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There have been remarkable advances in imaging drug nanocarriers, but there are few real-time imaging strategies to determine if the cargo has been released from the carrier. This is important because the pharmacokinetics and pharmacodynamics of the carrier can often be dramatically decoupled from that of the cargo. Thus, new tools are clearly needed to image the timing and quantity of drug release from nanocarriers. Here, we describe a simple strategy for photoacoustic monitoring of drug release based on the redox chemistry of methylene blue, which offers predictable redox chemistry: It can transition from the oxidized state with a bright blue color and robust photoacoustic signal to the reduced state that the transparent with no photoacoustic signal. We locked this drug-dye conjugate into a reduced state inside of a nanoparticle with no photoacoustic signal. As the drug is released from the carrier, the dye is oxidized for quantification with photoacoustic imaging. We first prepared paclitaxel-methylene blue conjugate (PTX-MB) with strong absorbance at 640 nm and photoacoustic intensity proportional to its concentration. This cargo was co-encapsulated in a poly(lactic-co-glycolic acid) nanoparticle with a dithiothreitol reducing agent. The IC(50) of PTX-MB-loaded NPs (PTX-MB @ PLGA NPs) was 78 μg mL(−1). We then used the redox reaction of PTX-MB to monitor its release from poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs). In vitro drug-release in phosphate buffer saline with 20% v/v normal mice serum showed a 670-fold increase in photoacoustic signal. The particles showed an initial burst release (25%) during the first 24 hours. After 24 hours, a sustained release was observed through 120 hours leading to cumulative release of 40.6% of PTX-MB. In vivo drug release study in mice for a duration of 12 hours showed a photoacoustic signal enhancement of up to 649% after 10 hours. We then used this system to treat an orthotopic model of colon cancer via luciferase-positive CT26 cells. Our data showed that tumor burden decreased by 44.7% ± 4.8% when treated with the PTX-MB @ PLGA NPs versus the empty PLGA carrier. This work presents a direct strategy to simultaneously monitor drug release biodistribution.

Heparin is an indispensable drug in anticoagulation therapy but with a narrow therapeutic window, which dictates regular testing and dose adjustment. However, current monitoring tools have a long turnaround time or are operator intensive. In this work, we describe a cellulose-based photoacoustic sensor for heparin. The sensors have a turnaround time of 6 min for whole blood samples and 3 min for plasma samples regardless of heparin concentration. These sensors have a limit of detection of 0.28 U/ml heparin in human plasma and 0.29 U/ml in whole blood with a linear response (Pearson's r = 0.99) from 0 to 2 U/ml heparin in plasma and blood samples. The relative standard deviation was

We recently reported a real-time method to measure heparin in human whole blood based on the photoacoustic change of methylene blue (MB). Intriguingly, the MB behaved unlike other "turn on" photoacoustic probes-the absorbance decreased as the photoacoustic signal increased. The underlying mechanism was not clear and motivated this study. We studied the binding mechanism of MB and heparin in water and phosphate buffer saline (PBS) with both experimental and computational methods. We found that the photoacoustic enhancement of the MB-heparin mixture was a result of MB-heparin aggregation due to charge neutralization and resulting sequestration of MB in these aggregates. The sequestration of MB in the MB-heparin aggregates led to decreased absorbance-there was simply less free dye in solution to absorb light. The highest photoacoustic signal and aggregation occurred when the number of negatively charged sulfate groups on heparin was approximately equal to the number of positively charged MB molecule. The MB-heparin aggregates dissociated when there were more sulfated groups from heparin than MB molecules because of the electrostatic repulsion between negatively charged sulfate groups. PBS facilitated MB dimer formation regardless of heparin concentration and reprecipitated free MB in aggregates due to ionic strength and ionic shielding. Further molecular dynamics experiments found that binding of heparin occurred at the sulfates and glucosamines in heparin. Phosphate ions could interact with the heparin via sodium ions to impair the MB-heparin binding. Finally, our model found 3.7-fold more MB dimerization upon addition of heparin in MB solution confirming that heparin facilitates MB aggregation. We conclude that the addition of heparin in MB decreases the absorbance of the sample because of MB-heparin aggregation leading to fewer MB molecules in solution; however, the aggregation also increases the PA intensity because the MB molecules in the MB-heparin aggregate have reduced degrees of freedom and poor heat transfer to solvent.

Synthetic melanin nanoparticles have value in metal chelation, photoprotection, and biocompatibility. Applications of these materials have been reported in optics, biomedicine, and electronics. However, precise size control has remained relatively difficult-especially for materials below 1000 nm. In this paper we describe the synthesis of ultrasmall synthetic nanoparticles with size of 9.4-31.4 nm in weakly acidic and neutral conditions via UV-irradiation. Size control of these particles was possible by varying the pH from 6.4-10.0. We then used UV-vis, FTIR, and nuclear magnetic resonance to investigate the mechanism of UV-induced polymerization. The data show that reactive oxygen species from UV irradiation oxidizes intermediates of the reaction and accelerates the formation of these synthetic melanin structures.

We developed and designed a near-infrared (NIR) absorbing diboronate xanthene dye ((E)-1,3,3-trimethyl-2-(2-(6-((4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzyl)oxy)-2,3-dihydro-1H-xanthen-4-yl)vinyl)-3H-indol-1-ium) for photoacoustic imaging that is sensitive to reactive oxygen and nitrogen species (RONS). We initially used the probe with an OPO-based laser but found that the photoacoustic intensity degrades by 2.6-fold within 3150 laser pulses. Therefore, we adopted the probe to a LED-based excitation source (The average and standard deviation of photoacoustic intensity in presence of 3150 LED pulses is 118.08 and 1.67 (1.4% variation) respectively) and found that hydrogen peroxide (H2O2), superoxide radical (O2˙ −), and peroxynitrite (ONOO−) produce absorption at 700 nm, which was used for photoacoustic excitation. We observed a photoacoustic intensity increase of 2.1-, 1.9-, and 1.75-fold with addition of ONOO−, O2˙ −, and H2O2, (50 ), respectively. The dye is not sensitive to OCl − and ˙OH. At vascular compartment, formation of ONOO− is based on the reaction of nitric oxide (˙NO) with superoxide radical (O2˙ −) and formed ONOO− xidize plasmatic components as well as reaction with intracellular. We then tested the photoacoustic response of various concentrations of ONOO− (25, 50, 125, 185, 250, 375, and 500) in whole human plasma and blood. Concentrations of 50 ONOO− were also easily detectable with this probe. Finally, we examined the capability of new molecular probe for detection of endogenous RONS via SKOV3 (ovarian cancer) cell media. The RONS from these cells activated the probe but media treated with N-acetylcystein (NAC) (RONS scavenger) did not. In pre-incubated cells with NAC, we observed 2.5-fold decrease in photoacoustic intensity versus untreated cells.

We recently reported a real-time method to measure heparin in blood based on photoacoustic (PA) signal from methylene blue (MB). The PA enhancement was surprisingly accompanied by a decrease in absorbance. Here, we describe a mechanistic study of the MB-heparin binding in water and phosphate buffered saline. The addition of 0.79 mg/mL heparin decreased the nuclear magnetic resonance (NMR) magnitude of 0.90 mg/mL MB by 63% with a 0.25 ppm downshift—this indicated formation of MB aggregates due to π-π staking of MB. We also observed nanoscale MB/heparin aggregates under transmission electron microscopy (TEM). Spectroscopic analysis of the isolated aggregates found that the percentage of MB inside the MB-heparin aggregate increased from 3.6% to 82.5% when heparin concentration was increased from 0.16 mg/mL to 0.79 mg/mL. Meanwhile, the photoacoustic intensity increased 25-fold. The signal increase was largely due to the aggregates rather than free MB in the solution. These trends suggested that the MB-heparin aggregation was responsible for the PA enhancement likely due to the decreased degrees of freedom for MB. Molecular dynamics simulations revealed MB dimer formation on heparin and indicated that electrostatic binding occurred between the central thiazine ring of MB and the sulfates and glucosamines in heparin via electrostatic interaction. These findings elucidate the binding process of MB and heparin and provide strategies for immobilizing MB-like molecules on implantable devices for intravascular heparin sensing.

We recently described a technique to monitor heparin anticoagulation therapy in real-time using methylene blue and photoacoustic imaging. The photoacoustic signal of methylene blue was significantly amplified in the presence of heparin, but the exact mechanism underlying this novel photoacoustic behavior remains unclear. Here, we showed that the signal amplification was due to the aggregation of methylene blue. Methylene blue formed different aggregates in water and phosphate buffered saline (PBS). In water, the absorbance maximum of methylene blue with heparin from 0 to 3 U/mL blue shifted from 660 to 570 nm and the corresponding fluorescence intensity decreased 6-fold, which indicated the methylene blue aggregated from monomer to dimer and eventually to high order aggregates. Furthermore, the corresponding 0.04 ppm chemical shift of the proton in the phenothiazine ring of methylene blue from the nuclear magnetic resonance spectrum suggested the electron delocalization and self-aggregation of methylene blue. The coupling of methylene blue molecules results in extra vibrational relaxations within the split exciton states, and this causes enhanced photoacoustic signal. In PBS, we observed the aggregation of methylene blue/heparin complex using transmission electron microscopy (size=150.5 nm), but the absorbance maximum reversed back to 660 nm. This suggested the methylene blue formed monomer bound to heparin—the heparin could not self-aggregate due to electrostatic repulsive forces. The methylene blue bound monomers experienced less degree of freedom than free monomers and therefore caused excess photoacoustic signal.

Conjugated polymer blends are widely used for realization of white light emitting diodes (WPLEDs). The theoretical understanding of the transfer of energy from high band gap host to the low band gap dopant is very important for designing an efficient and stable white LED system. Novel geometric isomers of thienylene vinylene are studied here for resonant energy transfer between the components of the blends. In this study we report a novel methodology for a simple and pictorial depiction of resonant energy transfer between conjugated polymers in blend for a quick screening of prospective constituents prior to synthesis of these polymeric materials. The resonant energy transfer is also supported by the experimental observations which show a spectral overlap. The sequence of energy transfer predicted by theory is in agreement with the order of band gap of the constituent polymers. The resulting fluorescence spectrum of the blends gave a broad emission in the visible region and is of great significance in the development of white light emission.

The personal ultraviolet (UV) dosimeter is a useful measurement tool to prevent UV induced dermal damages; however, conventional digital dosimeters are either bulky or require external power sources. Here, a wearable, colorimetric UV film dosimeter that provides color transition, from purple to transparent, is reported to indicate the UV dose. The film dosimeter is made of a purple photodegradable dye ((2Z,6Z)-2,6-bis(2-(2,6-diphenyl-4H-thiopyran-4-ylidene)ethylidene)cyclohexanone or DTEC) blended in low density polyethylene film. The DTEC film discolored 3.3 times more under the exposure of UV light (302 nm) than visible light (543 nm), and a UV bandpass filter is developed to increase this selectivity to UV light. The DTEC film completely discolors to transparency in 2 h under an AM 1.5 solar simulator, suggesting the potential as an indicator for individuals with types I-VI skin to predict interventions to avoid sunburn. Finally, the DTEC film is integrated with the UV bandpass filter on a wristband to function as a wearable dosimeter for low cost and convenient monitoring of sunlight exposure.