 24h after its application. Conversely, the disinfectants had no residual effect against Pseudomonas aeruginosa ATCC 15442. CONCLUSIONS The antimicrobial activity of the disinfectants tested against pathogenic and spoilage microorganisms was affected according to the type of organic matter present. SANICIP Q5 had a greater residual effect than the other disinfectants evaluated. Moreover, the residual effect of a disinfectant is greater on SS than on PP-B and is dependent on the microorganism tested.

Biofilm Formation by Staphylococcus aureus Isolated from Food Contact Surfaces in the Dairy Industry of Jalisco, Mexico

Staphylococcus aureus is an important food-borne pathogen able to form biofilms. This pathogen is responsible for outbreaks of food-borne illnesses associated with the consumption of milk and dairy products. The aim of this study was to evaluate the biofilm-forming ability of S. aureus isolates, recovered from food contact surfaces in the dairy industry of Jalisco, Mexico. A total of 84 S. aureus strains were evaluated. The isolates were characterized phenotypically by culture on Congo red agar plates. The ability of the strains to form biofilms was investigated in 96-well flat-bottomed microtiter polystyrene plates. Stainless-steel coupons were used as an experimental surface. Biofilm formation was observed, using epifluorescence microscopy and scanning electron microscopy. Detection of the icaADBC genes in S. aureus was performed by the PCR technique. A total of 52.3% (44/84) of the S. aureus strains contained the icaADBC gene that synthesizes polysaccharide intercellular adhesion (PIA) molecules. On Congo red agar, 75% (63/84) of the S. aureus isolates were biofilm producers, 16.6% (14/84) were non-biofilm formers, and 8.3% (7/84) showed a noncharacteristic phenotype. The biofilm production of the S. aureus strains SA-4E, SA-9, SA-13, and SA-19 on stainless-steel coupons was investigated at 25°C for 8 days, and the detected cell population density was approximately 7.15–7.82 log CFU cm−2. In addition to the ability of biofilm production, it is important to highlight that these strains are potential enterotoxin producers as se genes have been previously detected in their genomes. A part of the ability of biofilm production and the determination of the presence of virulence determinants in the genome of S. aureus can contribute to the pathogenicity of strains. Therefore, vigilant food safety practices need to be implemented in the dairy industries regarding FCS to prevent food-borne infections and intoxications due to S. aureus contamination.

In Vitro Bioactivity of Methanolic Extracts from Amphipterygium adstringens (Schltdl.) Schiede ex Standl., Chenopodium ambrosioides L., Cirsium mexicanum DC., Eryngium carlinae F. Delaroche, and Pithecellobium dulce (Roxb.) Benth. Used in Traditional Medicine in Mexico

Seven out of eight methanolic extracts from five plants native to Mexico were inactive against ten bacterial strains of clinical interest. The fruit extract of Chenopodium ambrosioides inhibited the bacteria Enterococcus faecalis (MIC = 4375 μg/ml), Escherichia coli (MIC = 1094 μg/ml), and Salmonella typhimurium (MIC = 137 μg/ml). The fruit extract of C. ambrosioides was with CC50 = 45 μg/ml most cytotoxic against the cell-line Caco-2, followed by the leaf extract from Pithecellobium dulce (CC50 = 126 μg/ml); interestingly, leaves of C. ambrosioides (CC50 = 563 μg/ml) and bark of P. dulce (CC50 = 347 μg/ml) extracts were much less cytotoxic. We describe for the first time the cytotoxic effect from extracts of the aerial parts and the flowers of Cirsium mexicanum (CC50 = 323 μg/ml and CC50 = 250 μg/ml, resp.). Phytochemical analysis demonstrated for both extracts high tannin and saponin and low flavonoid content, while terpenoids were found in the flowers. For the first time we report a cytotoxicological study on an extract of Eryngium carlinae (CC50 = 356 μg/ml) and likewise the bark extract from Amphipterygium adstringens (CC50 = 342 μg/ml). In conclusion the fruit extract of C. ambrosioides is a potential candidate for further biological studies.