Exploring the maximum spatial resolution achievable in far‐field optical imaging, we show that applying solid immersion lenses (SIL) in stimulated emission depletion (STED) microscopy addresses single spins with a resolution down to 2.4 ± 0.3 nm and with a localization precision of 0.09 nm.

In optical microscopy, most red-emitting dyes provide only moderate performance due to unspecific binding, poor labeling efficiency, and insufficient brightness. Here we report on four novel red fluororescent dyes, including the first phosphorylated dye, created by combining a rigidized rhodamine backbone with various polar groups. They exhibit large fluorescence quantum yields and improved NHS ester stability. While these fluorophores are highly suitable for fluorescence microscopy in general, they excel in stimulated emission depletion (STED) microscopy, providing < 25 nm spatial resolution in raw images of cells.

No abstract

Bright and photostable fluorescent dyes with large Stokes shifts are widely used as sensors, molecular probes, and light-emitting markers in chemistry, life sciences, and optical microscopy. In this study, new 7-dialkylamino-4-trifluoromethylcoumarins have been designed for use in bioconjugation reactions and optical microscopy. Their synthesis was based on the Stille reaction of 3-chloro-4-trifluoromethylcoumarins and available (hetero)aryl- or (hetero)arylethenyltin derivatives. Alternatively, the acylation of 2-trifluoroacetyl-5-dialkylaminophenols with available (hetero)aryl- or (hetero)arylethenylacetic acids followed by intramolecular condensation afforded coumarins with 3-(hetero)aryl or 3-[2-(hetero)aryl]ethenyl groups. Hydrophilic properties were provided by the introduction of a sulfonic acid residue or by phosphorylation of a primary hydroxy group attached at C-4 of the 2,2,4-trimethyl-1,2-dihydroquinoline fragment fused to the coumarin fluorophore. For use in immunolabeling procedures, the dyes were decorated with an (activated) carboxy group. The positions of the absorption and emission maxima vary in the ranges 413-480 and 527-668 nm, respectively. The phosphorylated dye, 9,CH=CH-2-py,H, with the 1-(3-carboxypropyl)-4-hydroxymethyl-2,2-dimethyl-1,2-dihydroquinoline fragment fused to the coumarin fluorophore bearing the 3-[2-(2-pyridyl)ethenyl] residue (absorption and emission maxima at 472 and 623 nm, respectively) was used in super-resolution light microscopy with stimulated emission depletion and provided an optical resolution better than 70 nm with a low background signal. As a result of their large Stokes shifts, good fluorescence quantum yields, and adequate photostabilities, phosphorylated coumarins enable two-color imaging (using several excitation sources and a single depletion laser) to be combined with subdiffractional optical resolution.

The enthalpies of formation [Delta(g)] of tricyclo[8.2.2.2(4,7)]hexadeca-1(13),2,4(16),5,7(15),10(14),11-heptaene (2, 1,2-dehydro[2.2]paracyclophane or [2.2]paracyclophane-1-ene) and tricyclo[8.2.2.2(4,7)]hexadeca-1(13),2,4(16),5,7(15),8,10(14),11-octaene (3, 1,2,9,10-dehydro[2.2]paracyclophane or [2.2]paracyclophane-1,9-diene) have been determined by measuring their heats of combustion in a microcalorimeter and their heats of sublimation by the transpiration method. Values of the strain energies (SE) [SE(2) = 34.7 kcal mol(-)(1), SE(3) = 42.0 kcal mol(-)(1)] have been derived from the gas-phase heats of formation and are compared with those from MM3 and PM3 calculations and with the corresponding value SE(1) = 30.1 kcal mol(-)(1) for the parent tricyclo[8.2.2.2(4,7)]hexadeca-1(13),4(16),5,7(15),10(14),11-hexaene (1, [2.2]paracyclophane). The higher strain energies of 2 and 3 (by 4.6 and 11.9 kcal mol(-)(1)) are in accord with the well-known increased reactivities of their aromatic rings as a consequence of their increased bending. As revealed by an X-ray crystal structure analysis, the bending in the monoene 2 corresponds to that of 1 and 3 at one of two bridging corners.

EBC-23 (2), a prostate anticancer agent, was isolated from the fruit of Cinnamomum laubatii (family Lauraceae) in the Australian tropical rainforest. Extensive NOE experiments enabled the relative stereochemistry of the proposed EBC-23 (2) structure to be determined. Total synthesis of both enantiopodes over nine linear steps, involving challenging RCM and spiroacetal cyclizations, confirmed the gross structure and relative and absolute stereochemistry.

EBC-23, 24, 25, 72, 73, 75 and 76 were isolated from the fruit of Cinnamomum laubatii (family Lauraceae) in the Australian tropical rainforests. EBC-23 (1) was synthesized stereoselectively, in nine linear steps in 8 % overall yield, to confirm the reported relative stereochemistry and determine the absolute stereochemistry. Key to the total synthesis was a series of Tietze-Smith linchpin reactions. The novel spiroacetal structural motif, exemplified by EBC-23 (1), was found to inhibit the growth of the androgen-independent prostate tumor cell line DU145 in the mouse model, indicating potential for the treatment of refractory solid tumors in adults.

Diastereomeric meso‐ and d,l‐bis(bicyclopropylidenyl) (5) were obtained upon oxidation with oxygen of a higher‐order cuprate generated from lithiobicyclopropylidene (4) in 50 and 31 % yield, respectively. Their perdeuterated analogues meso‐[D14]‐ and d,l‐[D14]‐5 were obtained along the same route from perdeuterated bicyclopropylidene [D8]‐3 (synthesized in six steps in 7.4 % overall yield from [D8]‐THF) in 20.5 % yield each. Dehalogenative coupling of 1,1‐dibromo‐2‐cyclopropylcyclopropane (6) gave a mixture of all possible stereoisomers of 1,5‐dicyclopropylbicyclopropylidene 16 in 69 % yield, from which (Z)‐cis‐16 was separated by preparative gas chromatography (26 % yield). The crystal structure of meso‐5 looks like a superposition of the crystal structures of two outer bicyclopropylidene units (3) and one inner s‐trans‐bicyclopropyl unit, whereas the two outer cyclopropyl moieties adopt a gauche orientation with respect to the cyclopropane rings at the inner bicyclopropylidene units in (Z)‐cis‐16. Birch reduction with lithium in liquid ammonia of meso‐5 and d,l‐5 gave two pairs of diastereomeric quatercyclopropanes trans,trans‐(R*,S*,R*, S*)‐17/cis,trans‐(R*,S*,R*,R*)‐18 and trans,trans‐(R*,S*,S*,R*)‐19/cis,trans‐(R*,S*,S*,S*)‐20 in 97 and 76 % yield, respectively, in a ratio 9:1 for every pair. The latter diastereomer was also obtained as the sole product by Birch reduction of (Z)‐cis‐16 in 96 % yield. Under the same conditions, tetradecadeuterio analogues trans,trans‐[D14]‐(R*,S*,R*,S*)‐17/cis,trans‐[D14]‐(R*, S*,R*,R*)‐18 (8:1) and trans,trans‐[D14]‐(R*,S*,S*,R*)‐19/cis,trans‐[D14]‐(R*,S*,S*,S*)‐20 (12:1) were prepared from meso‐[D14]‐5 and d,l‐[D14]‐5 in 37 and 63 % yield, respectively. Reduction of meso‐5 with diimine gave the cis,cis‐quatercyclopropane (S*,S*,R*,R*)‐21 as the main product (58 % yield) along with the cis,trans‐diastereomer (S*,S*,R*,S*)‐18 (29 % yield). Thus, five of the six possible diastereomeric quatercyclopropanes were obtained from meso‐5, d,l‐5, and (Z)‐cis‐16. The X‐ray crystal structure analyses of trans,trans‐(R*,S*,R*,S*)‐17 and cis,cis‐(S*,S*,R*,R*)‐21 revealed for the both an unusual conformation in which the central bicyclopropyl unit adopts an s‐trans‐(antiperiplanar) orientation with ϕ=180.0°, and the two terminal bicyclopropyl moieties adopt a synclinal conformation with ϕ=49.8 and 72.0°, respectively. In solution the vicinal coupling constants 〈3JH,H〉 in trans,trans‐(R*,S*,R*,S*)‐[D14]‐17, trans,trans‐(R*,S*,S*,R*)‐[D14]‐19, trans,cis‐(R*,S*,R*,R*)‐[D14]‐18 and trans,cis‐(R*,S*,S*,S*)‐[D14]‐20 were found to be 4.1, 4.7, 5.9 and 5.9 Hz, respectively. This indicates a predominance of the all‐gauche conformer in (R*,S*,R*,S*)‐17 and a decreasing fraction of it in this sequence of the other diastereomers.