Sir2/Uth1/TOR
Silent information regulator 2 (Sir2) – parishin significantly increased SIR2 gene expression.
Superoxide dismutase (SOD1 and SOD2) – parishin increased SOD enzyme activity; the replicative lifespan of sod1 and sod2 mutants was not affected by parishin.
UTH1 and SKN7 – parishin did not affect the replicative lifespan of uth1 and skn7 mutants.
TOR signaling pathway (TORC1, RPS26A, RPL9A) – parishin significantly decreased gene expressions of TORC1, RPS26A, and RPL9A. [1]

K6001 yeast's replicative lifespan is prolonged by parishin at 3, 10, and 30 μM [1].

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parishin significantly extended the replicative lifespan of K6001 yeast at concentrations of 3, 10, and 30 µM (mean lifespan: 8.83±0.56, 9.20±0.52, and 8.98±0.58 generations, respectively) compared to untreated control (7.38±0.44 generations); positive control resveratrol at 10 µM gave 9.23±0.59 generations (p<0.05 and p<0.01). [1]
parishin significantly increased SIR2 gene expression in normal condition (p<0.05, p<0.05, and p<0.01 at 3, 10, 30 µM respectively). [1]
Under oxidative stress (H₂O₂), parishin increased the survival rate of BY4741 yeast: at 3, 10, and 30 µM, survival rates were significantly higher than the positive control (p<0.001 for all). [1]
parishin increased SOD enzyme activity in yeast: after 24 h treatment only at 30 µM (p<0.05); after 48 h treatment at 3 µM (p<0.01), 10 µM (p<0.05), and 30 µM (p<0.05). [1]
parishin decreased ROS levels in yeast at 10 µM (p<0.05) and 30 µM (p<0.01). [1]
parishin decreased MDA levels in yeast at 24 h: 10 µM and 30 µM (p<0.001 each); at 48 h: 3 µM (p<0.001), 10 µM (p<0.001), and 30 µM (p<0.05). [1]
parishin did not affect the replicative lifespan of sod1, sod2, uth1, and skn7 mutants of K6001 yeast. [1]
parishin significantly decreased gene expression of TORC1 at 3, 10, and 30 µM (p<0.05, p<0.05, p<0.01 respectively); decreased RPS26A and RPL9A at 10 and 30 µM (p<0.05 each). [1]
In uth1 mutant and uth1,sir2 double mutant of BY4741, parishin decreased RPS26A and RPL9A gene expressions significantly (p<0.01, p<0.01 for RPS26A; p<0.01, p<0.05 for RPL9A respectively). [1]
In uth1 mutant of K6001, parishin significantly decreased TORC1 gene expression (p<0.05). [1]

For replicative lifespan assay, K6001 yeast strain was resuscitated in galactose medium and incubated at 160 rpm for 24‑28 h at 28 °C. About 1 mL of culture was centrifuged at 1,500 rpm for 3 min, washed three times with PBS, and diluted. Approximately 4,000 cells were plated on glucose medium agar plates containing resveratrol (10 µM, positive control) or parishin (0, 3, 10, 30 µM). Plates were incubated at 28 °C for 2 days, and 40 microcolonies were randomly observed under a microscope. Daughter cells produced by each mother cell were counted. The same method was used for sod1, sod2, uth1, and skn7 mutants with K6001 background. [1]
For antioxidative stress assay (spotting method), BY4741 yeast was treated with parishin (0, 3, 10, 30 µM) or resveratrol (10 µM) at 28 °C for 48 h. Then about 0.1 OD of yeast cultures was spotted on agar plates containing 9 mM H₂O₂. Growth was observed and photographed after incubation at 28 °C for 3 days. [1]
For antioxidative stress assay (propidium iodide staining), BY4741 yeast was incubated with parishin (0, 3, 10, 30 µM) or resveratrol (10 µM) for 24 h, then treated with 0 or 180 mM H₂O₂ for 3 h. Approximately 0.1 OD of yeast was washed with cold PBS three times and treated with 15% ethanol for 20 min. The cells were then incubated with 10 µg/mL propidium iodide at 37 °C for 20 min in the dark after washing with PBS. Fluorescence microscope (excitation 535 nm, emission 615 nm) was used to observe cells. Approximately 100 cells were used to calculate survival rate. [1]
For SOD and MDA assays, BY4741 yeast cells were cultured in glucose medium with parishin (0, 3, 10, 30 µM) for 24 or 48 h. About 1×10⁹ cells were washed three times with PBS and resuspended in 1 mL PBS. Cells were ultrasonicated (1 min each) for five times, followed by freeze‑thaw (5 min in liquid nitrogen then 2 min at 37 °C water bath) and repeated sonication five times. Lysates were centrifuged at 12,000 rpm at 4 °C for 15 min, and the supernatant was used for SOD activity and MDA level measurements using assay kits according to manufacturer’s instructions. [1]
For ROS assay, BY4741 yeast cells were incubated with parishin (0, 3, 10, 30 µM) in glucose medium at 28 °C for 23 h. DCFH‑DA was added to a final concentration of 40 µM and incubated for 1 h at 28 °C in the dark. Cells were washed three times with PBS, and DCF fluorescence of 1×10⁷ cells was detected by a fluorescent plate reader (excitation 488 nm, emission 525 nm). [1]
For RT‑PCR analysis, BY4741, uth1 mutant, and uth1,sir2 double mutant (BY4741 background) were treated with control or parishin and cultured overnight at 28 °C with shaking. Wild type and uth1 mutant of K6001 were cultured in galactose medium overnight. Cells were collected, and RNA was extracted via the hot‑phenol method. RNA was purified, and reverse transcription was performed with 5 µg total RNA. RT‑PCR was performed with thermal cycling: for RPS26A and RPL9A – 40 cycles of 95 °C for 15 s and 60 °C for 35 s; for SIR2 – 40 cycles of 94 °C for 15 s, 60 °C for 25 s, and 72 °C for 20 s; for TORC1 – 40 cycles of 95 °C for 15 s, 59 °C for 25 s, and 72 °C for 20 s. Primers used: RPS26A (sense 5′‑TCAGAAACTTTGTAGACGCCG‑3′, antisense 5′‑ACAATTCTGGCGTGAATAGCAC‑3′), RPL9A (sense 5′‑ATGGTGCCAAATTCATTGAAGTC‑3′, antisense 5′‑AGTTACCTGACAAGACAATTTCG‑3′), SIR2 (sense 5′‑CGTTCCCCAAGTCCTGATTA‑3′, antisense 5′‑CCACATTTTTGGGCTACCAT‑3′), TORC1 (sense 5′‑TTGGTACAAGGCATGGCATA‑3′, antisense 5′‑TACCGTCAATCCGCACATTA‑3′), TUB1 (sense 5′‑CCAAGGGCTATTTACGTGGA‑3′, antisense 5′‑GGTGTAATGGCCTCTTGCAT‑3′). Relative gene expression was analyzed using 2^‑ΔΔCT method, with TUB1 as normalization control. [1]

[1]. Parishin from Gastrodia elata Extends the Lifespan of Yeast via Regulation of Sir2/Uth1/TOR Signaling Pathway. Oxid Med Cell Longev. 2016;2016:4074690.

Reports indicate that Parishin A has been found in Artemisia annua and Gastrodia elata, and relevant data is available for reference.

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parishin was isolated from the rhizomes of Gastrodia elata (dry weight 200 g) by methanol extraction, partitioned between EtOAc and H₂O, followed by ODS column chromatography, silica gel column chromatography, and HPLC (Developal ODS‑UG‑5, MeOH/H₂O 28:72, flow rate 8 mL/min, t_R = 36 min). The structure was confirmed by ¹H NMR, MS, and optical rotation. [1]
parishin exhibits antiaging effects in yeast via antioxidative stress, increasing SIR2 gene expression, and inhibiting the Uth1/TOR signaling pathway. Uth1 can upregulate TOR signaling pathway. The study suggests parishin might be a lead compound for drug discovery against age‑related diseases. [1]

C45H56O25

996.9112

996.311

62499-28-9

10557926

Off-white to light yellow solid

1.6±0.1 g/cm3

1137.0±65.0 °C at 760 mmHg

330.0±27.8 °C

0.0±0.3 mmHg at 25°C

1.657

-3.37

13

25

23

70

1540

15

O1[C@]([H])([C@@]([H])([C@]([H])([C@@]([H])([C@@]1([H])C([H])([H])O[H])O[H])O[H])O[H])OC1C([H])=C([H])C(C([H])([H])OC(C(C([H])([H])C(=O)OC([H])([H])C2C([H])=C([H])C(=C([H])C=2[H])O[C@@]2([H])[C@@]([H])([C@]([H])([C@@]([H])([C@@]([H])(C([H])([H])O[H])O2)O[H])O[H])O[H])(C([H])([H])C(=O)OC([H])([H])C2C([H])=C([H])C(=C([H])C=2[H])O[C@@]2([H])[C@@]([H])([C@]([H])([C@@]([H])([C@@]([H])(C([H])([H])O[H])O2)O[H])O[H])O[H])O[H])=O)=C([H])C=1[H]

WYKQPGOKTKQHQG-SHGJSZTHSA-N

InChI=1S/C45H56O25/c46-15-27-32(51)35(54)38(57)41(68-27)65-24-7-1-21(2-8-24)18-62-30(49)13-45(61,44(60)64-20-23-5-11-26(12-6-23)67-43-40(59)37(56)34(53)29(17-48)70-43)14-31(50)63-19-22-3-9-25(10-4-22)66-42-39(58)36(55)33(52)28(16-47)69-42/h1-12,27-29,32-43,46-48,51-59,61H,13-20H2/t27-,28-,29-,32-,33-,34-,35+,36+,37+,38-,39-,40-,41-,42-,43-/m1/s1

tris[[4-[(2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl]oxyphenyl]methyl] 2-hydroxypropane-1,2,3-tricarboxylate

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: This product requires protection from light (avoid light exposure) during transportation and storage.

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ~125 mg/mL (~125.39 mM)
H2O : ~100 mg/mL (~100.31 mM)

Solubility in Formulation 1: ≥ 6.25 mg/mL (6.27 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 62.5 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 6.25 mg/mL (6.27 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 62.5 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 3: ≥ 6.25 mg/mL (6.27 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 62.5 mg/mL clear DMSO stock solution to 900 μL corn oil and mix evenly.

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 (Please use freshly prepared in vivo formulations for optimal results.)