In vitro assessment of antioxidant, neuroprotective, anti-urease and anti-tyrosinase capacities of Tamarix africana leaves extracts

doi:10.19852/j.cnki.jtcm.20230105.003

Journal of Traditional Chinese Medicine ›› 2023, Vol. 43 ›› Issue (2): 252-264.DOI: 10.19852/j.cnki.jtcm.20230105.003

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In vitro assessment of antioxidant, neuroprotective, anti-urease and anti-tyrosinase capacities of Tamarix africana leaves extracts

Esma Anissa Trad Khodja¹^,²(), Abd El Hamid Khabtane³, Rabah Arhab⁴, Djamila Benouchenne⁵^,⁶, Mohamed Sabri Bensaad⁷^,⁸, Chawki Bensouici⁹, Ramazan Erenler¹⁰

1 Biotechnology, Water, Environment and Health Laboratory, Faculty of Natural and Life Sciences, University Abbes Laghrour, Khenchela 40000, Algeria
2 Laboratory of Natural Substances, Biomolecules and Biotechnological Applications, Faculty of Natural and Life Sciences, University Larbi Ben M Hidi, Oum El-Bouaghi 04000, Algeria
3 Biotechnology, Water, Environment and Health Laboratory. Faculty of Natural and Life Sciences, University Abbes Laghrour, Khenchela 40000, Algeria
4 Laboratory of Natural Substances, Biomolecules and Biotechnological Applications, Faculty of Natural and Life Sciences, University Larbi Ben M Hidi, Oum El-Bouaghi 04000, Algeria
5 Genetics, Biochemistry and Plant Biotechnology Laboratory, Faculty of Natural and Life Sciences, University Constantine 1, Constantine 25000, Algeria
6 Pharmaceutical Sciences Research Center, Constantine 25000, Algeria
7 Laboratory of Cellular and Molecular Physio-Toxicology-Pathology and Biomolecules (LPTPCMB), Faculty of Natural and Life Sciences, University Batna 2, Batna 05000, Algeria
8 Laboratory of Biotechnology of Bioactive Molecules and Cellular Physiopathology (LBMBPC), Faculty of Natural and Life Sciences University Batna 2, Batna 05000, Algeria
9 Biotechnology Research Center Ali Mendjli UV 03, Constantine 25000, Algeria
10 Department of Chemistry, Faculty of Science and Literature, University TokatGaziosmanpasa, Tokat 60250, Turkey

Received:2022-03-22 Accepted:2022-06-18 Online:2023-04-15 Published:2023-03-14
Contact: Esma Anissa Trad Khodja, Biotechnology, Water, Environment and Health Laboratory. Faculty of Natural and Life Sciences, University Abbes Laghrour, Khenchela 40000, Algeria; Laboratory of Natural Substances, Biomolecules and Biotechnological Applications, Faculty of Natural and Life Sciences, University Larbi Ben M Hidi, Oum El-Bouaghi 04000, Algeria. tradkhodja.esma@univ-khenchela.dz. Telephone: +213666712451

Abstract

Abstract:

OBJECTIVE: To characterize the chemical profile of methanolic crude extract and its fractions (Ethyl acetate, n-butanol and aqueous) using liquid chromatography-mass spectrometry (LC-MS) analysis, to evaluate their biological and pharmacological properties: antioxidant (1, 1-diphenyl-2-pycrylhydrazyl (DPPH), 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulphonic) (ABTS^·+), galvinoxyle free radical scavenging, reducing power, phenanthroline and β carotene-linoleic acid bleaching assays), enzymes inhibitory ability against several enzymes [acetyl-cholinesterase (AChE), buthyrylcholinesterase (BChE), urease and tyrosinase].

METHODS: Secondary metabolites were extracted from Tamarix africana air-dried powdered leaves by maceration, the crude extract was fractionated using different solvents with different polarities (Ethyl acetate, n-butanol and aqueous). The amount of polyphenols, flavonoids and tannins (hydrolysable and condensed) were determined using colorimetric assays. A variety of in vitro biochemical tests were carried out to assess antioxidant and oxygen radical scavenging properties using DPPH, ABTS, galvinoxyle free radical scavenging, reducing power, phenanthroline and β carotene-linoleic acid bleaching methods. Neuroprotective effect was examined against acetylcholinesterase and buthy-rylcholinesterase enzymes. The anti-urease and anti-tyrosinase activities were performed against urease and tyrosinase enzymes respectively. The extract's components were identified using LC-MS and compared to reference substances.

RESULTS: The results indicated that Tamarix africana extracts presented a powerful antioxidant activity in all assays and exhibited a potent inhibitory effect against AChE and BChE as well as urease and tyrosinase enzymes. LC-MS analysis identified amount of eight phenolic compounds were revealed in this analysis; Apigenin, Diosmin, Quercetin, Quercetine-3-glycoside, Apigenin 7-O glycoside, Rutin, Neohesperidin and Wogonin in methanolic extract and its different fractions of Tamarix africana from leaves.

CONCLUSIONS: Based on these findings, it is reasonable to assume that Tamarix africana could be considered as a potential candidate for pharmaceutical, cosmetics, and food industries to create innovative health-promoting drugs.

Key words: antioxidants, anti-tyrosinase, anti-urease, liquid chromatography-mass spectrometry, neuroprotection, phenolic content, Tamarix africana

Esma Anissa Trad Khodja, Abd El Hamid Khabtane, Rabah Arhab, Djamila Benouchenne, Mohamed Sabri Bensaad, Chawki Bensouici, Ramazan Erenler. In vitro assessment of antioxidant, neuroprotective, anti-urease and anti-tyrosinase capacities of Tamarix africana leaves extracts[J]. Journal of Traditional Chinese Medicine, 2023, 43(2): 252-264.

Figures/Tables 12

Table 1 Phenolic contents of Tamarix africana leaves extracts and fractions (

\bar{x} \pm s

)

Extracts and fractions	Total phenolic (μg GAE/mg)	Total flavonoids (μg QE/mg)	Total condensed tannin (μg CE/mg)	Total hydrolysable tannin (μg TAE/mg)
Methanol	634.5±3.9	188.9±1.6	8.8±0.8	581.3±4.1
Ethyl acetate	861.0±2.9	334.9±2.5	9.7±1.1	811.0±4.9
n-butanol	896.2±3.6	399.1±3.0	5.9±1.5	740.9±3.6
Aqueous	482.9±2.9	117.1±2.1	1.0±0.4	310.1±2.9

Table 1 Phenolic contents of Tamarix africana leaves extracts and fractions ( $\bar{x} \pm s$ )

Extracts and fractions	Total phenolic (μg GAE/mg)	Total flavonoids (μg QE/mg)	Total condensed tannin (μg CE/mg)	Total hydrolysable tannin (μg TAE/mg)
Methanol	634.5±3.9	188.9±1.6	8.8±0.8	581.3±4.1
Ethyl acetate	861.0±2.9	334.9±2.5	9.7±1.1	811.0±4.9
n-butanol	896.2±3.6	399.1±3.0	5.9±1.5	740.9±3.6
Aqueous	482.9±2.9	117.1±2.1	1.0±0.4	310.1±2.9

Table 2 Identification and quantification of phenolic compounds in different extracts and fractions of Tamarix africana leaves by LC-MS analysis

NO	Phenolic substance	Retention Time (min)	MeOH extract (ppb)	Ethyl acetate extract (ppb)	n-BuOH extract (ppb)	Aqueous extract (ppb)
01	Tangeretin	9.088	ND	ND	ND	ND
02	Gardenin-B	9.46	ND	ND	ND	ND
03	Gentisic acid	2.895	ND	ND	ND	ND
04	chlorogenic acid	3.658	ND	ND	ND	ND
05	cafic acid	2.776	ND	ND	ND	ND
06	Rutin	6.512	ND	ND	0.3674	ND
07	p-Coumaric acid	4.747	ND	ND	ND	ND
08	Ferulic acid	3.884	ND	ND	ND	ND
09	hesperidin	6.392	ND	ND	ND	ND
10	Apigenin 7-O glycoside	6.796	ND	ND	0.1038	ND
11	Quercetin	7.561	ND	ND	1.917	0.506
12	naringenin	7.441	ND	ND	ND	ND
13	kamempferol	7.943	ND	ND	ND	ND
14	protocatechuic acid	2.855	ND	ND	ND	ND
15	gallic acid	1.149	ND	ND	ND	ND
16	4-OH benzoic acid	4.403	ND	ND	ND	ND
17	4-OH benzaldehyde	5.429	ND	ND	ND	ND
18	Taxifolin	5.993	ND	ND	ND	ND
19	diosmet	8.084	ND	ND	ND	ND
20	diosmin	6.734	0.0735	0.079	ND	ND
21	naringin	6.292	ND	ND	ND	ND
22	polydatin	5.851	ND	ND	ND	ND
23	Fisetin	7.078	ND	ND	ND	ND
24	galangin	8.808	ND	ND	ND	ND
25	Quercetin-3-glycoside	6.573	ND	0.037	0.9799	ND
26	neohesperidin	6.453	ND	ND	0.0608	ND
27	apigenin	8.023	0.066	0.055	0.4448	0.09
28	Biochanin A	8.628	ND	ND	ND	ND
29	salicylic acid	4.384	ND	ND	ND	ND
30	wogonin	8.627	ND	ND	0.1542	ND
31	Sinapic acid	4.145	ND	ND	ND	ND
32	Resveratrol	6.655	ND	ND	ND	ND
33	Cinnamic acid	5.469	ND	ND	ND	ND
34	vanilla acid	2.223	ND	ND	ND	ND

Figure 1 LC-MS chromatograms of Tamarix africana MeOH extract A: apigenin; B: diosmin. LC-MS: liquid chromatography-mass spectrometry; MeOH: methanol extract.

Figure 2 LC-MS chromatograms of Tamarix africana ethyl acetate fraction A: diosmin; B: quercetin-3- glikozit; C: apigenin. LC-MS: liquid chromatography-mass spectrometry.

Figure 3 LC-MS chromatograms of Tamarix africana n-BuOH fraction A: rutin; B: apigenin-7-O-glikozit; C: quercetin-3-glikozit; D: neohesperidin; E: wogonin; F: quercetin; G: apigenin. LC-MS: liquid chromatography-mass spectrometry; n-BuOH: n-butanol fraction.

Figure 4 LC-MS chromatograms of Tamarix africana Aqueous fraction A: quercetin; B: apigenin. LC-MS: liquid chromatography-mass spectrometry.

Table 3 Antioxidant capacity (inhibition percentage %) of Tamarix africana extracts and fractions (

\bar{x} \pm s

)

Assays	Extracts and Fractions	Dilution (μg/mL)
Assays	Extracts and Fractions	0.781	1.562	3.125	6.25	12.5	25	50
DPPH	MeOH Ethyl acetate n-BuOH Aqueous BHT BHA	0.16±0.53^ab 2.83±0.60^cd 16.17±1.09^ab 1.95±0.81^cd 20.09±2.33 8.52±1.67	11.22±0.64^ab 15.58±0.18^c 32.36±1.02^cde 9.31±0.76^cde 31.30±1.37 3.97±1.92	15.60±0.54^a 27.74±0.82^d 52.35±1.25^ab 15.09±0.08^cd 37.71±3.01 12.94±4.21	42.27±0.38^ce 54.18±0.60^ed 87.62±1.26^ab 25.76±2.21^ce 47.54±0.13 26.68±0.18	75.78±0.38^a 81.71±1.38^ce 88.05±0.83^cd 53.42±2.72^ce 62.16±2.11 47.12±2.95	87.55±0.08^a 86.97±0.60^b 88.69±0.56^b 82.83±0.68^a 77.60±0.83 68.69±1.17	87.62±0.15^a 87.06±0.52^a 89.09±1.01^ab 87.42±0.51^ce 88.33±0.38 83.69±0.54
ABTS	MeOH Ethyl acetate n-BuOH Aqueous BHT BHA	- 9.59±2.75^ab 6.65±1.49^ce 8.49±0.17^cde 35.61±0.75 31.47±1.60	1.71±0.11^ce 18.62±2.40^ab 8.13±0.96^cd 13.68±0.43^b 58.56±3.22 34.13±1.34	55.19±2.71^cd 37.41±0.26^ab 26.89±1.81^ce 26.68±1.57^cd 75.57±8.07 40.28±2.88	88.59±0.54^a 70.36±0.17^b 53.58±2.79^ns 39.78±0.80^a 92.29±1.60 49,71±0.12	91.35±0.86^b 92.86±0.10^ab 90.40±0.63^a 87.80±2.20^d 93.15±0.19 63.72±2.02	92.04±0.57^a 93.03±0.49^b 91.80±0.97^b 90.20±0.17^c 94.06±0.63 78.52±0.80	92.48±0.00^a 93,20±0.10^b 93.80±1.09^ab 92.37±0.17^b 97.31±0.26 96.81±0.37
β-carotene	MeOH Ethyl acetate n-BuOH Aqueous BHT BHA	- 19.00±0.68^cd 9.77±0.97^ce 0.81±1.95^ns 63.76±0.43 65.55±0.80	10.76±0.50^ab 23.75±0.47^cd 16.46±1.20^cde 9.31±0.76^c 72.52±0.81 74.24±0.24	11.62±1.29^ab 37.98±1.10^c 18.47±0.28^ce 15.09±0.08^c 81.14±0.84 84.23±1.14	20.13±0.46^cd 70.88±1.28^ce 25.33±1.32^ab 25.76±1.21^ab 86.0.9±1.04 90.11±0.68	26.52±1.02^c 88.90±2.09^ns 37.80±0.22^b 53.42±2.72^ns 87.52±4.24 94.59±0.77	42.66±0.95^c 93.76±1.93^ce 61.39±0.01^a 82.83±0.68^b 91.67±0.52 96.09±0.02	71.21±1.95^ab 99.67±0.00^a 87.27±1.70^ab 87.42±0.51^a 94.11±0.4 97.35±1.08
GOR	MeOH Ethyl acetate n-BuOH Aqueous BHT BHA	- 2.65±0.58^ce 3.23±0.23^cd - 27.66±1.62 25.99±2.56	- 4.73±0.10^c 9.82±0.77^ce - 39.11±2.34 39.15±0.88	7.99±1.03^cde 13.89±1.19^ab 14.98±0.17^c 3.50±1.63^cde 58.67±0.94 46.67±0.27	30.82±1.61^c 45.12±0.45^b 40.76±2.06^cd 28.58±0.09^c 69.65±0.04 62.27±1.40	69.21±0.52^cd 73.31±0.95^ce 67.65±0.86^ab 55.67±1.43^cd 70.02±0.50 71.46±0.29	72.71±0.24^a 77.68±1.63^b 74.24±1.01^ce 76.22±1.22^c 72.44±0.23 73.25±0.41	74.70±1.20^ab 82.49±1.05^a 78,01±0.90^b 77.71±0,11^ce 73.61±0.10 73.78±0.17
CUPRAC	MeOH Ethyl acetate n-BuOH Aqueous BHT BHA	0.13±0.01^ab 0.17±0.01^c 0.17±0.01^ab 0.13±0.01^cd 0.23±0.01 0.33±0.02	0.15±0.01^ce 0.23±0.03^ce 0.20±0.01^cd 0.16±0.01^ab 0.32±0.01 0.60±0.04	0.21±0.03^a 0.25±0.02^ce 0.25±0.02^ab 0.20±0.03^cd 0.48±0.03 0.91±0.02	0.30±0.01^ab 0.35±0.05^cde 0.37±0.01^b 0.27±0.03^cde 0.76±0.02 1.35±0.07	0.46±0.05^a 0.56±0.08^cd 0.57±0.02^ce 0.41±0.01^ab 1.08±0.05 2.03±0.09	0.79±0.05^c 0.86±0.04^ce 0.86±0.07^cde 0.66±0.02^ab 1.56±0.08 2.91±0.37	1.54±0.16^cd 1.37±0.11^ab 1.35±0.12^ce 1.01±0.05^b 2.20±0.03 3.60±0.19
Reducing power	MeOH Ethyl acetate n-BuOH Aqueous BHT BHA Ascorbic acid	0.17±0.00^a 0.14±0.01^a 0.18±0.01^ab 0.16±0.03^ce 0.07±0.00 0.09±0.00 0.09±0.00	0.21±0.02^c 0.19±0.02^ce 0.25±0.05^ab 0.17±0.01^ce 0.08±0.00 0.11±0.01 0.11±0.00	0.28±0.01^ce 0.35±0.06 0.40±0.05^ab 0.24±0.02^ce 0.10±0.01 0.18±0.02 0.16±0.01	0.46±0.04^ce 0.50±0.03^a 0.86±0.03^ab 0.29±0.01^ab 0.13±0.02 0.36±0.04 0.33±0.04	0.69±0.07^cd 1.08±0.06 1.00±0.04^ce 0.49±0.08^ab 0.22±0.04 0.78±0.07 0.76±0.16	1.06±0.08^ab 1.56±0.05^c 1.32±0.09^cd 0.89±0.05^a 0.28±0.05 1.74±0.07 2.02±0.23	1.44±0.23^ab 2.63±0.19^a 1.82±0.52^ce 1.15±0.03^c 0.43±0.02 3.53±0.19 3.87±0.27
Phenanthroline	MeOH Ethyl acetate n-BuOH Aqueous BHT BHA	0.43±0.09^ce 0.44±0.03^c 0.26±0.01^a 0.40±0.01^ab 0.47±0.01 0.49±0.01	0.51±0.04^c 0.46±0.02^ce 0.30±0.01^a 0.45±0.01^a 0.47±0.01 0.59±0.01	0.59±0.10^ab 0.51±0.02^cde 0.37±0.01^ab 0.47±0.00^a 0.53±0.03 0.73±0.02	0.60±0.07^b 0.56±0.05^c 0.39±0.02^ab 0.49±0.00^a 1.23±0.02 0.93±0.01	0.67±0.04^ab 0.62±0.08^ce 0.39±0.01^ab 0.61±0.04^ce 1.84±0.01 1.25±0.04	0.97±0.14^b 0.79±0.08^b 0.41±0.00^a 0.84±0.07^b 3.48±0.03 2.10±0.05	1.28±0.12^ab 1.20±0.11^ce 0.54±0.00^a 1.25±0.22^ab 4.84±0.01 4.89±0.06

Table 3 Antioxidant capacity (inhibition percentage %) of Tamarix africana extracts and fractions ( $\bar{x} \pm s$ )

Assays	Extracts and Fractions	Dilution (μg/mL)
Assays	Extracts and Fractions	0.781	1.562	3.125	6.25	12.5	25	50
DPPH	MeOH Ethyl acetate n-BuOH Aqueous BHT BHA	0.16±0.53^ab 2.83±0.60^cd 16.17±1.09^ab 1.95±0.81^cd 20.09±2.33 8.52±1.67	11.22±0.64^ab 15.58±0.18^c 32.36±1.02^cde 9.31±0.76^cde 31.30±1.37 3.97±1.92	15.60±0.54^a 27.74±0.82^d 52.35±1.25^ab 15.09±0.08^cd 37.71±3.01 12.94±4.21	42.27±0.38^ce 54.18±0.60^ed 87.62±1.26^ab 25.76±2.21^ce 47.54±0.13 26.68±0.18	75.78±0.38^a 81.71±1.38^ce 88.05±0.83^cd 53.42±2.72^ce 62.16±2.11 47.12±2.95	87.55±0.08^a 86.97±0.60^b 88.69±0.56^b 82.83±0.68^a 77.60±0.83 68.69±1.17	87.62±0.15^a 87.06±0.52^a 89.09±1.01^ab 87.42±0.51^ce 88.33±0.38 83.69±0.54
ABTS	MeOH Ethyl acetate n-BuOH Aqueous BHT BHA	- 9.59±2.75^ab 6.65±1.49^ce 8.49±0.17^cde 35.61±0.75 31.47±1.60	1.71±0.11^ce 18.62±2.40^ab 8.13±0.96^cd 13.68±0.43^b 58.56±3.22 34.13±1.34	55.19±2.71^cd 37.41±0.26^ab 26.89±1.81^ce 26.68±1.57^cd 75.57±8.07 40.28±2.88	88.59±0.54^a 70.36±0.17^b 53.58±2.79^ns 39.78±0.80^a 92.29±1.60 49,71±0.12	91.35±0.86^b 92.86±0.10^ab 90.40±0.63^a 87.80±2.20^d 93.15±0.19 63.72±2.02	92.04±0.57^a 93.03±0.49^b 91.80±0.97^b 90.20±0.17^c 94.06±0.63 78.52±0.80	92.48±0.00^a 93,20±0.10^b 93.80±1.09^ab 92.37±0.17^b 97.31±0.26 96.81±0.37
β-carotene	MeOH Ethyl acetate n-BuOH Aqueous BHT BHA	- 19.00±0.68^cd 9.77±0.97^ce 0.81±1.95^ns 63.76±0.43 65.55±0.80	10.76±0.50^ab 23.75±0.47^cd 16.46±1.20^cde 9.31±0.76^c 72.52±0.81 74.24±0.24	11.62±1.29^ab 37.98±1.10^c 18.47±0.28^ce 15.09±0.08^c 81.14±0.84 84.23±1.14	20.13±0.46^cd 70.88±1.28^ce 25.33±1.32^ab 25.76±1.21^ab 86.0.9±1.04 90.11±0.68	26.52±1.02^c 88.90±2.09^ns 37.80±0.22^b 53.42±2.72^ns 87.52±4.24 94.59±0.77	42.66±0.95^c 93.76±1.93^ce 61.39±0.01^a 82.83±0.68^b 91.67±0.52 96.09±0.02	71.21±1.95^ab 99.67±0.00^a 87.27±1.70^ab 87.42±0.51^a 94.11±0.4 97.35±1.08
GOR	MeOH Ethyl acetate n-BuOH Aqueous BHT BHA	- 2.65±0.58^ce 3.23±0.23^cd - 27.66±1.62 25.99±2.56	- 4.73±0.10^c 9.82±0.77^ce - 39.11±2.34 39.15±0.88	7.99±1.03^cde 13.89±1.19^ab 14.98±0.17^c 3.50±1.63^cde 58.67±0.94 46.67±0.27	30.82±1.61^c 45.12±0.45^b 40.76±2.06^cd 28.58±0.09^c 69.65±0.04 62.27±1.40	69.21±0.52^cd 73.31±0.95^ce 67.65±0.86^ab 55.67±1.43^cd 70.02±0.50 71.46±0.29	72.71±0.24^a 77.68±1.63^b 74.24±1.01^ce 76.22±1.22^c 72.44±0.23 73.25±0.41	74.70±1.20^ab 82.49±1.05^a 78,01±0.90^b 77.71±0,11^ce 73.61±0.10 73.78±0.17
CUPRAC	MeOH Ethyl acetate n-BuOH Aqueous BHT BHA	0.13±0.01^ab 0.17±0.01^c 0.17±0.01^ab 0.13±0.01^cd 0.23±0.01 0.33±0.02	0.15±0.01^ce 0.23±0.03^ce 0.20±0.01^cd 0.16±0.01^ab 0.32±0.01 0.60±0.04	0.21±0.03^a 0.25±0.02^ce 0.25±0.02^ab 0.20±0.03^cd 0.48±0.03 0.91±0.02	0.30±0.01^ab 0.35±0.05^cde 0.37±0.01^b 0.27±0.03^cde 0.76±0.02 1.35±0.07	0.46±0.05^a 0.56±0.08^cd 0.57±0.02^ce 0.41±0.01^ab 1.08±0.05 2.03±0.09	0.79±0.05^c 0.86±0.04^ce 0.86±0.07^cde 0.66±0.02^ab 1.56±0.08 2.91±0.37	1.54±0.16^cd 1.37±0.11^ab 1.35±0.12^ce 1.01±0.05^b 2.20±0.03 3.60±0.19
Reducing power	MeOH Ethyl acetate n-BuOH Aqueous BHT BHA Ascorbic acid	0.17±0.00^a 0.14±0.01^a 0.18±0.01^ab 0.16±0.03^ce 0.07±0.00 0.09±0.00 0.09±0.00	0.21±0.02^c 0.19±0.02^ce 0.25±0.05^ab 0.17±0.01^ce 0.08±0.00 0.11±0.01 0.11±0.00	0.28±0.01^ce 0.35±0.06 0.40±0.05^ab 0.24±0.02^ce 0.10±0.01 0.18±0.02 0.16±0.01	0.46±0.04^ce 0.50±0.03^a 0.86±0.03^ab 0.29±0.01^ab 0.13±0.02 0.36±0.04 0.33±0.04	0.69±0.07^cd 1.08±0.06 1.00±0.04^ce 0.49±0.08^ab 0.22±0.04 0.78±0.07 0.76±0.16	1.06±0.08^ab 1.56±0.05^c 1.32±0.09^cd 0.89±0.05^a 0.28±0.05 1.74±0.07 2.02±0.23	1.44±0.23^ab 2.63±0.19^a 1.82±0.52^ce 1.15±0.03^c 0.43±0.02 3.53±0.19 3.87±0.27
Phenanthroline	MeOH Ethyl acetate n-BuOH Aqueous BHT BHA	0.43±0.09^ce 0.44±0.03^c 0.26±0.01^a 0.40±0.01^ab 0.47±0.01 0.49±0.01	0.51±0.04^c 0.46±0.02^ce 0.30±0.01^a 0.45±0.01^a 0.47±0.01 0.59±0.01	0.59±0.10^ab 0.51±0.02^cde 0.37±0.01^ab 0.47±0.00^a 0.53±0.03 0.73±0.02	0.60±0.07^b 0.56±0.05^c 0.39±0.02^ab 0.49±0.00^a 1.23±0.02 0.93±0.01	0.67±0.04^ab 0.62±0.08^ce 0.39±0.01^ab 0.61±0.04^ce 1.84±0.01 1.25±0.04	0.97±0.14^b 0.79±0.08^b 0.41±0.00^a 0.84±0.07^b 3.48±0.03 2.10±0.05	1.28±0.12^ab 1.20±0.11^ce 0.54±0.00^a 1.25±0.22^ab 4.84±0.01 4.89±0.06

Table 4 Antioxidant capacities (IC50 and A0.50 μg/mL) of standards and leaves plant extracts and fractions (

\bar{x} \pm s

)

Extracts/Fractions/standards	Antioxydant capacities
Extracts/Fractions/standards	DPPH assay IC₅₀ (μg/mL)	ABTS assay IC₅₀ (μg/mL)	β-carotene assay IC₅₀ (μg/mL)	GOR assay IC₅₀ (μg/mL)	Reducing power assay A_0.50(μg/mL)	CUPRAC assay A_0.50(μg/mL)	Phenanthroline assay A_0.50(μg/mL)
Ethylacetate	5.68±0.06^a	4.34±0.03^a	4.22±0.02^a	7.84±0.20^a	2.73±0.16^c	11.13±2.65^a	0.75±0.23
n-BuOH	2.91±0.08^a	4.73±0.56^a	19.01±0.12^a	8.92±0.11^a	1.72±0.05^c	10.41±0.43^a	10.44±0.20^a
MeOH	7.80±0.11^a	2.01±0.11^ns	31.90±0.71^a	5.99±0.22^b	3.93±0.22^c	13.94±0.72^a	0.20±0.08^a
Aqueous	11.49±0.83^a	7.08±0.21^a	21.87±1.28^a	7.44±0.75^b	6.44±0.32^c	17.19±0.92^a	1.52±0.13^ns
BHA	6.14±0.41	1.81±0.10	1.05±0.03	5.38±0.06	8.41±0.67	5.35±0.71	0.93±0.07
BHT	12.99±0.41	1.29±0.30	0.91±0.01	3.32±0.18	>50	8.97±3.94	2.24±0.17
Ascorbic acid	NT	NT	NT	NT	9.01±1.46	NT	NT

Table 4 Antioxidant capacities (IC50 and A0.50 μg/mL) of standards and leaves plant extracts and fractions ( $\bar{x} \pm s$ )

Extracts/Fractions/standards	Antioxydant capacities
Extracts/Fractions/standards	DPPH assay IC₅₀ (μg/mL)	ABTS assay IC₅₀ (μg/mL)	β-carotene assay IC₅₀ (μg/mL)	GOR assay IC₅₀ (μg/mL)	Reducing power assay A_0.50(μg/mL)	CUPRAC assay A_0.50(μg/mL)	Phenanthroline assay A_0.50(μg/mL)
Ethylacetate	5.68±0.06^a	4.34±0.03^a	4.22±0.02^a	7.84±0.20^a	2.73±0.16^c	11.13±2.65^a	0.75±0.23
n-BuOH	2.91±0.08^a	4.73±0.56^a	19.01±0.12^a	8.92±0.11^a	1.72±0.05^c	10.41±0.43^a	10.44±0.20^a
MeOH	7.80±0.11^a	2.01±0.11^ns	31.90±0.71^a	5.99±0.22^b	3.93±0.22^c	13.94±0.72^a	0.20±0.08^a
Aqueous	11.49±0.83^a	7.08±0.21^a	21.87±1.28^a	7.44±0.75^b	6.44±0.32^c	17.19±0.92^a	1.52±0.13^ns
BHA	6.14±0.41	1.81±0.10	1.05±0.03	5.38±0.06	8.41±0.67	5.35±0.71	0.93±0.07
BHT	12.99±0.41	1.29±0.30	0.91±0.01	3.32±0.18	>50	8.97±3.94	2.24±0.17
Ascorbic acid	NT	NT	NT	NT	9.01±1.46	NT	NT

Table 5 Neuroprotective activity of Tamarix africana extracts and fractions (

\bar{x} \pm s

)

Assays	Extracts/ Fractions/ Standard	Dilution (μg/mL)
Assays	Extracts/ Fractions/ Standard	3.125 μg	6.25μg	12.5 μg	25 μg	50 μg	100 μg	200 μg
AChE	Galantamine	35.93±2.28	43.77±0.00	68.50±0.31	80.69±0.41	85.78±1.63	91.80±0.20	94.77±0.34
	MeOH	33.21±0.72^a	37.96±0.04^c	43.43±2.12^b	47.12±1.31^b	60.52±0.72^b	69.85±1.48^b	76.25±0.17^b
	AE	NA	9.05±0.50^b	21.71±1.69^b	25.67±0.18^b	26.68±0.28^b	51.40±0.63^b	64.37±0.76^b
	n-BuOH	13.09±0.27^b	23.49±0.27^b	28.09±0.07^c	32.35±0.32^b	35.35±0.72^b	46.45±0.53^b	77.88±0.19^b
	Aqueous	30.78±0.17^c	38.48±0.16^c	81.89±0.68^c	57.92±1.91^b	67.09±0.13^c	74.27±0.08^b	74.57±0.38^c
BChE	Galantamine	3.26±0.62	6.93±0.62	24.03±2.94	45.13± 2.60	63.87± 2.85	73.57± 0.77	78.95± 0.58
	MeOH	40.21±0.02^b	47.06±0.14^b	62.97±1.82^b	71.22±0.83^b	76.84±0.96^c	82.76±0.55^a	82.76±0.55^c
	AE	2.89±0.0^b	5.09±0.02^ns	10.77±1.77^c	15.50±0.00^b	23.61±1.37^b	54.58±0.60^ac	79.22±0.99^a
	n-BuOH	43.09±0.27^b	50.49±0.17^b	58.19±1.07^c	65.35±1.32^b	71.69±0.12^c	81.51±2.02^ac	89.17±0.43^b
	Aqueous	10.78±0.16^b	29.48±0.76^b	36.34±0.23^c	49.95±1.90^a	68.08±0.25^a	78.60±0.08^b	81.94±0.75^a

Table 5 Neuroprotective activity of Tamarix africana extracts and fractions ( $\bar{x} \pm s$ )

Assays	Extracts/ Fractions/ Standard	Dilution (μg/mL)
Assays	Extracts/ Fractions/ Standard	3.125 μg	6.25μg	12.5 μg	25 μg	50 μg	100 μg	200 μg
AChE	Galantamine	35.93±2.28	43.77±0.00	68.50±0.31	80.69±0.41	85.78±1.63	91.80±0.20	94.77±0.34
	MeOH	33.21±0.72^a	37.96±0.04^c	43.43±2.12^b	47.12±1.31^b	60.52±0.72^b	69.85±1.48^b	76.25±0.17^b
	AE	NA	9.05±0.50^b	21.71±1.69^b	25.67±0.18^b	26.68±0.28^b	51.40±0.63^b	64.37±0.76^b
	n-BuOH	13.09±0.27^b	23.49±0.27^b	28.09±0.07^c	32.35±0.32^b	35.35±0.72^b	46.45±0.53^b	77.88±0.19^b
	Aqueous	30.78±0.17^c	38.48±0.16^c	81.89±0.68^c	57.92±1.91^b	67.09±0.13^c	74.27±0.08^b	74.57±0.38^c
BChE	Galantamine	3.26±0.62	6.93±0.62	24.03±2.94	45.13± 2.60	63.87± 2.85	73.57± 0.77	78.95± 0.58
	MeOH	40.21±0.02^b	47.06±0.14^b	62.97±1.82^b	71.22±0.83^b	76.84±0.96^c	82.76±0.55^a	82.76±0.55^c
	AE	2.89±0.0^b	5.09±0.02^ns	10.77±1.77^c	15.50±0.00^b	23.61±1.37^b	54.58±0.60^ac	79.22±0.99^a
	n-BuOH	43.09±0.27^b	50.49±0.17^b	58.19±1.07^c	65.35±1.32^b	71.69±0.12^c	81.51±2.02^ac	89.17±0.43^b
	Aqueous	10.78±0.16^b	29.48±0.76^b	36.34±0.23^c	49.95±1.90^a	68.08±0.25^a	78.60±0.08^b	81.94±0.75^a

Table 6 Neuroprotective activities (IC50 μg/mL) of standard and leaves plant extracts and fractions (

\bar{x} \pm s

)

Extracts/Fractions/ standard	Acetylcholinestease Inhibitory Activity	Butyrylcholinesterase inhibitory activity
Ethylacetate	100.64±1.77^a	85.42±2.06^a
n-BuOH	124.95±0.58^a	7.01±0.28^a
MeOH	9.46±0.12^b	8.54±0.32^a
Aqueous	28.39±0.62^a	27.14±0.84^c
Galantamine	6.27±1.15	34.75±1.99

Table 6 Neuroprotective activities (IC50 μg/mL) of standard and leaves plant extracts and fractions ( $\bar{x} \pm s$ )

Extracts/Fractions/ standard	Acetylcholinestease Inhibitory Activity	Butyrylcholinesterase inhibitory activity
Ethylacetate	100.64±1.77^a	85.42±2.06^a
n-BuOH	124.95±0.58^a	7.01±0.28^a
MeOH	9.46±0.12^b	8.54±0.32^a
Aqueous	28.39±0.62^a	27.14±0.84^c
Galantamine	6.27±1.15	34.75±1.99

Table 7 Urease inhibitory activity of Tamarix africana extracts and fractions (

\bar{x} \pm s

)

Extracts/ Fractions/ Standard	Dilution (μg/mL)
Extracts/ Fractions/ Standard	3.125 μg	6.25 μg	12.5 μg	25 μg	50 μg	100 μg	200 μg
Thiourea	4.49±0.78	19.85±2.74	55.64±4.24	94.17±0.15	98.42±0.19	98.49±0.41	98.90±0.05
MeOH	45.01±0.12^a	48.06±0.44^a	52.89±0.85^a	60.89±0.67^a	67.60±1.19^b	85.11±0.09^a	86.33±0.96^b
AE	24.15±0.56^a	30.56±0.00^a	36.89±1.03^a	49.90±0.10^a	55.01±0.37^a	63.76±0.04^b	66.89±1.15^a
n-BuOH	43.09±0.27^a	49.49±0.10^b	57.10±0.07^ns	60.71±0.08^a	66.02±1.33^a	74.82±1.53^b	82.40±2.71^b
Aqueous	18.90±0.06^b	25.89±0.16^b	31.23±0.66^c	38.30±0.49^a	43.17±0.97^a	50.10±1.24^a	57.89±1.59^a

Table 7 Urease inhibitory activity of Tamarix africana extracts and fractions ( $\bar{x} \pm s$ )

Extracts/ Fractions/ Standard	Dilution (μg/mL)
Extracts/ Fractions/ Standard	3.125 μg	6.25 μg	12.5 μg	25 μg	50 μg	100 μg	200 μg
Thiourea	4.49±0.78	19.85±2.74	55.64±4.24	94.17±0.15	98.42±0.19	98.49±0.41	98.90±0.05
MeOH	45.01±0.12^a	48.06±0.44^a	52.89±0.85^a	60.89±0.67^a	67.60±1.19^b	85.11±0.09^a	86.33±0.96^b
AE	24.15±0.56^a	30.56±0.00^a	36.89±1.03^a	49.90±0.10^a	55.01±0.37^a	63.76±0.04^b	66.89±1.15^a
n-BuOH	43.09±0.27^a	49.49±0.10^b	57.10±0.07^ns	60.71±0.08^a	66.02±1.33^a	74.82±1.53^b	82.40±2.71^b
Aqueous	18.90±0.06^b	25.89±0.16^b	31.23±0.66^c	38.30±0.49^a	43.17±0.97^a	50.10±1.24^a	57.89±1.59^a

Table 8 Tyrosinase inhibitory activity of Tamarix africana extracts and fractions (

\bar{x} \pm s

)

Extracts/ Fractions/ Standard	Dilution (μg/mL)
Extracts/ Fractions/ Standard	3.125 μg	6.25 μg	12.5 μg	25 μg	50 μg	100 μg	200 μg
Kojic acid	6.91±0.75	19.22±0.75	36.29±2.92	49.46±2.24	58.32±0.37	64.36±0.65	66.95±2.24
MeOH	37.96±0.54^a	40.99±0.89^a	46.80±1.18^a	48.22±1.79^a	49.52±0.81^c	51.19±0.67^b	59.85±1.79^c
AE	42.53±0.00^a	47.11±0.57^a	48.92±1.70^a	49.08±0.57^b	50.50±0.65^b	51.86±0.49^b	52.84±0.65^a
n-BuOH	46.52±0.82^a	49.63±0.41^a	50.03±0.94^a	57.47±0.00^a	58.54±0.61^b	61.58±0.25^c	68.69±0.20^b
Aqueous	43.66±0.56^a	44.95±0.43^a	45.52±0.49^a	46.51±0.99^b	48.93±0.25^c	50.92±1.23^a	52.35±0.00^a

Table 8 Tyrosinase inhibitory activity of Tamarix africana extracts and fractions ( $\bar{x} \pm s$ )

Extracts/ Fractions/ Standard	Dilution (μg/mL)
Extracts/ Fractions/ Standard	3.125 μg	6.25 μg	12.5 μg	25 μg	50 μg	100 μg	200 μg
Kojic acid	6.91±0.75	19.22±0.75	36.29±2.92	49.46±2.24	58.32±0.37	64.36±0.65	66.95±2.24
MeOH	37.96±0.54^a	40.99±0.89^a	46.80±1.18^a	48.22±1.79^a	49.52±0.81^c	51.19±0.67^b	59.85±1.79^c
AE	42.53±0.00^a	47.11±0.57^a	48.92±1.70^a	49.08±0.57^b	50.50±0.65^b	51.86±0.49^b	52.84±0.65^a
n-BuOH	46.52±0.82^a	49.63±0.41^a	50.03±0.94^a	57.47±0.00^a	58.54±0.61^b	61.58±0.25^c	68.69±0.20^b
Aqueous	43.66±0.56^a	44.95±0.43^a	45.52±0.49^a	46.51±0.99^b	48.93±0.25^c	50.92±1.23^a	52.35±0.00^a

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In vitro assessment of antioxidant, neuroprotective, anti-urease and anti-tyrosinase capacities of Tamarix africana leaves extracts

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