Higher stability and better photoluminescence quantum yield of cesium lead iodide perovskites nanoparticles in the presence of CTAB ligand.

Inorganic halide perovskites, such as CsPbI3, have unique optoelectronic properties which made them promising candidates for several applications. Unfortunately, these perovskites undergo rapid chemical decomposition and transformation into yellow δ-phase. Thus, the synthesis of stable cesium lead iodide perovskites remains an actual challenging field and it is imperative to develop a stabilized black phase for photovoltaic applications. For this purpose, a surfactant ligand was used to control the synthesis of inorganic perovskite CsPbI3 nanoparticles. Herein we demonstrate a new avenue for lead halide perovskites with the addition of either hexadecyltrimethylammonium bromide (CTAB) or silica nanoparticles to maintain in the first place; the stability of the α-CsPbI3 phase, and later on to boost their photoluminescence quantum yield (PLQY). The prepared perovskites were characterized using UV-visible absorption spectroscopy, fluorescence spectroscopy, scanning electron microscopy, thermogravimetric analysis and X-Ray diffraction technique. Results show higher stability of α-CsPbI3 phase and improvement in PLQY % to reach 99% enhancement in presence of CTAB. Moreover, the photoluminescence intensity of CsPbI3 nanoparticles was higher and was maintained for a longer duration in the presence of CTAB.

Interaction of Curcumin with Poly Lactic-Co-Glycolic Acid and Poly Diallyldimethylammonium Chloride By Fluorescence Spectroscopy.

Poly Lactic-Co-Glycolic Acid (PLGA) and Poly Diallyldimethylammonium Chloride (PDDA) are widely being used for drug delivery and curcumin is being studied as potential drug molecule for its anti-oxidant, anti-inflammatory and anti-cancer activities. The interaction between PLGA, PDDA and curcumin was investigated by fluorescence spectroscopy. The modified Stern-Volmer equation was used to estimate the value of the binding constant Ka and the van't Hoff equation was used to estimate the corresponding thermodynamic parameters (ΔHo, ΔSo, and ΔGo). The obtained results showed that the binding constant between PLGA and Curcumin is due to the formation of hydrogen bonds and van der Waals forces. However, PDDA interacts with curcumin through hydrophobic interactions. Moreover, zeta potential measurements were obtained for these polymers and the surface charge was compared in presence and absence of the negatively charged curcumin molecules. It was found that the results obtained by zeta potential measurements are in agreement with those obtained by fluorescence spectroscopy. It is also found that binding of curcumin with PDDA is further encouraged in the presence of PLGA.

Curcumin Modulates 1,2-dibehenoyl-sn-glycero-3-phosphocholine (DBPC) Liposomes: Chitosan Oligosaccharide Lactate Influences Membrane Fluidity But Does Not Alter Phase Transition Temperature of DBPC Liposomes.

1,2-dibehenoyl-sn-glycero-3-phosphocholine (DBPC) is one of the important phospholipids found in cell membrane but not studied well. Importance of curcumin as a dietary supplement and for its medicinal properites is getting widely recoginsed. The present study for the first time explores the effect of curcumin on properties of DBPC liposomes by monitoring the fluorescence properties of curcumin. The phase transition temperature (Tm) of DBPC is assessed which confirms increase in curcumin concentration causes a slight drop in the Tm value. Chitosan is being applied for various drug delivery uses. The study establishes new insight on effect of chitosan oligosaccharide lactate on DBPC liposomes. It is found that in the absence of chitosan oligosaccharide lactate, curcumin partitions more strongly in the liquids crystalline phase than in the solid gel phase, however, the opposite result is obtained with the presence of chitosan oligosaccharide lactate which penetrates into the DBPC liposomes membranes at higher temperature, blocking thus the passage of curcumin into the lipid bilayer. However, addition of chitosan oligosaccharide lactate had no effect on the Tm. Fluorescence quenching study of curcumin establishes that the location of curcumin to be in the hydrophobic cavity of DBPC membrane.

Curcumin-embedded DBPC liposomes coated with chitosan layer as a fluorescence nanosensor for the selective detection of ribonucleic acid.

One of the limitations of fluorescence probe molecules during biomedical estimation is their lack of ability to selectively determine the targeted species. To overcome this there have been various approaches that involve attaching a functional group or aptamers to the fluorescence probe. However, encapsulating probe molecules in a matrix using nanotechnology can be a viable and easier method. Curcumin (Cur) as a fluorescence marker cannot distinguish DNA and RNA. This research reports a novel selective approach involving the use of nanocapsules composed of liposomal curcumin coated with chitosan for the selective detection of RNA molecules using a fluorescence method. The increase in RNA concentration enhanced the electrostatic interaction between the negatively charge surface of RNA and the positively charged nanocapsule, which was further verified by zeta potential measurement. This method had a low limit of detection (36 ng/ml) and higher linear dynamic ranges compared with other studies found in the literature. Moreover, the method was not affected by DNA and was selective for the detection of RNA molecules for which the site of interaction was confined only to uracil. The selectivity for RNA molecules towards other analogues species was also examined and recovery range found was between 99 and 100.33%.

Curcumin encapsulated colloidal amphiphilic block co-polymeric nanocapsules: colloidal nanocapsules enhance photodynamic and anticancer activities of curcumin.

Curcumin-based novel colloidal nanocapsules were prepared from amphiphilic poly(ethylene oxide)-block-poly(propylene oxide)-block-poly(ethylene oxide) (F108). These colloidal nanocapsules appeared as spherical particles with size ranging between 270 and 310 nm. Curcumin fluorescence spectra exhibited an aggregation-induced 23 nm red-shift of the emission maximum in addition to the enhancement of the fluorescence quantum yield in these nanocapsules. The cytotoxicity of curcumin and colloidal nanocapsules was assessed using human derived immortalized cell lines (A549 and A375 cells) in the presence and absence of light irradiation. The nanocapsules exhibited a >30-fold decrease in IC50, suggesting enhanced anticancer activity associated with curcumin encapsulation. Higher toxicity was also reported in the presence of light irradiation (as shown by the IC50 data), indicating their potential for future application in photodynamic therapy. Finally, A375 cells treated with curcumin and the nanocapsules showed a significant increase in single- and/or double-strand DNA breaks upon exposure to light, indicating promising biological effects.

Fluorescence Sensing of Nucleic Acid by Curcumin Encapsulated Poly(Ethylene Oxide)-Block-Poly(Propylene Oxide)-Block-Poly(Ethylene Oxide) Based Nanocapsules.

In a novel approach, curcumin has been encapsulated inside Poly(Ethylene Oxide)-Block-Poly(Propylene Oxide)-Block-Poly(Ethylene Oxide) (F108) nanocapsules. FTIR spectra have indicated a type of hydrogen bonding and dipole interaction between curcumin and F108. Fluorescence and UV-visible absorption profiles of curcumin in nanocapsules have indicated location of curcumin in more hydrophobic microenvironment. The relative fluorescence yield has increased by 6 times in the nanocapsules, which renders them as more sensitive probes to be used later on in sensing study. Therefore, based on the functionality of curcumin as a fluorescent transducer, encapsulated curcumin is used in biomedical application as DNA and RNA sensing. Detection limits are detected as 50 µM and 60 µM for DNA and RNA respectively. Linear dynamic concentration range obtained in this proposed method is much higher than reported in literature. The interaction between the nanocapsules and targeted DNA/RNA molecules is further approved by zeta potential studies. Furthermore, the real interaction of DNA with the encapsulated curcumin is confirmed by the interaction of the adenine and cytosine nucleotides. This has been verified through zeta potential measurements. Moreover, our prepared nanocapsules has presented a high percentage recovery of DNA and RNA (96-101%). Finally, stability results have illustrated a high photostability of encapsulated curcumin, indicating that proposed nanocapsules can be considered as a stable sensor during measurement time.

Introducing Principal Coordinate Analysis (PCoA) Assisted EEMF Spectroscopic Based Novel Analytical Approach for the Discrimination of Commercial Gasoline Fuels.

In the present work, a novel analytical procedure by integrating principal coordinate analysis (PcoA) with excitation-emission matrix fluorescence (EEMF) spectroscopy was introduced for discriminating the commercial gasoline fuels. The PcoA technique involved analysis of the distance matrices containing the dissimilarity information and it can serve as an efficient tool for capturing the major as well as subtle compositional differences among the analyzed commercial gasoline samples. The utility of the proposed PcoA assisted EEMF analytical procedure was successfully tested by discriminating gasoline fuel samples belonging to five different industrial brands. The obtained results clearly showed that combination of PcoA and EEMF could provide a simple, sensitive and economical analytical procedure to carry out the rapid analyses of the gasoline samples belonging to different brands.

Interaction of carbon nanotubes with curcumin: Effect of temperature and pH on simultaneous static and dynamic fluorescence quenching of curcumin using carbon nanotubes.

Curcumin (Cur) has medicinal properties, undergoes hydrolysis, and has low water solubility that limits its bioavailability and industrial usage. Different host molecules such as carbon nanotubes (CNT) can be useful in improving solubility and stabilizing Cur, therefore understanding the interaction of Cur with host molecules such as CNT is crucial. In this study, UV-visible light absorption and fluorescence spectroscopic techniques have been applied to reveal the interaction of Cur with CNT. Visible light absorption of Cur increases with CNT concentration, whereas fluorescence intensity of Cur is quenched in the presence of CNT. The obtained results confirm that fluorescence reduction is due to both static and dynamic quenching and is a result of the ground state and excited-state complex formation. The pH environment influences the quenching rate due to deprotonation of Cur at higher pH; excess OH- ion concentration in the solution further discourages electrostatic interaction between the deprotonated form of Cur with CNT. It is found that at lower temperatures (<35°C) dynamic quenching is much more dominant and at higher temperatures (45°C) static quenching is more dominant. The interaction is further supported using X-ray diffraction patterns and Fourier transform infrared spectra in the solid state, and suggests encapsulation of curcumin within the CNT. It is further evident that fluorescence quenching of Cur using CNT is further enhanced in the presence of several salts, as increase in ionic strength of the solution pushes the hydrophobic Cur molecule towards the CNT core by increasing the proximity between them.

Glutathione-capped CuO nanoparticles for the determination of cystine using resonance Rayleigh scattering spectroscopy.

Ascorbic acid was used to reduce cystine to cysteine that induces the aggregation of glutathione-capped copper oxide nanoparticles. The aggregation of CuO NPs was optimized through resonance Rayleigh scattering and dynamic light scattering measurements. The high specificity toward cysteine from other amino acids and biomolecules was due to its mercapto group that binds to the surface of CuO NPs and the electrostatic interaction between the cysteine zwitterions on the surface of CuO NPs. Accordingly, glutathione-capped copper oxide nanoparticles was used as a sensing probe for cystine based on resonance Rayleigh scattering (RRS) technique. Increase in the RRS signal of CuO NPs was observed with increasing cystine concentration. A linear calibration plot was obtained in the range 2-20 µM with a limit of detection of 4.55 ± 0.5 nM, which is lower than literature value. The applicability of the proposed sensing strategy toward cystine was established, and the recovery percentage was between 99.8 ± 0.4 and 101.0 ± 2.1 for n = 3. Graphical Abstract .

Spectroscopic Evaluation of Novel Adenine/Thymine-Conjugated Naphthalenediimides: Preference of Adenine-Adenine over Thymine-Thymine Intermolecular Hydrogen Bonding in Adenine- and Thymine-Functionalized Naphthalenediimides.

The synthesis and spectroscopic characterization of novel nucleobase (adenine/thymine)-conjugated naphthalenediimides (NDIs), namely, NDI-AA, NDI-TT, and NDI-AT have been successfully achieved. NDI-AA, NDI-TT and NDI-AT have similar absorption in the 300-400 nm region. The effect of solvent on the absorption spectrum indicates aggregation, either through intermolecular π-σ interaction among the main chromophore or through intermolecular hydrogen bonding between adenine and adenine group. Addition of water does not assist hydrogen bond formation between thymine-thymine, rather increasing the polarity of the solvent encourages π-σ interaction among NDI-TTs. No spectral change for NDI-TT with increasing temperature confirms hydrogen bonding is not playing a crucial role in NDI-TT. A fluorescence study on NDI-AA also establishes excimer formation along with ground state aggregation. As the water content in the solvent mixture increases, aggregation of NDI-AA is discouraged due to adenine-adenine hydrogen bonding in accordance with earlier results. At the same time, the NDI-TT emission spectrum does not shift to the blue region and the intensity of the peak around 535 nm increases at the expense of fluorescence in 411 nm. Thus, increasing water content in the solvent mixture facilitates aggregation through π-σ interaction in NDI-TT as thymine-thymine hydrogen bonding is less pronounced.

Gold and silver nanoparticles in resonance Rayleigh scattering techniques for chemical sensing and biosensing: a review.

This review (with 116 refs.) summarizes the state of the art in resonance Rayleigh scattering (RRS)-based analytical methods. Following an introduction into the fundamentals of RRS and on the preparation of metal nanoparticles, a first large section covers RRS detection methods based on the use of gold nanoparticles, with subsections on proteins (albumin, bovine serum albumin and ovalbumin, glycoproteins, folate receptors, iron binding-proteins, G-proteins-coupled receptors, transmembrane proteins, epidermal growth factor receptors), on pesticides, saccharides, vitamins, heavy metal ions (such as mercury, silver, chromium), and on cationic dyes. This is followed by a section on RRS methods based on the use of silver nanoparticles, with subsections on the detection of nucleic acids and insecticides. Several Tables are presented where an RRS method is compared to the performance of other methods. A concluding section summarizes the current status, addresses current challenges, and gives an outlook on potential future trends. Graphical Abstract Change in the resonance Rayleigh scattering (RRS) intensity when mixing the nanoparticles with the specific analyte.

Tuning the surface of Au nanoparticles using poly(ethylene glycol)-block-poly(propylene glycol)-block-poly(ethylene glycol): enzyme free and label free sugar sensing in serum samples using resonance Rayleigh scattering spectroscopy.

Poly(ethylene glycol)-block-poly(propylene glycol)-block-poly(ethylene glycol) (F-108) functionalized gold nanoparticles (Au NPs) have been successfully synthesized. During synthesis it is found that an increase in the F-108 concentration contributes to agglomeration in the media, increasing the size of the Au particles, and boosting the curcumin concentration leads to a higher density of functional groups, resulting in smaller Au NPs. FT-IR analysis reveals that the hydroxyl and phenolic groups of curcumin and F-108 are involved during the functionalization of Au surfaces. Enhancement in the fluorescence/RRS intensity is due to the combination of the influence of the shape/size of the Au NPs as well as the extent of curcumin conjugation at the interface of the Au NP surface and F-108. The presence of sugar molecules remarkably boosts the RRS intensity without significantly affecting the fluorescence and surface plasmon absorbance of the Au NPs; in contrast, the RRS intensity of standard CTAB functionalized Au NPs is unaffected by glucose molecules indicating that the functionalization of F-108 at Au surfaces is crucial. Interestingly, no interference from other potential interferents and antioxidant substances like ascorbic acid, creatinine and acetaminophen is observed. This method is simple and fast, and offers a wider linear dynamic range, 0-10 mM, that is applicable under physiological conditions and in serum samples. It is stable and provides an excellent recovery for serum samples, thus, potentially it can be useful in this field due to its low energy consumption, enzyme free assay, fast response time, better selectivity and sensitivity.

Nanosensing of ATP by fluorescence recovery after surface energy transfer between rhodamine B and curcubit[7]uril-capped gold nanoparticles.

The authors describe a method for functionalization of gold nanoparticles (AuNPs) with the supramolecular host molecule, curcubit[7]uril (CB[7]) which can bind rhodamine B (RhB). The fluorescence of RhB is quenched by the AuNPs via surface energy transfer. On addition of ATP, a dimeric RhB-ATP complex is formed and RhB is pushed out of CB[7]. Hence, fluorescence increases by a factor of 8. This fluorescence recovery effect has been utilized to develop a new detection scheme for ATP. The assay, measured at fluorescence excitation and emission wavelengths of 500 nm and 574 nm respectively, works in the 0.5-10 µM concentration range and has a 100 nM detection limit. The method is not interfered by UTP, GTP, CTP, TTP, ascorbic acid and glutathione. Graphical abstract Schematic of a method for determination of ATP in the 500 nM to 10 µM concentration range by using fluorescence recovery after surface energy transfer (SET) between rhodamine B (RhB) and gold nanoparticles capped with curcubit[7]uril (CB[7]).

Gold nanoparticles functionalized with Pluronic are viable optical probes for the determination of uric acid.

The authors describe the preparation of gold nanoparticles (AuNPs) coated with poly(ethylene glycol)-block-poly(propylene glycol)-block-poly(ethylene glycol) (Pluronic F-108) by reducing Au3+ to Au0 using curcumin, a natural and non-toxic food spice, in water of pH ~7 in the presence of F-108 and Ag+ ion. The coated AuNPs display strong resonance Rayleigh scattering (RRS) and fluorescence that results from the functionalization of the gold surface with curcumin and Pluronic F-108. The molar mass of Pluronic F-108 affects the particle size of the AuNPs formed, and small AuNPs are formed when using low molar weight F-108 that was purified by centrifugation or dialysis. The coated AuNPs were employed in an optical method for the determination of uric acid. The combination of uric acid with the AuNPs boosts both the RRS signal and the fluorescence of the AuNPs. However, higher concentrations of uric acid shift the fluorescence peak to shorter wavelengths. The method is simple, and fluorescence, best measured at excitation/emission wavelengths of 425/534 nm, increases linearly in the 50 µM to 50 mM uric acid concentration range, with a 0.14 µM detection limit which is lower than reported for other methods in the literature. Graphical abstract Pluronic F-108 capped gold nanoparticles prepared by reducing Au3+ to Au0 using curcumin can estimate uric acid in 50 µM to 50 mM concentration range.

The role of OH- in the formation of highly selective gold nanowires at extreme pH: multi-fold enhancement in the rate of the catalytic reduction reaction by gold nanowires.

There is a quest to understand the mechanism governing the morphology and geometry control of the particle growth of nanomaterials for their optical and catalytic applications. In the available literature, the role of OH- in dictating the size and shape of Au nanowires is unknown. As one of the first examples, herein, we explore how excess OH- ions in CTAB micelles play a significant role during the highly selective formation of gold nanowires having controlled diameters of ∼20-25 nm and length >1 µm, by reducing Au3+ to Au0 in a one pot, simple synthesis procedure in the presence of Ag+ ions. At pH 4-11, the same procedure does not harvest Au NWs, but Au NPs of diameter 50-70 nm, indicating that excess OH- is needed for nanowire formation. XRD, TGA, DSC, EDX, FT-IR and fluorescence spectroscopic analysis confirm that both CTAB and curcumin act as capping and stabilizing agents for Au NWs as well as Au NPs - there is no remarkable difference in the curcumin/CTAB content between Au NWs and NPs prepared in different pH environments. However, changing the CTAB micellar media to DPPC liposome media inhibits the formation of nanowires at pH ∼13; the growth of the Au NPs diminishes in DPPC liposomes, offering smaller NPs of diameter ∼25 to 30 nm, suggesting that the role of CTAB is necessary in nanowire formation. The rate of NW formation has been found to be 0.13 h-1 and the growth mechanism advocates elongation in the [110] facet of Au [110] as opposed to the [100] or [111] facets. Curcumin capped Au nanowires serve as excellent nano-catalysts for the reduction of nitro-compounds and the rate of reduction of 4-nitrophenol, a model compound, by curcumin capped Au NWs is found to be ∼10 fold higher, compared to Au NPs, which signifies that catalytic activities can be dictated by the size and shape of Au NPs.

Cyclic azacyanines: experimental and computational studies on spectroscopic properties and unique reactivity.

The absorption and fluorescence properties of cyclic azacyanine (CAC) derivatives were examined in several solvents. The presence of electron donating or withdrawing groups on the CAC impacts spectroscopic properties. The general solvent relaxation displayed by azacyanine derivatives is in accordance with Lippert-Mataga's prediction but exception is noted in the case of protic solvent due to specific hydrogen bonding interactions. Fluorescence lifetime decay studies indicate a relaxation time in the nanosecond timescale with mono exponential decay. Donating substituents markedly increase the excited state lifetime, whereas withdrawing groups marginally decrease the excited state lifetime. Quantum chemical computations were used to explore the origins of the reactivity and spectroscopic properties of CACs; results are consistent with a model in which regioselectivity results from differences in mechanistic steps occurring after initial attack by hydroxide on the CAC.

Combining time-resolved fluorescence with synchronous fluorescence spectroscopy to study bovine serum albumin-curcumin complex during unfolding and refolding processes.

We investigated the complex interaction between bovine serum albumin (BSA) and curcumin by combining time-resolved fluorescence and synchronous fluorescence spectroscopy. The interaction was significant and sensitive to fluorescence lifetime and synchronous fluorescence characteristics. Binding of curcumin significantly shortened the fluorescence lifetime of BSA with a bi-molecular quenching rate constant of k(q) = 3.17 × 10(12) M(-1) s(-1). Denaturation by urea unfolded the protein molecule by quenching the fluorescence lifetime of BSA. The tyrosine synchronous fluorescence spectra were blue shifted whereas the position of tryptophan synchronous fluorescence spectra was red shifted during the unfolding process. However, denaturation of urea had little effect on the synchronous fluorescence peak of tyrosine in curcumin-BSA complex except in the low concentration range; however, it shifted the peak to the red, indicating that curcumin shifted tryptophan moiety to a more polar environment in the unfolded state. Decreases in the time-resolved fluorescence lifetime and curcumin-BSA complex during unfolding were recovered during refolding of BSA by a dilution process, suggesting partial reversibility of the unfolding process for both BSA and curcumin-BSA complex.

Influence of substituent and solvent on the radiative process of singlet excited states of novel cyclic azacyanine derivatives.

The photophysical properties of novel cyclic azacyanine derivatives have been investigated in acetonitrile, N-butyronitrile, methanol, ethanol, DMF and water. Introduction of electron donating or accepting groups on the cyclic azacyanine has a direct impact on the spectroscopic and photophysical properties. Irrespective of the nature of the substitution, azacyanine shows a general solvent relaxation in accordance with Lippert-Mataga's prediction; however, in protic solvent, specific interactions are encountered. Fluorescence lifetime decay suggests a relaxation in the nanosecond time scale with monoexponential decay in polar solvents and biexponential decay in non polar solvents. The fluorescence lifetime of azacyanines are found to be longer than popular cy3 dyes. An electron donating substituent increases the fluorescence lifetime and influences the radiative process, whereas an electron withdrawing group marginally increases the excited state lifetime but remarkably enhances the radiative process. The fluorescence quantum yield of substituted cyclic azacyanine in water is noted to be at least five fold higher than the popular cy3 dye.

Fluorescence modulation of 1,7-bis(4-hydroxy-3-methoxyphenyl)-1,6-heptadiene-3,5-dione by silver nanoparticles and its possible analytical application.

The effects of silver nanoparticles on the photophysical properties of 1,7-bis(4-hydroxy-3-methoxyphenyl)-1,6-heptadiene-3,5-dione, popularly known as curcumin, have been investigated using optical absorption and fluorescence techniques. Although absorption spectroscopy suggests a ground-state complex formation, fluorescence quenching data confirms a simultaneous static and dynamic quenching, inferring ground as well as excited-state complex formation. The recovery of fluorescence quenching of the curcumin-silver nanoparticle complex in the presence of ascorbic acid or uric acid emphasizes a strong interaction between the silver nanoparticles and ascorbic acid/uric acid, suggesting that fluorescence recovery after the quenching of curcumin-silver nanoparticle complexes has potential for ascorbic acid or uric acid assay development.

Curcumin-PLGA based nanocapsule for the fluorescence spectroscopic detection of dopamine.

The main purpose of this paper is to design curcumin loaded PLGA nanocapsules for the selective detection of dopamine using fluorescence spectroscopy. In the present work curcumin loaded PLGA nanocapsules were synthesized using a solid-in-oil-in water (s/o/w) emulsion technique. The prepared nanocapsules were coated with a poly(diallyldimethylammonium)chloride (PDDA) polymer to increase the entrapment of curcumin into the core of PLGA polymer. PLGA-Cur-PDDA nanocapsules were characterized using different microscopic and spectroscopic techniques. Unlike free curcumin, the formed CUR-PLGA-PDDA NCs were established as nanoprobes for the selective detection of dopamine molecules. The selectivity and specificity of nanocapsules toward dopamine was achieved by measuring the fluorescence emission spectra of the NCs in the presence of other interference molecules such as tryptophan, melamine, adenine, etc. It was noticed that increasing the concentration of the different molecules had no significant change in the fluorescence signal of the nanocapsules. These results confirm the strong quenching between dopamine and curcumin in the nanocapsules. Hence, this fluorescence emission technique was found to be selective, easy and fast with low cost for the determination of dopamine in a concentration range up to 5 mM with a detection limit equal to 22 nM.