Saikosaponin C targets amyloid beta (Aβ) and tau proteins, acting as a dual-target anti-Alzheimer agent. No IC50/Ki/EC50/DC50 values reported.

Saikosaponin C significantly suppressed the secretion of Aβ1-40 and Aβ1-42 into cell culture supernatants in various neuronal models including SH-SY5Y, H4, primary mouse cortical neurons, organotypic hippocampal slices, PC12 cells, and SH-SY5Y cells stably expressing APPwt or APPswe. [1]
Saikosaponin C did not affect BACE1 enzymatic activity or protein expression, nor did it affect APP phosphorylation at Thr668. [1]
Saikosaponin C inhibited Aβ25-35-induced apoptotic cell death in bovine microvascular endothelial cells (BMVECs) as shown by trypan blue exclusion, sub-G1 fraction, annexin V binding, and LDH release assays in a dose-dependent manner. [1]
Saikosaponin C attenuated abnormal tau hyperphosphorylation at Thr231 and Ser396 induced by wortmannin and GF-109203X in SH-SY5Y and SK-N-SH neuroblastoma cells. [1]
Saikosaponin C promoted NGF-induced neurite outgrowth in PC12 cells, increasing the percentage of differentiated cells and average neurite length. [1]
Saikosaponin C increased microtubule assembly in PC12 cells, as evidenced by increased polymerized tubulin levels. [1]
Saikosaponin C increased synaptic marker proteins synaptophysin (p=0.0523) and PSD-95 (p=0.0157) in primary cortical neurons. [1]

Beta-secretase (BACE1) activity assay: Cells were lysed in extraction buffer, supernatants collected, and protein concentration determined. Supernatants were incubated with beta-secretase substrate and reaction buffer at 37°C in darkness for 1 hour. Fluorescence was measured with excitation at 335-355 nm and emission at 495-510 nm. Beta-secretase activity was expressed as relative fluorescence units per μg protein sample. Saikosaponin C treatment (up to 10 μM) showed no effect on BACE1 activity in SH-SY5Y APPwt and APPswe cells. [1]

Aβ secretion quantification: Cells were treated with indicated concentrations of Saikosaponin C for 36 hours. Culture media were collected and levels of Aβ1-40 and Aβ1-42 were determined using colorimetric sandwich ELISA kits. Samples diluted 1:2 to 1:5 in diluent buffer or standards were incubated on plates pre-coated with an Aβ N-terminus specific monoclonal capturing antibody and an Aβ C-terminus specific biotinylated rabbit detecting antibody for 3 hours at 25°C. After washing, immobilized Aβ was incubated with horseradish peroxidase-labeled streptavidin for 30 minutes. Colorimetric substrate was added and optical density measured at 450 nm. Concentrations calculated using standard curves. [1]
Cell death assay (Trypan blue exclusion): BMVECs were challenged with 10 μM Aβ25-35 for 72 hours in presence or absence of Saikosaponin C (12-hour pre-treatment). Viable cells were determined based on ability to exclude trypan blue. [1]
Flow cytometric detection of sub-G1 fraction: Cells were treated with 10 μM Aβ25-35 for 60 hours with/without Saikosaponin C. Cells were fixed with ice-cold 70% ethanol, washed, stained with propidium iodide (50 μg/mL) for 30 minutes at 25°C in dark, and 10,000 cells measured per sample using flow cytometry. [1]
Annexin V apoptosis assay: Cells were treated with 10 μM Aβ25-35 for 48 hours with/without Saikosaponin C. Cells were stained with propidium iodide and annexin V-FITC for 10 minutes at 25°C in dark and analyzed by flow cytometry. Annexin V-positive cells defined as apoptotic. [1]
LDH release assay: Cells were treated with 10 μM Aβ25-35 for 60 hours with/without Saikosaponin C. Supernatants collected and LDH activity determined by measuring conversion of tetrazolium salt into formazan product at 492 nm. LDH release calculated as percentage of LDH in supernatant compared to total LDH in lysed cells plus supernatant. [1]
Immunoblot analysis: Cells were solubilized using lysis buffer containing protease and phosphatase inhibitors. Protein samples (10 μg/lane) separated by SDS-PAGE, transferred to PVDF membranes, and probed with specific antibodies against BACE1, p-APP (Thr668), synaptophysin, PSD-95, actin, tubulin, APP, Tau-5, AT180 (p-Thr231-Tau), PHF-13 (p-Ser396-Tau). Primary antibodies detected with HRP-conjugated secondary antibody and chemiluminescence. Band intensities quantified using ImageJ. [1]
Neurite outgrowth assay: PC12 cells were serum-starved, pre-treated with Saikosaponin C for 12 hours, then co-treated with 50 ng/mL NGF for 50 hours. Cells fixed with 4% paraformaldehyde for 10 minutes. Differentiated cells defined as cells with at least one neurite length equal to cell body diameter. Percentage of differentiated cells and average neurite length per differentiated cell quantified from 10 random images per triplicate experiment. [1]
Tubulin polymerization assay: PC12 cells treated with Saikosaponin C for 24 hours, lysed with hypotonic buffer at 37°C for 5 minutes. Lysate centrifuged at 18,000 g for 10 minutes at 25°C. Supernatant (soluble/unpolymerized tubulin) and pellet (polymerized tubulin resuspended in buffer) were separated by SDS-PAGE and immunoblotted with anti-tubulin antibody. [1]

[1]. A potential therapeutic effect of saikosaponin C as a novel dual-target anti-Alzheimer agent. J Neurochem. 2016 Mar;136(6):1232-1245.

Saikosaponin C is a potential novel dual-target anti-Alzheimer agent targeting both amyloid beta and tau pathologies. It suppresses Aβ secretion and abnormal tau hyperphosphorylation, promotes neurite outgrowth and microtubule assembly, increases synaptic protein expression, and protects brain endothelial cells from Aβ-induced apoptosis. These findings suggest SSc might be a therapeutic tool for Alzheimer's disease and other neurodegenerative diseases. [1]
The molecular mechanism by which Saikosaponin C inhibits Aβ secretion does not involve BACE1 or APP Thr668 phosphorylation, and remains to be identified. [1]

C48H78O17

927.12

942.518

20736-08-7

167927

White to off-white solid powder

1.4±0.1 g/cm3

310-314ºC

1.632

2.35

10

17

8

65

1780

25

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 : ~100 mg/mL (~107.86 mM)

Solubility in Formulation 1: ≥ 2.5 mg/mL (2.70 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 25.0 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: ≥ 2.5 mg/mL (2.70 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 25.0 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: ≥ 2.5 mg/mL (2.70 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 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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