Substrate recycling amplification enabled by hierarchically mesoporous MOFs for the specific screening of pheochromocytoma and paraganglioma

The sensitive and specific diagnosis of pheochromocytoma and paraganglioma (PPGL) remains challenging due to the low abundance of their detectable biomarker of catecholamines (CAs) in serum. Herein, we established an enzyme-assisted substrate recycling amplification model based on a hierarchically mesoporous UiO-66-NH₂ (HMAUiO) to address these difficulties. Such porous platform was constructed by the Hofmeister ion-assisted nanoemulsion method with mesoporous size beyond 20 nm, in which Pt (II) meso-tetra(4-carboxyphenyl)porphyrin (TCPP-Pt) was incorporated within the microporous framework to form an HMAUiO-Pt sensing probe. The expanded mesopores not only afforded sufficient spaces for the accommodation of multiple enzymes but also facilitated the diffusion of enzyme-substrate of CAs. Tyrosinase and glucose dehydrogenase (GDH), along with its coenzyme nicotinamide adenine dinucleotide (NAD), were co-immobilized in the HMAUiO-Pt, where CAs were catalytically converted by tyrosinase into CAs-quinone which could be converted by the NAD-dependent GDH back to CAs, accompanied by the recycling consumption of oxygen. The oxygen-sensitive reporter of TCPP-Pt in the nearby microporous skeleton was swiftly triggered by the depletion of oxygen to yield fluorescence response, thereby realizing the sensitive detection of CAs with the limit of detection for epinephrine, dopamine, and norepinephrine below 1.35, 1.68, and 1.87 nM, respectively. The developed sensing system can quantitatively distinguish between PPGL, primary hypertension patients, and healthy individuals with high accuracy and reliability. These findings not only prefigure the potential of the proposed sensing model for screening PPGL patients, but also enable the possible diagnosis of different diseases by altering the encapsulated enzymes to recognize different biomarkers.