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2 protocols using 4 4 6 dimethoxy 1 3 5 triazin 2 yl 4 methyl morpholinium chloride

1

Synthesis of Dendrimer-Drug Conjugates

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EDA core PAMAM dendrimer G4.5 caboxylate sodium salt was purchased from Dendritech (Midland, MI). 2,2’-Bipyridine, 4-dimethylaminopyridine (DMAP), copper(II) sulfate (CuSO4), (+)-sodium L-ascorbate, 4-(4,6-dimethoxy-(1,3,5)triazin-2-yl)-4-methyl-morpholinium chloride (DMTMM), deuterated solvents, dichloromethane (DCM), dimethyl sulfoxide (DMSO), and other organic solvents were purchased from Acros (Morris Plains, NJ). (S)-(+)-Camptothecin (CPT), propargylamine (PPA), copper bromide (CuBr), N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride (EDC), trifluoroacetic acid (TFA) and silica gel 60 (40-63 μm, 230-400 mesh) were purchased from Sigma-Aldrich (St. Louis, MO). 1-Azido-3,6,9,12,15-pentaoxaoctadecan-18-oic acid (APO) and methoxypoly(ethylene glycol) amine (mPEG-NH2, 2000 g/mol) were purchased from Biomatrik (Jiaxing, Zhejiang, China) and JenKem Technology USA (Plano, TX), respectively. SnakeSkin dialysis tubing 3.5 kDa and 7 kDa MWCO, acetonitrile (ACN), HPLC grade water, phosphate-buffered saline (PBS), and magnesium sulfate (MgSO4) were purchased from Thermo Fisher Scientific (Pittsburg, PA).
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2

Comprehensive Metabolomics Sample Preparation

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Methanol, acetonitrile, perchloric
acid (PCA), trichloroacetic
acid (TCA), hydrochloric acid (HCl), sodium hydroxide (NaOH), (2-bromoethyl)
trimethylammonium bromide, dimethylformamide (DMF), 3-(trimethylsilyl)propionic
acid-2,2,3,3-d4 sodium salt (TSP) were
all obtained from Sigma-Aldrich (St. Louis, MO). 4-(4,6-Dimethoxy[1,3,5]triazin-2-yl)-4-methylmorpholinium
chloride (DMTMM) was obtained from Acros Organic (Pittsburgh, PA),
while 15N-phthalimide potassium and deuterium oxide were
obtained from Cambridge Isotope Laboratories (Andover, MA). All chemicals
were used without further purification. Pooled human serum sample
was obtained from Innovative Research, Inc. (Novi, MI). Deionized
(DI) water was purified using an in-house Synergy Ultrapure Water
System from Millipore (Billerica, MA). Centrifugal filters (3-kDa
cutoff; Amicon Microcon, YM-3) were purchased from Sigma-Aldrich.
Standard compounds used for spiking and confirming the peak assignments
were 1-methylhistidine, 2-hydroxybutyrate, 2-hydroxyisocaproate, 2-hydroxyisovalerate,
2-oxocaproate, 3-methylbutyrate, 3-methylhistidine, 3-methyl-2-oxobutanoate,
arginine, benzoate, betaine, carnitine, citrulline, dimethylglycine,
pyridoxine, pyroglutamate, ornithine, sarcosine, serine and succinate
(all from Sigma-Aldrich).
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