New planar assay for streamlined detection and quantification of β-glucuronidase inhibitors applied to botanical extracts

Mahran, Ehab; Keusgen, Michael; Morlock, Gertrud E.

doi:10.1016/j.acax.2020.100039

Cited by 10 publications

(8 citation statements)

References 51 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The β-glucuronidase inhibition assay was run as described recently ( Mahran et al, 2020 ) The chromatogram was pre-wetted with potassium phosphate buffer (0.5 ml; 9.34 mg/ml K 2 HPO 4 and 6.31 mg/ml KH 2 PO 4 ; green nozzle, level 6). Then, 750 µL enzyme solution (25 U/ml in potassium phosphate buffer with 1 mg/ml BSA) were sprayed onto the chromatogram (green nozzle, level 6).…”

Section: Methodsmentioning

confidence: 99%

Is Our Natural Food Our Homeostasis? Array of a Thousand Effect-Directed Profiles of 68 Herbs and Spices

et al. 2021

View full text Add to dashboard Cite

The beneficial effects of plant-rich diets and traditional medicines are increasingly recognized in the treatment of civilization diseases due to the abundance and diversity of bioactive substances therein. However, the important active portion of natural food or plant-based medicine is presently not under control. Hence, a paradigm shift from quality control based on marker compounds to effect-directed profiling is postulated. We investigated 68 powdered plant extracts (botanicals) which are added to food products in food industry. Among them are many plants that are used as traditional medicines, herbs and spices. A generic strategy was developed to evaluate the bioactivity profile of each botanical as completely as possible and to straightforwardly assign the most potent bioactive compounds. It is an 8-dimensional hyphenation of normal-phase high-performance thin-layer chromatography with multi-imaging by ultraviolet, visible and fluorescence light detection as well as effect-directed assay and heart-cut of the bioactive zone to orthogonal reversed-phase high-performance liquid chromato-graphy−photodiode array detection−heated electrospray ionization mass spectrometry. In the non-target, effect-directed screening via 16 different on-surface assays, we tentatively assigned more than 60 important bioactive compounds in the studied botanicals. These were antibacterials, estrogens, antiestrogens, androgens, and antiandrogens, as well as acetylcholinesterase, butyrylcholinesterase, α-amylase, α-glucosidase, β-glucosidase, β-glucuronidase, and tyrosinase inhibitors, which were on-surface heart-cut eluted from the bioautogram or enzyme inhibition autogram to the next dimension for further targeted characterization. This biological-physicochemical hyphenation is able to detect and control active mechanisms of traditional medicines or botanicals as well as the essentials of plant-based food. The array of 1,292 profiles (68 samples × 19 detections) showed the versatile bioactivity potential of natural food. It reveals how efficiently and powerful our natural food contributes to our homeostasis.

show abstract

Section: Methodsmentioning

confidence: 99%

Is Our Natural Food Our Homeostasis? Array of a Thousand Effect-Directed Profiles of 68 Herbs and Spices

et al. 2021

View full text Add to dashboard Cite

show abstract

“…In particular, the detection of estrogen-like, androgen-like, genotoxic, and agonistic/antagonistic effects, which have been developed recently [ 46 ], should be of interest. As demonstrated in previous studies, the individual active compounds can further be characterized by HPTLC-HESI-HRMS and elucidated in its structure [ 64 , 70 ]. Quantification of the discovered activities can be performed by external standards or a well-known reference (equivalency calculation), when a standard compound is not available or the bioactive compound is unknown/unidentified [ 63 , 71 ].…”

Section: Resultsmentioning

confidence: 99%

“…The PC was tetracycline (0.5, 1.5, and 3 μL/band, 0.004 mg/mL in ethanol). For the β-glucuronidase assay [ 64 ], 0.5 mL potassium phosphate buffer (0.1 M, pH 7.0, for prewetting) and then 3.0 mL of β-glucuronidase solution (25 U/mL in previous buffer) were sprayed (yellow nozzle), followed by incubation at 37 °C for 15 min. For generation of the colorless (white) inhibition bands on an indigo-blue background, the plate was sprayed (red nozzle) with 1.0 mL 5-bromo-4-chloro-3-indolyl-β- d -glucuronide solution (2 mg/mL in water) and incubated at 37 °C for 1 h. The PC was d -saccharolactone (0.5, 1.5, and 3 μL/band, 0.1 mg/mL in water).…”

Section: Methodsmentioning

confidence: 99%

HI-HPTLC-UV/Vis/FLD-HESI-HRMS and bioprofiling of steviol glycosides, steviol, and isosteviol in Stevia leaves and foods

Morlock

Heil

2020

Anal Bioanal Chem

Self Cite

View full text Add to dashboard Cite

Food products and botanicals on the global market need to be investigated in a more comprehensive way to detect effects, falsifications or adulterations. This is especially true for such ones containing Stevia leaves, Stevia extracts, or steviol glycosides. A multi-imaging profiling was developed exploiting hydrophilic interaction liquid chromatography (HILIC). A minimalistic sample preparation, different mixtures of acetonitrile and water/buffer on the silica gel phase as well as derivatization reagents and optional hyphenation with high-resolution mass spectrometry were exploited. The hydrophilic interaction high-performance thin-layer chromatography (HI-HPTLC) development took 10 min for 48 analyses. It was used to screen Stevia leaf extracts and 20 different food products. For the first time, the biological and biochemical profiling of Stevia leaf products by HI-HPTLC-UV/ Vis/FLD-assay pointed to 19 different bioactive compound bands found in the more natural multicomponent Stevia leaf extracts, whereas most of these activities were not existent for the steviol glycosides. The chemically isolated, purified, and EU-regulated steviol glycosides ease risk assessment and food product development. However, multipotent botanicals may have subtle impact on homeostasis via several metabolic pathways, providing benefits for the consumer's health. Analyzed side by side by means of the effect-directed profiling, their individual activity profiles were visualized as image and individual substances of importance were pointed out. Multi-imaging (comprehensive detection) plus non-targeted bioprofiling (focus on known and unknown bioactivity) allows for a fast detection of questionable product changes that occur along the global food chain and are particularly related to food safety.

show abstract

“…Furthermore, high intestinal microflora β-GUS activity has been recognized as an important factor that can increase the risk of a panel of intestinal diseases, such as colorectal inflammation and colorectal cancer [28]. Therefore, it is urgent and necessary to find more efficacious EcGUS inhibitors possessing good safety profiles for the prevention and treatment of irinotecan-induced intestinal toxicity and other β-GUS-associated intestinal dysfunction [29,30].…”

Section: Discussionmentioning

confidence: 99%

Beta-Glucuronidase Inhibition by Constituents of Mulberry Bark

Bai

Chen

Cao³

et al. 2021

Planta Med

View full text Add to dashboard Cite

Intestinal bacterial β-glucuronidases, the key enzymes responsible for the hydrolysis of various glucuronides into free aglycone, have been recognized as key targets for treating various intestinal diseases. This study aimed to investigate the inhibitory effects and mechanisms of the Mulberry bark constituents on E. coli β-glucuronidase (EcGUS), the most abundant β-glucuronidases produced by intestinal bacteria. The results showed that the flavonoids isolated from Mulberry bark could strongly inhibit E. coli β-glucuronidase, with IC50 values ranging from 1.12 µM to 10.63 µM, which were more potent than D-glucaric acid-1,4-lactone. Furthermore, the mode of inhibition of 5 flavonoids with strong E. coli β-glucuronidase inhibitory activity (IC50 ≤ 5 µM) was carefully investigated by a set of kinetic assays and in silico analyses. The results demonstrated that these flavonoids were noncompetitive inhibitors against E. coli β-glucuronidase-catalyzed 4-nitrophenyl β-D-glucuronide hydrolysis, with Ki values of 0.97 µM, 2.71 µM, 3.74 µM, 3.35 µM, and 4.03 µM for morin (1), sanggenon C (2), kuwanon G (3), sanggenol A (4), and kuwanon C (5), respectively. Additionally, molecular docking simulations showed that all identified flavonoid-type E. coli β-glucuronidase inhibitors could be well-docked into E. coli β-glucuronidase at nonsubstrate binding sites, which were highly consistent with these agentsʼ noncompetitive inhibition mode. Collectively, our findings demonstrated that the flavonoids in Mulberry bark displayed strong E. coli β-glucuronidase inhibition activity, suggesting that Mulberry bark might be a promising dietary supplement for ameliorating β-glucuronidase-mediated intestinal toxicity.

show abstract

New planar assay for streamlined detection and quantification of β-glucuronidase inhibitors applied to botanical extracts

Cited by 10 publications

References 51 publications

Is Our Natural Food Our Homeostasis? Array of a Thousand Effect-Directed Profiles of 68 Herbs and Spices

Is Our Natural Food Our Homeostasis? Array of a Thousand Effect-Directed Profiles of 68 Herbs and Spices

HI-HPTLC-UV/Vis/FLD-HESI-HRMS and bioprofiling of steviol glycosides, steviol, and isosteviol in Stevia leaves and foods

Beta-Glucuronidase Inhibition by Constituents of Mulberry Bark

Contact Info

Product

Resources

About