Our aim was to assess the feasibility of a theranostic strategy that combines positron emission tomography (PET) with [64Cu]Cu-DOTA-panitumumab F(ab')2 to image HNSCC and predict the radiation equivalent doses to the tumour and normal organs from RIT with [177Lu]Lu-DOTA-panitumumab F(ab')2.Epidermal growth factor receptors (EGFR) are overexpressed on many head and neck squamous cell carcinoma (HNSCC). Radioimmunotherapy (RIT) with F(ab')2 were conjugated to DOTA and complexed to 64Cu or 177Lu in high radiochemical purity  and exhibited high affinity EGFR binding (Kd = 2.9 ± 0.7 × 10− 9 mol/L). Biodistribution (BOD) studies at 6, 24 or 48 h post-injection (p.i.) of [64Cu]Cu-DOTA-panitumumab F(ab')2  or [177Lu]Lu-DOTA-panitumumab F(ab')2  in NRG mice with s.c. HNSCC patient-derived xenografts (PDX) overall showed no significant differences in tumour uptake but modest differences in normal organ uptake were noted at certain time points. Tumours were imaged by microPET/CT with [64Cu]Cu-DOTA-panitumumab F(ab')2 or microSPECT/CT with [177Lu]Lu-DOTA-panitumumab F(ab')2 but not with irrelevant [177Lu]Lu-DOTA-trastuzumab F(ab')2. Tumour uptake at 24 h p.i. of [64Cu]Cu-DOTA-panitumumab F(ab')2 [14.9 ± 1.1% injected dose/gram (%ID/g) and [177Lu]Lu-DOTA-panitumumab F(ab')2 (18.0 ± 0.4%ID/g) were significantly higher (P < 0.05) than [177Lu]Lu-DOTA-trastuzumab F(ab')2 (2.6 ± 0.5%ID/g), demonstrating EGFR-mediated tumour uptake. There were no significant differences in the radiation equivalent doses in the tumour and most normal organs estimated for [177Lu]Lu-DOTA-panitumumab F(ab')2 based on the BOD of [64Cu]Cu-DOTA-panitumumab F(ab')2 compared to those estimated directly from the BOD of [177Lu]Lu-DOTA-panitumumab F(ab')2 except for the liver and whole body which were modestly underestimated by [64Cu]Cu-DOTA-panitumumab F(ab')2. Region-of-interest (ROI) analysis of microPET/CT images provided dose estimates for the tumour and liver that were not significantly different for the two radioimmunoconjugates. Human doses from administration of [177Lu]Lu-DOTA-panitumumab F(ab')2 predicted that a 2 cm diameter HNSCC tumour in a patient would receive 1.1–1.5 mSv/MBq and the whole body dose would be 0.15–0.22 mSv/MBq.Panitumumab F(ab')64Cu]Cu-DOTA-panitumumab F(ab')2 to image HNSCC tumours and predict the equivalent radiation doses in the tumour and normal organs from RIT with [177Lu]Lu-DOTA-panitumumab F(ab')2 is feasible. RIT with [177Lu]Lu-DOTA-panitumumab F(ab')2 may be a promising approach to treatment of HNSCC due to frequent overexpression of EGFR.A PET theranostic strategy combining [The online version contains supplementary material available at 10.1186/s41181-021-00140-1. In the es Chow, . For adves Chow, . Howeveres Chow,  and grades Chow, . ReductiThe epidermal growth factor receptor (EGFR) is overexpressed on 38–47% of HNSCC tumours and is a poor prognostic marker , Eβmax = 0.38 MeV (9.1%), Eβmax = 0.18 MeV (12.2%)] to anti-EGFR mAbs may be a promising approach for HNSCC, considering that EGFR are frequently overexpressed on tumours  with the corresponding [64Cu]Cu-labeled mAbs [Eβ+max = 0.65 MeV (17.4%)] in a “PET theranostic” strategy. An analogous SPECT theranostic strategy that uses [177Lu]Lu-labeled panitumumab to select patients for RIT is not as attractive due to the low abundance γ-photons emitted by 177Lu [Eγ = 113 keV (6%) and 208 keV (10%)] which combined with the collimation required for SPECT decreases the sensitivity for tumour imaging. In addition, the higher absorbed doses for 177Lu compared to 64Cu are not suitable for a theranostic approach in which imaging is performed to select patients for RIT. Although many HNSCC are EGFR-positive, PET with [64Cu]Cu-labeled mAbs would confirm lesions as EGFR-positive and importantly predict the tumour and normal tissue uptake of [177Lu]Lu-labeled anti-EGFR mAbs. Moreover, information on the tumour and normal organ uptake can be used to estimate the radiation equivalent doses from RIT which may predict the effectiveness and normal organ toxicity in an individual patient. Since 64Cu has a short half-life (t1/2 = 12.7 h), F(ab')2 fragments of mAbs that accumulate in tumours but are quickly eliminated from the blood and most normal organs in the feasible imaging time of 64Cu [up to 24 h post-injection (p.i.)] would be most attractive for a PET theranostic strategy. F(ab')2 also offer advantages for RIT because their more rapid elimination compared to IgG quickly decreases circulating radioactivity which is responsible for dose-limiting bone marrow toxicity from RIT due to the cross-fire effect from the 2 mm β-particles emitted by 177Lu  triacetic acid) imaged s.c. PANC-1 human pancreatic cancer tumours that overexpress EGFR in non-obese diabetic severe combined immunodeficiency (NOD/scid) mice 2 and Fab than for the intact IgG, but kidney uptake was much higher for Fab than F(ab')2, thus F(ab')2 are preferred for PET and RIT. [177Lu]Lu-DOTA-cetuximab F(ab')2 have been reported to be effective for RIT of human colorectal xenografts in female SWISS nu/nu mice  which links the β-particle-emitting radionuclide 64Cu]Cu-DOTA-panitumumab F(ab')2 must be very similar to that of [177Lu]Lu-DOTA-panitumumab F(ab')2. Moreover, the radiation equivalent doses in the tumour and normal organs predicted for [177Lu]Lu-DOTA-paniumumab F(ab')2 based on the biodistribution (BOD) of [64Cu]Cu-DOTA-panitumumab F(ab')2 assessed by PET need to accurately predict those estimated directly from the BOD of [177Lu]Lu-DOTA-panitumumab F(ab')2. Our aim was to compare the BOD of [64Cu]Cu-DOTA-panitumumab F(ab')2 and [177Lu]Lu-DOTA-panitumumab F(ab')2 in NOD-Rag1nullIL2rgnull (NRG) mice with subcutaneous (s.c.) patient-derived HNSCC tumour xenografts (PDX) and estimate the radiation equivalent doses in the tumour and normal organs for [177Lu]Lu-DOTA-panitumumab F(ab')2 based on the tumour and normal organ uptake of [64Cu]Cu-DOTA-panitumumab F(ab')2 assessed in BOD studies or by region-of-interest (ROI) analysis of microPET/CT images. We compared these predicted doses based on [64Cu]Cu-DOTA-panitumumab F(ab')2 to those estimated directly from the BOD of [177Lu]Lu-DOTA-panitumumab F(ab')2 to assess the feasibility of a PET theranostic strategy. The PDX mouse model of HNSCC used in this study is clinically relevant because these PDX recapitulate the properties of HNSCC tumours in patients   and engrafted subcutaneously (s.c.) on the right flank of NOD-Rag1nullIL2rgnull (NRG) mice. These patient-derived tumour xenografts (PDX) were serially propagated in NRG mice following an Animal Care Protocol (No. 1542.28) approved by the Animal Care Committee at the University Health Network and following Canadian Council on Animal Care guidelines. The PDX used in this study were between the 3rd to 5th passage from the initial engraftment of the HNSCC tumour in NRG mice.MDA-MB-468 human breast cancer cells (1.3 × 102 (MW ~ 110 kDa) were produced by proteolytic digestion of panitumumab IgG  using immobilized pepsin  as reported with minor modifications . The 100 mM NaHCO3 buffer, pH 8.2 buffer was purified from trace metals by passage through a column of Chelex-100 cation exchange resin . The F(ab´)2 concentration was measured spectrophotometrically at 280 nm (A280 of a 1 mg/mL solution = 1.40). The purity and homogeneity of F(ab´)2 were assessed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) on a 7.5% Mini-Protean Tris/glycine mini-gel (BioRad) under reducing (2-mercaptoethanol) and non-reducing conditions with bands stained with Biosafe Coomassie Blue G-250 (BioRad). Anti-human epidermal growth factor receptor-2 (HER2) trastuzumab F(ab´)2 was prepared by proteolytic digestion of trastuzumab IgG .Panitumumab F(ab´)2  in 100 mM NaHCO3 buffer, pH 8.2 were conjugated to 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) by reaction with a 20-fold molar excess of the N-hydroxysuccinimide ester  for 2 h at room temperature (RT) on a nutating mixer . DOTA-panitumumab F(ab´)2 were purified from excess DOTA and buffer-exchanged into 100 mM HEPES, pH 5.5 and re-concentrated to 20–28.5 mg/mL using an Amicon ultracentrifugal unit (MWCO = 10 kDa) centrifuged at 7500×g for 5 mins. This was repeated 8 times to ensure complete removal of excess DOTA. The DOTA substitution level [moles DOTA/mole F(ab´)2] was determined by trace-labeling an aliquot  of the impure conjugation mixture with [64Cu]CuCl2  and measuring the proportion of [64Cu]Cu-DOTA-panitumumab F(ab´)2 and [64Cu]Cu-DOTA by instant thin layer-silica gel chromatography  developed in 100 mM sodium citrate, pH 5.5, then multiplying this proportion by the 20:1 M ratio of NHS-DOTA:F(ab´)2 in the reaction 2 and free [64Cu]Cu-DOTA (or 64Cu) were 0.0 and 1.0, respectively.Panitumumab F(ab´) Reilly, . The Rf 2 (~ 600 μg in 30 μL) were labeled with 64Cu to a specific activity of 0.074–1.7 MBq/μg by incubation with [64Cu]CuCl2  for 1 h at 42 °C. No post-labeling purification was performed. The radiochemical purity (RCP) of [64Cu]Cu-DOTA-panitumumab F(ab´)2 was confirmed by ITLC and size exclusion (SE)-HPLC on an Agilent 1260 Infinity II HPLC system fitted with a BioSep SEC-S4000 column  eluted with NaH2PO4 buffer, pH 7.0 at a flow rate of 0.50 mL/min with UV detection (λ = 280 nm) and radioactivity detection by a Flowstar LB514 radioactivity detector fitted with a BGO-X flow cell . DOTA-panitumumab F(ab´)2 (~ 600 μg in 30 μL) were labeled at a SA = 0.07–0.2 MBq/μg with 177Lu by incubation wih [177Lu]LuCl3  at 42 °C for 3 h. The RCP of [177Lu]Lu-DOTA-panitumumab F(ab´)2 was determined by ITLC as described for [64Cu]Cu-DOTA-panitumumab F(ab´)2. Trastuzumab F(ab´)2 were similarly conjugated to DOTA and labeled with 177Lu.Purified DOTA-panitumumab F(ab´)d) and maximum number of binding sites (Bmax) for binding of [177Lu]Lu-DOTA-panitumumab F(ab´)2 to EGFR on MDA-MB-468 human breast cancer cells (1.3 × 106 EGFR/cell) 2 (0.098–200 nmoles/L) were incubated with 1 × 106 MDA-MB-468 cells in 200 μL of PBS, pH = 7.4 in 1.5 mL Eppendorf tubes at 4 °C for 3 h with gentle shaking every 30 mins. The samples were then centrifuged at 1000×g for 2 mins on an Eppendorf Centrifuge 5424  and the supernatant containing unbound 177Lu collected. The cell pellets were rinsed with ice cold PBS, pH 7.4, centrifuged again and the supernatant was collected and pooled with the previously collected supernatant. This was repeated twice. Total bound 177Lu (TB) in the cell pellets and unbound 177Lu in the supernatant were measured in a γ-counter . Non-specifically bound 177Lu (NSB) was assessed by repeating the assay in the presence of 50-fold molar excess of panitumumab IgG. Specifically bound 177Lu (SB) was calculated by subtracting NSB from TB. [177Lu]Lu-DOTA-panitumumab F(ab´)2 bound to MDA-MB-468 cells (pmoles) was plotted vs. the concentration of free (unbound) [177Lu]Lu-DOTA-panitumumab F(ab´)2 (nmoles/L) and the curve was fitted to a one-site-receptor-binding model using Prism Ver. 4.0 software .A saturation radioligand binding assay was performed to measure the dissociation constant (K64Cu]Cu-DOTA-panitumumab F(ab´)2 were determined by BOD and microPET/CT studies in NRG mice with subcutaneous (s.c.) HNSCC PDX. Groups of 3–4 tumour-bearing NRG mice were injected i.v. (tail vein) with [64Cu]Cu-DOTA-panitumumab F(ab´)2  and sacrificed at 6 h, 24 or 48 h post-injection (p.i.). The tumour and samples of blood and normal tissues were obtained, weighed and the radioactivity in each measured in a γ-counter using a window (425–640 keV) to include the 511 keV annihilation γ-photon of 64Cu. Tumour and normal organ uptake were expressed as percent injected dose/g (%ID/g). MicroPET/CT was performed after i.v. injection (tail vein) of 37 MBq (50 μg) of [64Cu]Cu-DOTA-panitumumab F(ab´)2 in a group of 4 NRG mice with s.c. HNSCC PDX. Mice were anaesthetized using 2% isoflurane in O2 and were imaged in a supine position on a NanoScan® SPECT/CT/PET system . PET images were acquired for 10, 20 and 40 mins at 6, 24 and 48 h p.i. of [64Cu]Cu-DOTA-panitumumab F(ab´)2, respectively. Images were reconstructed by an ordered subset expectation maximization (OSEM) algorithm and consisted of 4 subsets and 4 iterations with attenuation and scatter correction supported by an isotropic voxel size of 300 μm. Prior to PET, CT images were acquired with 50 kVp X-rays, 980 μA and 300 msec exposure time. CT scans were reconstructed using the medium voxel and slice thickness settings, resulting in an isotropic voxel size of 250 μm. PET and CT images were co-registered by the Mediso Nucline NanoScan 3.00.020.0000 software. PET and CT DICOM files were exported using Mediso’s Nucline Acquisition/Reconstruction Software to the Inveon Research Workplace Software 4.0 (Siemens) for analysis and quantification of 64Cu uptake (%ID/g) as well as estimation of the volume of the liver and tumour. Regions of interest (ROI) were drawn around the tumour and liver on the PET images aided by delineation of the anatomy on the CT images to compare the accuracy of PET for quantifying the tumour and liver uptake compared to BOD studies. The ROI for each organ was drawn two-dimensionally on axial slices of the image. To ensure the coverage of the entire organ, > 5 ROIs were drawn before the slices were convoluted to form a 3D volume that encompassed the entire organ.The tumour and normal tissue uptake of [177Lu]Lu-DOTA-panitumumab F(ab´)2 was determined by BOD and microSPECT/CT studies. [177Lu]Lu-DOTA-panitumumab F(ab´)2  were injected i.v. (tail vein) in NRG mice with s.c. HNSCC PDX and groups (n = 3–4) of mice were sacrificed at 6 h, 24 h or 48 h p.i. The tumour, and samples of blood and other tnormal issues were obtained, weighed and counted in a γ-counter using a window (130–470 kev) to include the γ-photons of 177Lu [Eγ = 113 keV (6.6%) and Eγ = 208 keV (11%)]. Tumour and normal organ uptake was expressed as %ID/g. The excised tumours were subjected to immunohistochemical (IHC) staining with anti-human EGFR antibodies  to confirm EGFR positivity. In addition, the tumour and normal tissue BOD of irrelevant [177Lu]Lu-DOTA-trastuzumab F(ab´)2  were determined in a separate group of 5 NRG mice bearing HNSCC PDX at 24 h p.i. The excised tumours in these mice were stained for HER2 using anti-human HER2 antibodies (Invitrogen Cat. No. MA5–14509). Representative mice were anaesthetized using 2% isoflurane in O2 and microSPECT/CT images were acquired in a supine position, at 6 h, 24 h and 48 h p.i. of [177Lu]Lu-DOTA-panitumumab F(ab´)2 on a NanoScan® SPECT/CT/PET system (Mediso). Images were acquired in a 256 × 256 matrix. A window (± 10%) was set around each of the γ-photopeaks  of 177Lu. A Mediso APT62 collimator (WB-HS standard) was affixed to each of the 4 detector NaI (TI) detector heads. Images were reconstructed by Monte Carlo methods with three subsets of data undergoing 48 iterations using the Mediso Nucline NanoScan acquisition and reconstruction software (ver 3.00.020.0000). Prior to SPECT, imaging CT images were acquired with 50 kVp X-rays, 980 μA and a 300 msec exposure time. CT scans were reconstructed using the medium voxel and slice thickness settings resulting in an isotropic voxel size of 250 μm. SPECT and CT were co-registered by the Mediso Nucline acquisition/reconstruction software. MicroSPECT/CT images were similarly obtained for NRG mice with HNSCC PDX at 24 h p.i. of [177Lu]Lu-DOTA-trastuzumab F(ab´)2. All animal studies were conducted under a protocol (AUP 2843.8) approved by the Animal Care Committee at the University Health Network following Canadian Council on Animal Care guidelines.The tumour and normal tissue uptake of [64Cu]Cu-DOTA-panitumumab F(ab´)2 or [177Lu]Lu-DOTA-panitumumab F(ab´)2 were estimated from the tumour and normal organ uptake of activity in source organs measured in BOD studies. The mean equivalent dose (D) was calculated as D = Ãs × S × WR, where Ãs is the time-integrated activity in the source organs or tumour and S are the Snyder values for mice  from the area-under-the-curve (AUC) up to 48 h p.i. of [64Cu]Cu-DOTA-panitumumab F(ab´)2 or [177Lu]Lu-DOTA-panitumumab F(ab´)2  derived from the activity vs. time curves. The activity/source organ at each time point t (s) was calculated using the formula %ID/g (t) × ID/100 × organ weight × exp.(−kt), where %ID/g was obtained from the BOD studies, ID was the injected dose in Bq, and k was the decay constant for 64Cu (1.52 × 10− 5 s− 1) or 177Lu (1.21 × 10− 6 s − 1). The time integrated activity from 48 h p.i. to infinity  was calculated by dividing the activity at 48 h p.i. by the decay constant for 64Cu or 177Lu, assuming further elimination of activity from source organs only by radioactive decay. The S-value for the tumour was estimated using the sphere model in OLINDA/EXM software based on the measured tumour mass 2. To predict the doses in normal organs in humans, the activity in source organs of a human adult female with a 2 cm diameter spherical tumour in the neck at the different times up to 48 h p.i. of the RICs were estimated based on proportional extrapolation of the activities in mice using the % kg/g method, i.e. (%ID/organ)human = [(%ID/organ)mouse × (mouse body weight/human body weight)]  and from 48 h p.i. to infinity  were calculated as described earlier for mice, and the combined A0h to ∞ for each human source organ was used to predict the equivalent doses in human female adults (mSv/Bq) using OLINDA/EXM 1.0 software 2 and [177Lu]Lu-DOTA-panitumumab F(ab´)2 were made using a Mann-Whitney U test (P < 0.05) and Prism 4.0 software (GraphPad). Statistical comparisons for radiation equivalent dose were made using a two-tailed unpaired Student’s t-test (P < 0.05) using PrismVersion 4.0 software (GraphPad).All results were reported as mean ± SEM. Statistical comparisons for normal organ and tumour uptake of Cu-DOTA-panitumumab F(ab´)2 2 (not shown). [177Lu]Lu-DOTA-panitumumab F(ab´)2 exhibited saturable binding to EGFR on MDA-MB-468 cells that was displaced by a 50-fold molar excess of panitumumab IgG   measured by ITLC-SG. SDS-PAGE analysis 2and a major band with the expected Mr ~ 150 kDa for trastuzumab IgG under non-reducing conditions. Under reducing conditions, trastuzumab F(ab´)2 migrated as a single band at Mr ~ 25 kDa, corresponding to the variable and constant region of the heavy chain (VH-CH) and the light chain (VL-CL) while trastuzumab IgG migrated as two main bands with Mr ~ 50 kDa and 25 kDa, corresponding to the dissociated heavy and light chains, respectively.SE-HPLC with UV detection at 280 nm demonstrated a single peak for panitumumab F(ab´)sis Fig.  revealed64Cu]Cu-DOTA-panitumumab F(ab´)2 in NRG mice with s.c. HNSCC PDX 2 2 was similar to [64Cu]Cu-DOTA-panitumumab F(ab´)2. At 6 h p.i., blood pool activity (mediastinum) and uptake into the liver were observed. Blood pool activity decreased greatly at 24 and 48 h p.i. and liver uptake diminished slightly. The tumour was visualized at all time points. MicroSPECT/CT images of NRG mice bearing the same s.c. HNSCC PDX were obtained at 24 h p.i. of [177Lu]Lu-DOTA-trastuzumab F(ab´)2 2 was similar to [177Lu]Lu-DOTA-panitumumab F(ab´)2 at 24 h p.i. but with higher kidney uptake. Tumours were not well-visualized by imaging with [177Lu]Lu-DOTA-trastuzumab F(ab´)2 in agreement with very low HER2 expression determined by IHC staining 2 or [177Lu]Lu-DOTA-panitumumab F(ab´)2 at 6, 24 or 48 h p.i. are shown in Table 64Cu]Cu-DOTA-panitumumab F(ab´)2 was 9.3 ± 1.0%ID/g at 6 h p.i., but increased to 14.9 ± 1.1%ID/g at 24 h p.i., then declined to 9.8 ± 1.3%ID/g at 48 h p.i. Similarly, tumour uptake of [177Lu]Lu-DOTA-panitumumab F(ab´)2 was 10.4 ± 2.0%ID/g at 6 h p.i., increased to 18.0 ± 0.4%ID/g at 24 h p.i., then declined to 15.3 ± 2.6%ID/g at 48 h p.i. There were no significant differences in tumour uptake of [64Cu]Cu-DOTA-panitumumab F(ab´)2 compared to [177Lu]Lu-DOTA-panitumumab F(ab´)2 at any time points. Activity in the blood for mice injected with [64Cu]Cu-DOTA-panitumumab F(ab´)2 decreased from 10.6 ± 0.6%ID/g at 6 h p.i. to 4.8 ± 0.3%ID/g at 24 h p.i., then to 1.9 ± 0.1%ID/g at 48 h p.i. For mice injected with [177Lu]Lu-DOTA-panitumumab F(ab´)2, blood activity decreased from 15.3 ± 2.0%ID/g at 6 h p.i. to 4.5 ± 0.1%ID/g at 24 h p.i. then to 1.1 ± 0.2%ID/g at 48 h p.i. Blood activity was not significantly lower for [64Cu]Cu-DOTA-panitumumab F(ab´)2 than [177Lu]Lu-DOTA-panitumumab F(ab´)2 at 6 h p.i. but was significantly higher (P = 0.03) at 48 h p.i.. The tumour/blood (T/B) ratio for [64Cu]Cu-DOTA-panitumumab F(ab´)2 increased from 0.9 ± 0.1 at 6 h p.i. to 3.1 ± 0.1 at 24 h p.i. then to 5.1 ± 0.8 at 48 h p.i. The T/B ratio for [177Lu]Lu-DOTA-panitumumab F(ab´)2 increased from 0.7 ± 0.1 at 6 h p.i. to 4.0 ± 0.1 at 24 h p.i. then to 14.7 ± 2.8 at 48 h p.i. The T/B ratio was significantly greater for [177Lu]Lu-DOTA-panitumumab F(ab´)2 at 48 h p.i. than [64Cu]Cu-DOTA-panitumumab F(ab´)2 (P = 0.03) but not at 6 or 24 h p.i. At 6 h p.i., there was low uptake in normal tissues in mice injected with [64Cu]Cu-DOTA-panitumumab F(ab´)2 and normal tissue uptake was not significantly different than [177Lu]Lu-DOTA-panitumumab F(ab´)2. (Table 177Lu]Lu-DOTA-panitumumab F(ab´)2 than [64Cu]Cu-DOTA-panitumumab F(ab´)2. Liver uptake was insignificantly higher for [177Lu]Lu-DOTA-panitumumab F(ab´)2. At 48 h p.i., there was significantly higher uptake in the blood and several normal tissues including the liver for [64Cu]Cu-DOTA-panitumumab F(ab´)2 than [177Lu]Lu-DOTA-panitumumab F(ab´)2 (P < 0.05). However, skin and muscle uptake were significantly higher for [177Lu]Lu-DOTA-panitumumab F(ab´)2. The tumour and normal tissue BOD of irrelevant [177Lu]Lu-DOTA-trastuzumab F(ab´)2 is also shown in Table 177Lu]Lu-DOTA-trastuzumab F(ab´)2 at 24 h p.i. was 2.6 ± 0.5%ID/g, which was 5.7 times (P = 0.04) and 6.9 times (P = 0.04) significantly lower than [64Cu]Cu-DOTA-panitumumab F(ab´)2 and [177Lu]Lu-DOTA-panitumumab F(ab´)2, respectively. Kidney uptake of [177Lu]Lu-DOTA-trastuzumab F(ab´)2 (27.4 ± 2.7%ID/g) was significantly higher than [64Cu]Cu-DOTA-panitumumab F(ab´)2  or [177Lu]Lu-DOTA-panitumumab F(ab´)2 . Liver uptake of [177Lu]Lu-DOTA-trastuzumab F(ab´)2 (8.0 ± 1.0%ID/g) was significantly lower than [177Lu]Lu-DOTA-panitumumab F(ab´)2  but not significantly different than [64Cu]Cu-DOTA-panitumumab F(ab´)2 . A comparison of the tumour and liver uptake of [64Cu]Cu-DOTA-panitumumab F(ab´)2 measured by ROI analysis of microPET/CT images or BOD studies is shown in Table 64Cu]Cu-DOTA-panitumumab F(ab´)2 at 48 h p.i.The tumour and normal tissue BOD of [ab´)2 or 77LuLu-DOab´)2 or 77LuLu-DOab´)2 or 77LuLu-DO64Cu]Cu-DOTA-panitumumab F(ab´)2 or [177Lu]Lu-DOTA-panitumumab F(ab´)2 in NRG mice with s.c. HNSCC PDXs was used to estimate the radiation equivalent doses (Table 177Lu]Lu-DOTA-panitumumab F(ab´)2 were estimated from the BOD of [64Cu]Cu-DOTA-panitumumab F(ab´)2 for comparison with those estimated from the BOD of [177Lu]Lu-DOTA-panitumumab F(ab´)2. We first calculated the doses from [64Cu]Cu-DOTA-panitumumab F(ab´)2 in NRG tumour-bearing mice. The liver was the normal organ which received the highest dose (0.10 ± 0.02 Sv/MBq) followed by the kidneys (0.07 ± 0.01 Sv/MBq), lungs (0.05 ± 0.01 Sv/MBq), spleen (0.04 ± 0.01 Sv/MBq), stomach (0.03 ± 0.01 Sv/MBq), heart (0.03 ± 0.01 Sv/MBq) and intestine (0.02 ± 0.00 Sv/MBq). The tumour received the highest dose (0.14 ± 0.03 Sv/MBq). The whole body equivalent dose was 0.05 ± 0.00 Sv/MBq. The doses for [177Lu]Lu-DOTA-panitumumab F(ab´)2 exhibited a similar trend but were higher. Among the normal organs, the highest doses were in the liver (1.82 ± 0.14 Sv/MBq), kidneys (0.75 ± 0.08 Sv/MBq), spleen (0.66 ± 0.18 Sv/MBq), lungs (0.38 ± 0.04 Sv/MBq), heart (0.32 ± 0.13 mSv/MBq), stomach (0.27 ± 0.04 mSv/MBq), intestines (0.21 ± 0.05 mSv/MBq) and pancreas (0.16 ± 0.04 Sv/MBq). The tumour dose and whole body dose were 2.5 ± 0.8 Sv/MBq and 0.34 ± 0.02 Sv/MBq, respectively which were about 7 times higher than those for [64Cu]Cu-DOTA-panitumumab F(ab´)2.The BOD of [es Table . In addi177Lu]Lu-DOTA-panitumumab F(ab´)2 based on the BOD of [64Cu]Cu-DOTA-panitumumab F(ab´)2 were made for comparison (Table 177Lu]Lu-DOTA-panitumumab F(ab´)2 based on the BOD of [64Cu]Cu-DOTA-panitumumab F(ab´)2 were not significantly different than those estimated from the BOD of [177Lu]Lu-DOTA-panitumumab F(ab´)2 except for the liver (P = 0.03) and whole body (P = 0.05).Estimates of the radiation equivalent doses for [on Table . The hig64Cu]Cu-DOTA-panitumumab F(ab´)2 or [177Lu]Lu-DOTA-panitumumab F(ab´)2 and were compared to doses calculated from BOD studies of [64Cu]Cu-DOTA-panitumumab F(ab´)2 or [177Lu]Lu-DOTA-panitumumab F(ab´)2. ROI-based estimates of the doses for [64Cu]Cu-DOTA-panitumumab F(ab´)2 in the tumour (0.12 ± 0.04 Sv/MBq) and liver (0.14 ± 0.02 Sv/MBq) did not differ significantly from those based on BOD studies of [64Cu]Cu-DOTA-panitumumab F(ab´)2 2 in the tumour (2.3 ± 0.4 Sv/MBq) and liver (1.45 ± 0.17 Sv/MBq) were not significantly different than those based on BOD studies of [64Cu]Cu-DOTA-panitumumab F(ab´)2 (2.0 ± 0.6 Sv/MBq and 1.22 ± 0.13 Sv/MBq espectively) or [177Lu]Lu-DOTA-panitumumab F(ab´)2 2 or [177Lu]Lu-DOTA-panitumumab F(ab´)2 were projected based on BOD studies or ROI analysis of PET images in mice with s.c. HNSCC PDX (Supplementary Table 64Cu]Cu-DOTA-panitumumab F(ab´)2 deposited the highest doses in the liver (0.065 ± 0.012 mSv/MBq), kidneys (0.043 ± 0.006 mSv/MBq), intestines (0.037 ± 0.002 mSv/MBq) and tumour (0.092 ± 0.017 mSv/MBq). The whole body dose was 0.030 ± 0.002 mSv/MBq. Doses based on ROI analysis of the images were estimated for the liver (0.080 ± 0.009 mSv/MBq) and tumour (0.065 ± 0.016 mSv/MBq). These were similar to those estimated using the BOD studies. Using the BOD, [177Lu]Lu-DOTA-panitumumab F(ab´)2 deposited the highest doses in the liver (1.05 ± 0.08 mSv/MBq), kidneys (0.44 ± 0.04 mSv/MBq), spleen (0.39 ± 0.11 mSv/MBq) and tumour (1.47 ± 0.46 mSv/MBq). The whole body dose was 0.22 ± 0.02 mSv/MBq. The doses deposited by [177Lu]Lu-DOTA-panitumumab F(ab´)2 were higher than those from [64Cu]Cu-DOTA-panitumumab F(ab´)2. The doses for [177Lu]Lu-DOTA-panitumumab F(ab´)2 based on BOD or ROI analysis of the PET images of [64Cu]Cu-DOTA-panitumumab F(ab´)2 were similar to those estimated from the BOD of [177Lu]Lu-DOTA-panitumumab F(ab´)2 except for the liver  and tumour  which were modestly underestimated 2 based on BOD studies and ROI analysis of microPET/CT images in NRG mice with s.c. HNSCC PDX tumours after i.v. administration of [64Cu]Cu-DOTA-panitumumab F(ab´)2. Our results indicate that a PET theranostic strategy employing [64Cu]Cu-DOTA-panitumumab F(ab´)2 to image HNSCC tumours and predict the doses to the tumour and normal organs from RIT with [177Lu]Lu-DOTA-panitumumab F(ab´)2 is feasible. The doses in the tumour and normal organs for [177Lu]Lu-DOTA-panitumumab F(ab´)2 that were estimated from the BOD studies of [64Cu]Cu-DOTA-panitumumab F(ab´)2 were not significantly different than those estimated directly from the BOD of [177Lu]Lu-DOTA-panitumumab F(ab´)2 except for the liver and tumour, which were modestly underestimated by [64Cu]Cu-DOTA-panitumumab F(ab´)2 (Tables 64Cu]Cu-DOTA-panitumumab F(ab´)2 2 2 Fig.  or micro2 were produced from panitumumab IgG in high purity    of [177Lu]Lu-DOTA-panitumumab F(ab´)2 to EGFR-positive MDA-MB-468 human breast cancer cells 2 was 3-fold higher than we previously reported for [177Lu]Lu-DOTA-panitumumab IgG (Kd = 1.0 ± 0.4 10− 9 mol/L) 2 but substitution of 64Cu for 177Lu is not expected to change the affinity. Irrelevant F(ab´)2 were produced in high purity from trastuzumab IgG  to provide specificity control RICs for imaging and BOD studies.F(ab´)ity Fig.  and wereIgG Fig. , conjuga64Cu]Cu-DOTA-panitumumab F(ab´)2 2 2 2 and [177Lu]Lu-DOTA-panitumumab F(ab′)2 than [177Lu]Lu-DOTA-trastuzumab F(ab′)2. These results indicated that tumour imaging with [64Cu]Cu-DOTA-panitumumab F(ab´)2 or [177Lu]Lu-DOTA-panitumumab F(ab′)2 was EGFR-mediated. There was higher kidney uptake of [64Cu]Cu-DOTA-trastuzumab F(ab')2 (27.4 ± 2.7%ID/g) than [64Cu]Cu-DOTA-panitumumab F(ab')2 2 were estimated to be 8.58 and 6.71, respectively. At physiological pH = 7.4, trastuzumab F(ab')2 would be cationic and panitumumab F(ab')2 would be slightly anionic. The cationic charge of some mAb fragments is known to promote kidney uptake 2. Although PDX in NRG mice were imaged with [177Lu]Lu-DOTA-panitumumab F(ab′)2, SPECT has poor sensitivity due to the low abundance of the γ-photons emitted by 177Lu [Eγ = 113 keV (6.4%) and Eγ = 208 keV(11%)]. In addition, the dosimetry of 177Lu is not favourable for imaging. The whole body equivalent dose for [177Lu]Lu-DOTA-panitumumab F(ab′)2 in NRG mice (0.34 ± 0.02 Sv/MBq) was 7-fold higher than [64Cu]Cu-DOTA-panitumumab F(ab´)2 2 than [64Cu]Cu-DOTA-panitumumab F(ab´)2 2 Fig. a-c. Thes´)2 Fig. d. HNSCC i. Table  we estim177Lu]Lu-DOTA-panitumumab F(ab′)2 based on the BOD of [64Cu]Cu-DOTA-panitumumab F(ab´)2. BOD studies in NRG mice with s.c. HNSCC PDX at 6 or 24 p.i. showed similar uptake of these two RICs (Table 64Cu]Cu-DOTA-panitumumab F(ab´)2 and [177Lu]Lu-DOTA-panitumumab F(ab′)2. At 6 h p.i., uptake of [64Cu]Cu-DOTA-panitumumab F(ab´)2 was lower than [177Lu]Lu-DOTA-panitumumab F(ab′)2 in the intestine, spleen, kidneys and skin but higher in the blood. At 24 h p.i., lung uptake of [64Cu]Cu-DOTA-panitumumab F(ab´)2 was higher than [177Lu]Lu-DOTA-panitumumab F(ab′)2 but liver uptake was lower. At 48 h p.i., the uptake of [64Cu]Cu-DOTA-panitumumab F(ab´)2 was higher than [177Lu]Lu-DOTA-panitumumab F(ab′)2 in the lungs, stomach, pancreas, intestines and blood but lower in the liver and skin (P < 0.05). These modest differences in BOD may be due to experimental variability in tissue uptake in BOD studies. It is possible that the lower stability constant of DOTA for complexing Cu2+ (log K = 23.3)  may cause some differences in BOD between the two RICs 2 in the liver than [177Lu]Lu-DOTA-panitumumab F(ab′)2 at 24 h p.i. was unexpected, since release of 64Cu from DOTA should increase liver uptake 2 and [177Lu]Lu-DOTA-panitumumab F(ab′)2 at any time point. In patients, the tumour and normal organ uptake of [64Cu]Cu-DOTA-panitumumab F(ab´)2 would be measured by ROI analysis to predict doses from [177Lu]Lu-DOTA-panitumumab F(ab′)2. Thus, we compared the uptake of the RICs into the tumour and liver measured by ROI analysis of microPET/CT images with that measured in BOD studies  and the Ã48 h – ∞ from 48 h p.i. to infinity (Bq × sec). The calculation of Ãs tends to minimize variability in BOD between [64Cu]Cu-DOTA-panitumumab F(ab´)2 and [177Lu]Lu-DOTA-panitumumab F(ab′)2 at individual time points. Consequently, we found no significant differences in the doses estimated for [177Lu]Lu-DOTA-panitumumab F(ab′)2 based on the BOD of [64Cu]Cu-DOTA-panitumumab F(ab´)2 compared to those based on the BOD of [177Lu]Lu-DOTA-panitumumab F(ab′)2 except for the liver and whole body, which were modestly underestimated by [64Cu]Cu-DOTA-panitumumab F(ab´)2 (Table 177Lu]Lu-DOTA-panitumumab F(ab’)2 than [64Cu]Cu-DOTA-panitumumab F(ab')2 is due to the higher liver uptake in BOD studies. The reason for the higher liver uptake of [177Lu]Lu-DOTA-panitumumab F(ab')2 is not known. However, DOTA forms higher coordination number and more stable complexes with 177Lu  than 64Cu  2 to estimate the doses in the tumour or liver yielded estimates for [177Lu]Lu-DOTA-panitumumab F(ab′)2 that were not significantly different than those based on the BOD of [64Cu]Cu-DOTA-panitumumab F(ab´)2 or [177Lu]Lu-DOTA-panitumumab F(ab′)2 (Table 177Lu]Lu-DOTA-panitumumab F(ab′)2 was 1.10 ± 0.32 mSv/MBq based on the BOD in mice of [64Cu]Cu-DOTA-panitumumab F(ab´)2 and 1.47 ± 0.46 mSv/MBq based on [177Lu]Lu-DOTA-panitumumab F(ab′)2 (Supplementary Information Table 64Cu]Cu-DOTA-panitumumab F(ab´)2 and 0.22 ± 0.01 mSv/MBq based on the BOD of [177Lu]Lu-DOTA-panitumumab F(ab′)2. Fractionated doses of external radiation are used to treat HNSCC  2 and the respective doses should be determined in a future clinical trial.The radiation equivalent doses in the tumour and normal organs were calculated as D = Ã)2 Table . The hig177Lu]Lu-DOTA-panitumumab F(ab′)2 2 and 1.05 ± 0.08 mSv/Bq based on the BOD of [177Lu]Lu-DOTA-panitumumab F(ab′)2 , although the estimated dose in the liver was 6500 mSv 2 may be safe and effective for treatment of HNSCC. However, our studies projected human doses based on the BOD of [177Lu]Lu-DOTA-panitumumab F(ab′)2 in NRG mice and panitumumab does not recognize mouse EGFR 2 to image HNSCC in patients and predict the radiation equivalent doses in a tumour and normal organs from RIT with [177Lu]Lu-DOTA-panitumumab F(ab′)2 is feasible. RIT with [177Lu]Lu-DOTA-panitumumab F(ab′)2 may be a promising approach to treatment of HNSCC due to frequent overexpression of EGFR.We conclude that a PET theranostic strategy employing [Additional file 1Fig. S1. SDS PAGE analysis of trastuzumab IgG and F(ab′)2 under non-reducing  or reducing  conditions on a 7.5% Tris/Glycine mini-gel stained with Coomassie blue. Table S1. Estimated radiation equivalent doses in a tumour-bearing NRG mouse for [64Cu]Cu-DOTA-panitumumab F(ab´)2. Table S2. Projected radiation equivalent doses for a female human adult with a 2 cm tumour in the neck for [64Cu]Cu-DOTA-panitumumab F(ab´)2 or [177Lu]Lu-DOTA-panitumumab F(ab´)2. (DOCX 244 kb)


The obtained I-NON was characterized by elemental analyses , high-resolution ESI+-MS, 1H and 13C{1H} NMR, FTIR, UV-vis spectroscopy, TGA, and X-ray crystallography (XRD). The copper(II) complexes bearing deprotonated I-NON were prepared as follows: cis-[Cu(I-NON–H)(I-NON)](I3) (1) was obtained by the reaction between Cu(NON-H)2 and I2 in CHCl3/MeOH, while trans-[Cu(I-NON–H)2] (2) was synthesized from I-NON and Cu(OAc)2 in MeOH. Crystals of trans-[Cu(I-NON–H)2(THF)2] (3) and trans-[Cu(I-NON–H)2(Py)2] (4) were precipitated from solutions of 2 in CHCl3/THF and Py/CHCl3/MeOH mixtures, respectively. The structures of 1 and 3–4 were additionally verified by X-ray crystallography. The characteristic feature of the structures of 1 and 3 is the presence of intermolecular halogen bonds with the involvement of the iodine center of the metal-bound deprotonated I-NON. The nature of the I···I and I···O contacts in the structures of 1 and 3, correspondingly, were studied theoretically at the DFT (PBE0-D3BJ) level using the QTAIM, ESP, ELF, NBO, and IGM methods.The 3-Iodo-1-nitrosonaphthalene-2-ol (I-NON) was obtained by the copper(II)-mediated iodination of 1-nitroso-2-naphthol (NON). The suitable reactants and optimized reaction conditions, providing 94% NMR yield of I-NON, included the usage of Cu(OAc) The o-n sensing  and poly sensing .2 strongly depends on the identity of a halogen and, in the case of less reactive I2, this reaction commonly requires Lewis acid catalysis, e.g., application of copper(II) salts, and it was conducted mainly for electron-rich arenes  reacted with 8 equivs of molecular iodine in MeOH to form the [Cu(I-NON–H)(I-NON)](I3) (1) complex bearing the iodinated ligands . We applied this reaction as a starting point to develop a synthetic route to I-NON  was obtained by the copper(II)-mediated iodination of 1-nitroso-2-naphthol (NON). Our initial experiments demonstrated that the copper complex  (2) was obtained by the reaction of I-NON with Cu(OAc)2·H2O in MeOH and it was isolated as a black solid, while cis-[Cu(I-NON–H)(I-NON)](I3) (1) was generated by the treatment of [Cu(NON–H)2] with 8-fold excess of I2 in a CHCl3/MeOH solution at RT. For details of characterization of 1 and 2, see ESI.I-NON, as the structural analog of NON, can be also applied as a ligand for metal binding. We studied the ligand properties of I-NON and thus synthesized two I-NON-based Cu2 from the MeOH/THF solution resulted in the release of trans-[Cu(I-NON–H)2(THF)2] (3), while the crystallization of trans-[Cu(I-NON–H)2] from MeOH/Py led to the precipitation of crystals of trans-[Cu(Py)2(I-NON–H)2] (4). Complex 1 was crystallized directly from the reaction mixture. Structures of these three complexes were studied by XRD ,cis- geometry, while 3 and 4 exist in the trans-,trans- form. In 1, the copper(II) center forms two Cu–N, two Cu–O, and one Cu–I coordination bonds, being in the square-pyramidal environment.The crystal structure of 3, copper(II) is in a distorted octahedral environment, where the equatorial Cu–N (1.964(5)–1.977(5) Å) and Cu–O (1.953(4)–1.957(4) Å) bonds are distinctly shorter than the axial Cu–OTHF bonds (2.448(4)–2.535(5) Å). In 4, the coordination octahedron is less distorted with quite similar distances of the Cu–N (2.018(4) Å) and Cu–NPy (2.028(4) Å) bonds and with a slightly longer Cu–O length (2.302(3) Å). The values of the distances N–O (1.240(6)–1.299(7) Å) and N–C (1.313(8)–1.354(6) Å) of the I-NON–H ligands in 3 and 4 and one I-NON–H ligand in 1 support the existence of these ligands predominantly in the nitrosophenolate form. In contrast, the ligated I-NON in 1 demonstrates the interatomic separations (N–O 1.330(7), N–C 1.313(8), and C–O 1.249(8) Å), which are more typical for the quinone monoxime form.The Cu–I distance (2.9417(8) Å) is slightly longer than those typical for the conventional Cu–I bonds (2.68 Å)  and exhi1 and 3–4  is greater than the corresponding sum of Bondi van der Waals radii (I3) (M1), the maximum values of ESP at the terminal ends of the C–I bonds calculated on the 1.1× van der Waals surface are 0.054 and 0.061 a.u.  at the HaB donor atom. Indeed, in the optimized structure of the monomer 061 a.u. A. Despit I2 atom B.The charge transfer (CT) plays only an insignificant role in these HaBs. The NBO analysis indicated that the main contribution in CT in HaB(I···I)1 and HaB(I···I)2 comes from the LP(I3)→σ*(I1–C) and LP(I4)→σ*(I2–C) transitions with the second order perturbation energies E(2) of only 1.3 kcal/mol. Meanwhile, such CT confirmed the attribution of both these interactions to the type II HaBs with the I1 and I2 atoms serving as HaB donors; the back CT was not significant. The charge transfer in HaB(I···O)1–HaB(I···O)3 is even less substantial with E(2) of 0.8–0.9 kcal/mol for the predominant LP(O)→σ*(I–C) transitions and it is negligible for HaB(I···O)4.int(BSSE) and Eint(S); see Computational details). The Eint values of the individual bonds are within the range of −4.0 to −10.4 kcal/mol that is typical for HaBs of weak to medium strength, with the I···I bonds being slightly more stable than the I···O ones  salts, in particular Cu(OAc)2·H2O, were found to be the best promotors of this reaction. The metal center played multiple roles in this iodination: Copper(II) stabilized NON in a deprotonated nitrosophenolate form by the coordination, thus preventing oxidative deoximation; the anionic aromatic NON–H− form is preferable for aromatic electrophilic substitution involving such halogen as iodine. In addition, copper(II) activated I2 as an electrophilic agent + requires 321.9341), 620.8707 ([2M + Na]+ requires 620.8784). νmax(KBr)/cм−1: 3434, 3355, 3176 m ν(OH); 2922 m, 2852 w ν(C–H); 1668 s ν(C=N or C=O); 1594 w, 1541 s δ(CHAr). 1H NMR  δ 17.16 , 8.40 , 8.34 , 7.58 , 7.50 , 7.44 . 13C NMR  δ 177.93 (C2), 156.62 (C4), 143.03 (C1), 131.55 (C7), 130.97 (C10), 130.21 (C9), 129.95 (C6), 129.08 (C5), 123.42 (C8), 97.74 (C3). 13C ss-NMR δ 177.34 (C2), 157.05 (C4), 140.49 (C1), 134.24 (C7), 132.63 (C10), 131.09 , 130.11 (C5), 125.45 (C8), 104.24 (C3). On heating in a capillary (2°/min), this complex turns black and it decomposes at 154–156 °С. TGA: 158–228 °C (mass loss 22.5%), 228–576 °C (mass loss 37.6%). Crystals suitable for XRD were obtained from the reaction mixture by its slow evaporation.Yield 40.6 mg, 94%. Dark brown residue. Anal. Calcd for C3) (1).[Cu(I-NON–H)(I-NON)](I Cu(C10H6NO2)2 (0.025 mmol) was added to a solution of I2 (0.2 mmol) in CHCl3/MeOH  placed in a 20-mL, round-bottomed flask. The reaction mixture was stirred at RT under ultrasonic treatment until the homogenization was complete and then left to stand at RT for slow evaporation. [Cu(I-NON–H)(I-NON)](I3)  was released as black crystals after 7–9 d.20H11CuI5N2O4: C, 23.07; H, 1.06; N, 2.69. Found: C, 23.21; H, 1.19, N, 2.43%. HRESI+-MS: m/z: 299.9514 ([I-NON + H]+ requires 299.9521), 321.9327 ([I-NON + Na]+ requires 321.9341), 533.9123 ([Cu(I-NON–H)(NON)]+ requires 533.9153), 620.8766 ([2I-NON + Na]+ requires 620.8784), 659.8088 ([Cu(I-NON–H)2 + H]+ requires 659.8104), 681.7916 ([Cu(I-NON–H)2 + Na]+ requires 681.7924), 1019.6751 ([Cu2(I-NON–H)3]+ requires 1019.6687). HRESI–MS, m/z: 126.9059 ([I]– requires 126.9045), 297.9354 ([I-NON − H]− requires 297.9365), 380.7113 ([I3]− requires 380.7134), 658.7954 ([Cu(I-NON–H)2]− requires 658.8026), 956.7297 ([Cu(I-NON–H)3]− requires 956.7391). νmax(KBr)/cм−1: 3443 m ν(O–H); 3018 w ν(Csp2–H); 1574 m-s and 1537 s ν and δ(CHAr). On heating in a capillary (2°/min), this complex turns black and it decomposes at 189–190 °С; this decomposition is accompanied with iodine elimination. TGA: 181–576 °C (weight loss 62.1%). Crystals suitable for XRD were obtained on slow evaporation of the reaction mixture.Yield 17 mg, 67%. Dark-brown-red crystals. Anal. Calcd for C2] (2).+ requires 533.9153), 555.8969 ([Cu(I-NON–H)(NON-H) + Na]+ requires 555.8957), 659.8121 ([M + H]+ requires 659.8104), 681.7948 ([M + Na]+ requires 681.7924), 1340.5958 ([2M + Na]+ requires 1340.5950). νmax(KBr)/cm−1: 3060 w, 2921 w ν(C–H); 1576 m-w and 1531 δ(CHAr), 1511 s δ(CHAr) and ν(N=O). On heating in a capillary (2°/min), this complex turns black and it decomposes at 220–221 °С. TGA: 233–577 °C (weight loss 48.8%). Crystals of 3 suitable for XRD were obtained from solution of 2 in CHCl3/THF on its slow evaporation. Crystals of 4 suitable for XRD were obtained from a solution of 2 and excess Py in CHCl3/MeOH on its slow evaporation.Yield 77 mg, 66%. Dark residue. Anal. Calcd for CThe calculations for the QTAIM, ESP, NBO, and IGM analyses were carried out using the crystallographic coordinates at the DFT level of theory with the PBE0 functional ,44 and t1BM1, 2BM1, 1BM3, 2,BM3 and 3BM3) with geometries corresponding to the XRD structures of 1 and 3 and triplet spin state were used in the calculations. The C–H bonds were fixed at 1.09 Å and the O–H bonds were fixed at 1.0 Å.Five bimolecular clusters (M1 was optimized at the PBE0-D3BJ/DZP(I-ADZP) level and the energies and ESP values were refined at the PBE0-D3BJ/DZP-DKH(I-ADZP-DKH) level.The structure of the monomeric complex The topological analyses of the electron density distribution with the help of the AIM method of Bader  were perint(BSSE)] was calculated as the difference of total energy of dimer and the sum of the energies of monomers with unrelaxed geometries with the corresponding basis set superposition error (BSSE) correction:n = 1 for the I···I bonds and n = 2 for the I···O bonds. BSSE was estimated using the counterpoise method  was calculated as the energy difference of two dimers:In the second approach, the interaction energy [EIn dimer1, the HaB donor I atom (I1 or I2) forming the halogen bond was substituted by the hydrogen atom and the length of this C–H bond was fixed at 1.09 Å. In dimer2, the equivalent I atom not forming the HaB was replaced by the H atom (see


Theelucidation of their crystal structures revealed the arrangement ofthree monomers , one dimer (4), and two coordination polymers (1 and 2) depending on the steric requirementsof the dPy and predominance of the ligands. Besides, the study oftheir photophysical properties in solution supported by TD-DFT calculationsenabled us to understand their electronic effects and the influenceof the structural arrangement on them.Since many factors influence thecoordination around a metal center,steric and electronic effects of the ligands mainly determine theconnectivity and, thus, the final arrangement. This is emphasizedon Hg(II) centers, which have a zero point stabilization energy and,thus, a flexible coordination environment. Therefore, the unrestrictedHg(II) geometry facilitates the predominance of the ligands duringthe structural inception. Herein, we synthesized and characterizeda series of six Hg(II) complexes with general formula (Hg(Pip) N-, N^N′-, and N^N^N-donor sites have been characterizedand their crystal structure elucidated. Steric crowding defined thepredominance of the linkers in the structural arrangement, varyingfrom monomers to coordination polymers. Their photophysical propertieswere analyzed through experimental and TD-DFT calculations, identifyingthe character and regions involved on each electronic transition.In addition, seclusion of the HOMO from the LUMO defined them as appropriateto selective band gap tuning.Six Hg(II)complexes containing 1,3-benzodioxole-5-carboxylicacid and pyridine derivatives bearing  While metal ionsact as nodes and mainly determine the dimensionality through its geometry,modulation of the organic linkers drives the assembly of different architectures and therefore functionalities.10 electronic configuration, and therefore zerocrystal field stabilization energy, stand out for their flexible coordinationenvironment and wide range of geometries,12 usually presenting severe distortions. They enable metal–ligandrearrangement during the structural inception, which gives rise tohighly ordered networks.15 Furthermore, complexes with d10 metal nodes excel at presenting photoluminescence propertiesand, hence, have potential applications as fluorescence-emitting materials.16Among all of the potential metal nodes, those belonging togroup12 with a d17 it tends to form low coordinated linear structures,18 with the formation of Hg(II) coordination polymers beingscarce compared to that of Zn(II) and Cd(II). In addition, Hg(II)complexes have potential applications in paper, paints, and cosmeticsindustry or in the production of manometers, mercury batteries, andenergy efficient fluorescent light bulbs.It is essential to stress the lack ofknowledge on the coordinationbehavior and photophysics of Hg(II) nodes. Even though Hg(II) canadopt coordination numbers from 2 to 10,22In terms of the linkers,carboxylic acids are a recognized classof versatile organic ligands for their potential coordination modes,from monodentate to bidentate bridging, which can form a large varietyof structures. Besides, the addition of pyridine derivative ligands(dPy) with different denticity, electronic donor properties, conjugation,and planarity or steric requirements could result in diverse arrangementsand functionalities.23 In particular, modulatingthe bite angle combined with a strong chelate effect using N^N-bidentate or N^N^N-tridentate chelate ligands has provento be a determining factor of the final molecular geometry with aconcomitant effect in the emission spectra.24In this scenario,the photophysical properties of these systemscould be improved by adding conjugated π aromatic linkers bytuning their band gap and also driving the crystal packing throughC–H···π and π···πinteractions.25 our groupstudiedand analyzed the structure and photophysics from the reaction of M(OAc)2 (M = Zn(II), Cd(II)) with 1,3-benzodioxole-5-carboxylic acid and two pyridine derivative ligands (dPy= 3-phenylpyridine (3-phpy) and 4-phenylpyridine (4-phpy)), whichresulted in the formation of two Zn(II) dimeric paddle-wheels ([Zn(μ-Pip)2(dPy)]2) and two Cd(II) dimers ([Cd(μ-Pip)(Pip)(dPy)2]2).In a previous paper,2(H2O)2]) and three coordination polymers(two of Cd(II): [Cd(μ-Pip)2(H2O)]n and [Cd3(μ-Pip)6(MeOH)2]n and one of Hg(II):[Hg(μ-Pip)2]n]). Allof them showed different topologies, coordination numbers, and modes,depending on the metal preferences.26Recently, we have reported the reactionbetween Zn(II), Cd(II),Hg(II), and HPip, leading to the formation of one Zn(II) monomer (n (1), [Hg(μ-Pip)2(4-phpy)]n (2), [Hg(Pip)2](3), [Hg(μ-Pip)(Pip)]2·C6H5F (4a), [Hg(Pip)2(terpy)]·EtOH(5), and [Hg(Pip)2(dpa)]·1/2H2O·1/2MeOH (6a) eOH (6a) 1, which Hg(II) complexes are toxic, and anymanipulation of the samples has to be carried out in the fume hoodand wearing gloves.2), 1,3-benzodioxole-5-carboxylic acid,3-phenylpyridine (3-phpy), 4-phenylpyridine (4-phpy), 2,2′-bipyridine, 1,10-phenanthroline , 2,2′:6′,2′-terpyridine(terpy), and di(2-picolyl)amine (dpa) ligands and methanol (MeOH),ethanol (EtOH), dichloromethane (CH2Cl2), diethylether (Et2O), and fluorobenzene  solvents were purchased from Sigma-Aldrich. Deuterated chloroform(CDCl3) and deuterated dimethyl sulfoxide (dmso-d6) were used for the NMR experiments and werepurchased from Eurisotop. All of them were used without further purification.Reactions and manipulation were carried out in air at room temperature(RT) for compounds 1, 2, and 6 and under reflux conditions for 3–5. Thermal decomposition temperature (d.T.) was measured on a StuartMelting Point Apparatus SMP30  with a heating rampof 2.0 °C/min in a temperature range from 20 to 210 °C.Elemental analyses  were carried out on a Euro Vector3100 instrument. HR-ESI+-MS measurements of complexes 1 and 2 in MeOH solution and 3–6 in MeOH/DMSO (80/20) were recorded in a MicroTOF-Q  instrument equipped with an electrosprayionization source (ESI) in positive mode. Na+ ions comefrom the MeOH solvent which can contain <50 ppb. The conditionswere those used in routine experiments. The nebulizer pressure was1.5 bar, the desolvation temperature was 180 °C, the dry gaswas at 6 L·min–1, the capillary counter-electrodevoltage was 5 kV, and the quadrupole ion energy was 5.0 eV. SimultaneousTG/DTA determination of compound 5 was carried out ina Netzsch STA 409 instrument, with an aluminum oxide powder (Al2O3) crucible and heating at 5 °C·min–1 from 25 to 350 °C, under a nitrogen atmospherewith a flow rate of 80 mL·min–1. Al2O3 (PerkinElmer 0419-0197) was used as a standard. TheFTIR-ATR spectra were recorded on a PerkinElmer spectrometer, equippedwith a universal attenuated total reflectance (ATR) accessory witha diamond window in the range 4000–500 cm–1. 1H, 13C{1H}, and DEPT-135 NMRspectra were recorded on an NMR-FT Bruker 360 MHz spectrometer inCDCl3 or dmso-d6 solution atRT. All chemical shifts (δ) are given in ppm. The electronicspectra in solution of MeOH (1–6)were run on an Agilent HP 8453 UV–vis spectrophotometer witha quartz cell having a path length of 1 cm in the range 200–600nm. Molar absorptivity values have been calculated as log(ε).Fluorescence measurements were carried out at 25 °C with a PerkinElmerLS 55 50 Hz fluorescence spectrometer using a 1 cm quartz cell, inMeOH solution. The samples were excited at their absorption maxima,and the emission was recorded between 200 and 440 nm. Dilution effectson data were corrected by Origin Pro 8 software.Hg(II)acetate (Hg(OAc)2  in MeOH (30 mL), a solutionof HPip  and 3-phpy  in MeOH(30 mL) was added dropwise under vigorous stirring at RT. Immediately,a white solid appeared. The reaction remained under stirring for 1h. The solid obtained was filtered and washed with 10 mL of cold diethylether. Suitable crystals were obtained by recrystallization in MeOHfor 15 days. Yield: 127 mg (59%). d.T. = 172 °C. Anal. Calcdfor C27H19HgNO8 (686.04 g mol–1): C, 47.27; H, 2.79; N, 2.04. Found: C, 46.99; H,2.84; N, 1.97%. HR-MS : m/z (%) = 156.0813 (100%) ; 554.9998 (100%) 2] +Na}+ = 554.9977); 1085.0047 (77%) 2]2 + Na}+ = 1085.0046). FTIR-ATR : 3090(w) [ν(CH)]ar, 3035(w)[ν(CH)]ar, 2918(w) [ν(CH)]al, 1628(w),1604(w) [νas(COO)], 1555(m) [νas(COO)], 1521(m), 1504(w), 1487(s) ,1475(m), 1461(m), 1437(s) [νs(COO)], 1416(m), 1372(s), 1343(s), 1258(s), 1238(s),1205(m), 1189(m), 1167(m), 1131(m), 1112(s) [ν(C—O—C)],1072(m), 1032(s) [δip(C—H)], 934 (m), 921(m),882(s), 839(w), 817(w), 804(m) [δoop(C—H)],772(s) [δoop(C—H)], 753(s) [δoop(C—H)], 721(m), 697(s), 680(s), 656(m), 625(m). 1H NMR : δ = 6.06 , 6.89 , 7.52 , 7.76 , 8.01 , 8.64 , 8.89 . 13C{1H} NMR : δ = 169.20 [O2C—C], 150.57 [O2C—C—(CH)2—C], 148.89 [N—CH—C], 148.05 [N—CH—CH], 147.49 [O2C—C—CH—C], 137.05 [N—CH—C—C], 136.42 [N—CH—C], 135.52 [N—(CH)2—CH], 129.59 [C—C—CH—CH—CH or C—C—(CH)2—CH]3-phpy, 128.74 [C—C—(CH)2—CH or C—C—CH—CH—CH]3-phpy, 127.58 [O2C—C], 127.26 [C—C—CH]3-phpy, 125.54 [O2C—C—CH—CH],124.80 [N—CH—CH], 109.91[O2C—C—CH—C],108.18 [O2C—C—CH—CH], 102.02 [O—CH2—O]. UV–vis (MeOH) λmax (ε)= 212 nm (4.52); 258 nm (4.43); 278 nm (4.13); 293 nm (4.19).To a solutionof Hg(OAc)2  in MeOH (30 mL), a solutionof HPip  and 4-phpy  in MeOH(30 mL) was added dropwise under vigorous stirring at RT. Immediately,a white solid appeared. The reaction remained under stirring for 1h. The solid obtained was filtered and washed with 10 mL of cold diethylether. Suitable crystals were obtained by recrystallization in MeOHfor 18 days. Yield: 109 mg (53%). d.T. = 171 °C. Anal. Calcdfor C27H19HgNO8 (686.04 g mol–1): C, 47.27; H, 2.79; N, 2.04%. Found: C, 46.99; H,2.64; N, 1.98%. HR-MS : m/z (%) = 156.0822 (100%) ; 554.9990 (100%) 2] +Na}+ = 554.9977); 1085.0045 (71%) 2]2 + Na}+ = 1085.0046). FTIR-ATR : 3073(w) [ν(CH)]ar, 2907(w)[ν(CH)]al, 1614(w) [νas(COO)], 1560(m),1550(m) [νas(COO)], 1502(m), 1485(m) , 1433(s) [νs(COO)], 1373(s), 1336(s), 1256(s), 1227(s),1168(m), 1109(m) [ν(C—O—C)], 1073(m), 1031(s)[δip(C—H)], 1012(w), 934 (w), 921(m), 878(m),845(m), 821(m), 804(m) [δoop(C—H)], 767(s)[δoop(C—H)], 761(s) [δoop(C—H)], 731(m), 719(m), 696(m), 677(s), 665(m), 621(m). 1H NMR : δ = 6.02 ,6.83 , 7.51 , 7.64 , 7.68 , 7.72 , 8.79. 13C{1H} NMR : δ = 168.97 [O2C—C], 150.26 [N—CH], 150.17 [O2C—C—CH—CH—C], 148.67 [N—CH—CH—C], 147.14 [O2C—C—CH—C], 136.49 [N—(CH)2—C—C], 129.83 [C—C—(CH)2—CH or C—C—CH—CH—CH], 129.34 [C—C—(CH)2—CH or C—C—CH—CH—CH], 127.51 [O2C—C], 127.08 [C—C—CH], 125.16 [O2C—C—CH—CH], 122.12 [N—CH—CH], 109.59 [O2C—C—CH—C], 107.82 [O2C—C—CH—CH], 101.66 [O—CH2—O]. UV–vis (MeOH) λmax (ε) = 205 nm (4.78); 259 nm (4.50); 293 nm (4.22).To a solution ofHg(OAc)2  and HPip  in EtOH (25 mL),a solution of 2,2′-bipy  in MeOH (10 mL)was added dropwise under vigorous stirring and kept under reflux conditionsfor 4 h. A white powder precipitated. The solid obtained was filteredand washed with 5 mL of cold MeOH and 5 mL of cold diethyl ether.Suitable crystals were grown by recrystallization in MeOH for 5 days.Yield: 160 mg (75%). d.T. = 184 °C. Anal. Calcd for C26H18HgN2O8 (687.02 g·mol–1): C, 45.45; H, 2.64; N, 4.08. Found: C, 45.30; H,2.58; N, 3.86%. HR-MS : m/z (%) = 523.0570 (100%) ]}+ = 523.0579). FTIR-ATR : 3107–3019 [ν(CH)]ar, 2999–2921(w)[ν(CH)]al, 1629(w), 1598(w), 1592(w), 1578(m), 1544(m)[νas(COO)], 1503(m), 1490(m) , 1470(m), 1432(m) [νs(COO)],1375(s) , 1337(m), 1314(m),1256(s), 1239(s), 1205(m), 1158(m), 1099(m) [ν(C—O—C)],1076(w), 1032(s) [δip(C—H)], 1019(m), 981(w),919 (m), 883(m), 819(m), 805(m) [δoop(C—H)],785(m) [δoop(C—H)], 769(s) [δoop(C—H)], 734(m) [δoop(C—H)], 720(m)[δoop(C—H)], 679(m), 651(m), 627(m), 586(m),540(m). 1H NMR : δ = 6.07 , 6.93 , 7.37 , 7.54, 7.76 , 8.20 , 8.63 , 8.87 . 13C{1H} NMR : δ = 169.31 [O2C—C], 150.84 [N—C—C], 150.06 [O2C—C—(CH)2—C], 149.73 [N—CH—CH], 147.11 [O2C—C—CH—C], 139.43 [N—(CH)2—CH], 128.20 [N—CH—CH], 125.87 [O2C—C], 124.91 [O2C—C—CH—CH], 122.11 [N—C—CH], 109.43 [O2C—C—CH—C], 107.77 [O2C—C—CH—CH], 101.63 [O—CH2—O]. UV–vis (MeOH) λmax (ε) = 203 nm (4.69); 215 nm (4.66); 244 nm (4.18);260 nm (4.25); 287 nm (4.29).To a colorless solution of Hg(OAc)2  and HPip  in EtOH(25 mL), a solution of 1,10-phen  in MeOH (5mL) was added dropwise under vigorous stirring and kept under refluxconditions for 8 h. A white powder precipitated. The solid obtainedwas filtered and washed with 5 mL of cold MeOH. Suitable crystalswere grown by recrystallization in fluorobenzene for 6 days, obtainingthe compound [Hg(μ-Pip)(Pip)]2·C6H5F (4a). The characterization correspondsto 4. Yield: 172 mg (76%). d.T. = 190 °C. Anal.Calcd for C56H36Hg2N4O16 (1422.09 g·mol–1): C, 47.28; H, 2.55;N, 3.94. Found: C, 47.22; H, 2.43; N, 3.82%. HR-MS : m/z (%) = 547.0576(100%) ]}+ = 523.0580).FTIR-ATR : 3064–3014 [ν(CH)]ar, 2903–2782(w) [ν(CH)]al, 1622(w)[νas(COO)], 1603(w), 1585(m), 1574(m), 1543(m) [νas(COO)], 1501(m), 1485(m) ,1436(s) [νs(COO)], 1367(s) , 1319(s), 1260(s), 1240(s), 1163(m), 1142(w),1107(m) [ν(C—O—C)], 1075(w), 1038(s) [δip(C—H)], 937(m), 917(m), 892(w), 879(w), 863(w), 853(m),821(w), 802(m) [δoop(C—H)], 768(s) [δoop(C—H)], 725(s) [δoop(C—H)],680(m) [δoop(C—H)], 640(w), 580(m). 1H NMR : δ = 5.97 , 6.76, 7.57 , 7.71 , 8.0 , 8.01 , 8.53 , 9.42 . 13C{1H} NMR : δ = 169.16[O2C—C], 150.37 [N—CH], 149.49 [O2C—C—CH—CH—C], 146.74 [O2C—C—CH—C], 139.51 [N—(CH)2—CH], 138.47 [N—C], 128.89 [N—C—C], 128.66 [O2C—C],127.07 [N—CH—CH], 125.47[N—C—C—CH], 124.60[O2C—C—CH—C],109.32 [O2C—C—CH—CH], 107.39 [O2C—C—CH—CH], 101.28 [O—CH2—O]. UV–vis (MeOH) λmax (ε) = 206 nm (4.78); 216 nm (5.03); 266 nm (5.05);294 nm (4.93); 326 nm (3.68).To a colorless solution of Hg(OAc)2  and HPip  in EtOH (25 mL),a colorless solution of terpy  in EtOH (15 mL)was added dropwise under vigorous stirring, and the solution turnedyellow. The reaction was kept under reflux conditions for 3 h. Thesolution was concentrated until half of the volume and cooled downin an ice bath until a yellowish powder precipitated. The solid wasfiltered and washed with 5 mL of cold methanol. Suitable crystalswere obtained by slow evaporation of mother liquors on air for 12days. Yield: 214 mg (89%). d.T. = 197 °C. Anal. Calcd for C33H26HgN3O9 (810.17 g·mol–1): C, 48.92; H, 3.36; N, 5.19. Found: C, 48.74; H,3.23; N, 5.04%. HR-MS : m/z (%) = 600.0837 (100%) (terpy)]}+ = 600.0846). FTIR-ATR : 3415(br) [ν(OH)], 3099–3016(w) [ν(CH)]ar, 2964–2789(w) [ν(CH)]al, 1624(w),1591(w), 1575(w), 1534(m) [νas(COO)], 1506(w), 1478(m), 1432(s) [νs(COO)], 1366(s) , 1347(s),1312(s), 1253(s), 1196(w), 1160(m), 1114(w) [ν(C—O—C)],1071(m), 1053(m), 1038(s) [δip(C—H)], 1010(m),974 (w), 936(m), 916(m), 882(m), 852(w), 820(m), 805(m) [δoop(C—H)], 776(s) [δoop(C—H)],740(w), 721(w), 680(m), 650(m), 636(w), 608(w), 586(m), 516(w), 507(w). 1H NMR : δ = 3.72 , 1.62 , 1.24 , 5.94,6.72 , 7.52 , 7.65 , 7.66, 8.0 , 8.23 , 9.23. 13C{1H} NMR : δ = 169.38 [O2C—C], 150.65 [N—CH], 149.85 [O2C—C—(CH)2—C], 147.27 [O2C—C—CH—C], 140.30[N—C—CH—CHside and N—C—CH—CHcenter], 129.93 [N—CH—CH and N—C—CHcenter], 126.90 [O2C—C], 125.09 [O2C—C—CH—CH], 123.78 [N—Cside and N—Ccenter], 123.22 [N—C—CHside], 109.95 [O2C—C—CH—C], 107.89 [O2C—C—CH—CH], 101.80 [O—CH2—O], 56.49 [HO—CH2], 19.02 [HO—CH2—CH3]. UV–vis (MeOH) λmax (ε) = 205 nm (4.85); 253 nm (4.28); 282 nm (4.30);298 nm (4.15); 322 nm (4.17).To a colorless solution of Hg(OAc)2 in MeOH (10 mL), a solution of HPip  in MeOH (20mL) was added under stirring. Then, a yellow solution of dpa  in MeOH (5 mL) was added and stirred for 4 h. Theresulting yellow solution was concentrated under vacuum until a darkyellow oil-like reaction crude was formed, which was dissolved in10 mL of CH2Cl2, forced to precipitate with15 mL of cold diethyl ether, and filtered (repeated twice). The finalbrownish powder was washed twice with 5 mL of cold diethyl ether.Recrystallization of the solid in MeOH and cooling down to 4 °Cfor 5 days resulted in suitable crystals of [Hg(Pip)2(dpa)]·1/2H2O·1/2MeOH (6a). The characterization corresponds to 6. Yield: 118 mg (54%). d.T. = 178 °C. Anal. Calcd forC28H23HgN3O8 (730.09 g·mol–1): C, 46.06; H, 3.18; N, 5.76. Found: C, 46.14; H,3.22; N, 5.80%. HR-MS : m/z (%) = 566.0994 (100%) (dpa)]}+ = 566.1001). FTIR-ATR : 3312(w) [ν(NH)], 3069 (w) [ν(CH)]ar, 2902(m) [ν(CH)]al, 1622(w), 1600(w), 1571(m) [νas(COO)], 1545(m), 1502(w), 1485(m) ,1433(s) [νs(COO)], 1365–1321(s) , 1285(m), 1252(s), 1237(s), 1159(m), 1126(w),1104(m) [ν(C—O—C)], 1072(w), 1036(s) [δip(C—H)], 1012(m), 986(w), 935 (m), 920(m), 885(w),880(w), 804(m) [δoop(C—H)], 769(s) [δoop(C—H)], 719(m) [δoop(C—H)],678(m), 638(w), 583(m), 537(w), 524(w), 505(w). 1H NMR: δ = 4.16, 5.10, 6.03 , 6.87 , 7.37 , 7.51 , 7.55 , 7.97 , 8.75 . 13C{1H} NMR : δ = 169.47 [O2C—C], 155.34 [N—C—CH2], 149.61 [O2C—C—(CH)2—C], 149.42 [N—CH], 147.25[O2C—C—CH—C], 139.50 [N—(CH)2—CH], 130.98 [O2C—C], 124.96 [O2C—C—CH—CH], 124.58 [N—C—CH + N—CH—CH], 110.04 [O2C—C—CH—C], 107.85 [O2C—C—CH—CH], 101.74 [O—CH2—O], 50.77 [N—CH2]. UV–vis (MeOH) λmax (ε) = 202 nm (4.17); 238 nm (4.03); 281 nm (3.88); 307 nm(3.99).To a solution of Hg(OAc)1 and 2), colorless prism-like, and yellowneedle-like (5) specimens were used for the X-ray crystallographicanalysis.The X-ray intensity data were measured on a D8 Venture system equippedwith a multilayer monochromator and a Mo microfocus (λ = 0.71073Å). For 1–6a, the frames wereintegrated with the Bruker SAINT Software package using a narrow-framealgorithm. For 1, the integration of the data using atriclinic unit cell yielded a total of 17,255 reflections to a maximumθ angle of 23.27° (0.90 Å resolution), of which 3230were independent  and 3127 (96.81%) were greater than 2σ(|F|2). The calculated minimum and maximum transmission coefficients are 0.5894 and 0.7449. For 2,the integration of the data using a triclinic unit cell yielded atotal of 17,834 reflections to a maximum θ angle of 27.12°(0.78 Å resolution), of which 2591 were independent  and 2437 (94.06%)were greater than 2σ(|F|2). Thecalculated minimum and maximum transmission coefficients  are 0.5114 and 0.7461.Colorlessneedle-like , of which 3445 were independent  and 3237 (93.96%) were greaterthan 2σ(|F|2). The calculated minimumand maximum transmission coefficients  are0.3609 and 0.7461. For 4a, the integration of the datausing a monoclinic unit cell yielded a total of 58,489 reflectionsto a maximum θ angle of 30.46° (0.70 Å resolution),of which 7936 were independent  and 7288 (96.83%) were greater than 2σ(|F|2). The calculated minimum and maximum transmissioncoefficients  are 0.5115 and 0.7461. For 5, the integration of the data using a monoclinic unit cellyielded a total of 64,064 reflections to a maximum θ angle of30.56° (0.70 Å resolution), of which 4497 were independentand 4274 (95.04%) were greater than 2σ(|F|2). The calculated minimum and maximum transmission coefficients are 0.6261 and 0.7461. For 6a, the integration of the data using a triclinic unit cell yieldeda total of 159,036 reflections to a maximum θ angle of 36.35°(0.60 Å resolution), of which 25,925 were independent  and 23005 (88.74%)were greater than 2σ(|F|2). Thecalculated minimum and maximum transmission coefficients  are 0.5108 and 0.7471.For 29 For 1, the final anisotropic full-matrix least-squaresrefinement on |F|2 with 334 variablesconverged at R1 = 1.18% for the observeddata and wR2 = 4.68% for all data. For 2, the final anisotropic full-matrix least-squares refinementon |F|2 with 128 variables converged at R1 = 4.59% for the observed data and wR2 = 13.46% for all data. For 3, the final anisotropic full-matrix least-squares refinement on |F|2 with 168 variables converged at R1 = 1.96% for the observed data and wR2 = 4.93% for all data. For 4a, the finalanisotropic full-matrix least-squares refinement on |F|2 with 334 variables converged at R1 = 2.68% for the observed data and wR2 = 6.10% for all data. For 5, the final anisotropicfull-matrix least-squares refinement on |F|2 with 224 variables converged at R1 =1.39% for the observed data and wR2 =3.27% for all data. For 6a, the final anisotropic full-matrixleast-squares refinement on |F|2 with755 variables converged at R1 = 1.87%for the observed data and wR2 = 4.33%for all data.The structures were solvedusing the Bruker SHELXTL software packageand refined using SHELX (version-2018/3).1–6a, thefinal cell constantsand volume are based upon the refinement of the XYZ-centroids of reflections above 20 σ(I). Datawere corrected for absorption effects using the multiscan method (SADABS).Crystal data and relevant details of structure refinement for compounds 1–6a are reported in 1), 2043892 (2), 2043890 (3), 2043893 (4a), 2043894 (5), and 2043895 (6a). Molecular graphics were generatedwith Mercury 4.2.0 software32 using the POV-Ray image package.33 The color codes for all of the molecular graphicsare as follows: light gray (Hg), light blue (N), red (O), yellow (F),gray (C), and white (H).For 34 Since the chemistry of Hg(II) is still largelyunexplored, there is a lack of systematic computational research tobetter understand experimental results, especially from an electronicperspective as in the field of photochemistry. It is worth mentioningthat the reported examples of TD-DFT calculations with Hg(II) complexesare scarce and no previously reported data have been found regardingelectronic transition calculations. Geometry optimization of the groundstate (GS) and vertical absorptions from the electronically excitedstate (EES) for 1–5 have been donewith density functional theory (DFT) and time-dependent DFT (TD-DFT),respectively, using the ωB97X-D36 functional. A correlation consistent basis setwas used for the Hg(II) atom, the effective core potential CrenbL,37 while the 6-311G39 basis set was used with C, H, N, and O atoms. MeOH solvation effectswere incorporated using the polarizable continuum model-linear response(PCM-LR).41 The frequencies were also computed for eachoptimized structure to ensure that the geometries corresponded toan energy minimum. All of the optimized geometries in MeOH solutionare similar to their corresponding ones determined by the single crystalX-ray diffraction method. The most significant conformational changesare observed in 5. For instance, this is reflected inits pyridyl–pyridyl torsion angles. Unfortunately, all of theefforts to reach the geometry energy minima of complex 6 were unsuccessful. The highest occupied molecular orbital (HOMO)and lowest unoccupied molecular orbital (LUMO) were first examinedand the band gap energies calculated. For 1–5, the first 60 vertical absorptions from the ground stateto the excited states have been calculated and only the most probabletransitions  have been selectedfor the electronic analysis.All of the calculationshave been performed using Gaussian 09 software, version D.01.42 is a useful tool to identify and represent the molecular orbitalsimplied on the transition. The generation of NTOs is based on separatelyperforming unitary transformation for occupied MOs and virtual MOsoutstanding those pairs with predominant contributions, which makesthe orbitals’ inspection much easier. NTOs of the selectedtransitions for 1–5 have been generatedusing an isovalue of 0.02. Likewise, the identification of each transitiontype—(i) charge transfer (CT), in which the transition leadsto evident movement of charge density, or (ii) local excitation, wherethe electronic movement occurs in the same spatial region (LE)—isnot straightforward, and it has been afforded by calculating the Δr index43  define likely LE modes, while highervalues (>2.0) pertain to charge transfers) and mapping the transitiondensity matrix (TDM) as a color filled 2D plot. The combination ofboth allows the electronic excitation mode to be established. Thediagonal terms of the 2D plot highlight the primary atoms involvedin the transition, and the spatial extent can be graphically evaluated. The Δr index and TDM mapping have been calculatedusing Multiwfn 3.7 software.44Electronic transitions are composedof multiple non-negligible contributions from molecular orbitals (MOs),and their representation and evaluation as canonical orbitals couldbe intricated. Therefore, the useof natural transition orbitals (NTOs)1–6 were prepared via combination of Hg(OAc)2, HPip, and the corresponding dPy. Reactions were performedin a 1:2:4 (Hg:Pip:dPy) molar ratio for compounds 1 and 2, while 3–6 were synthesizedin a 1:2:1 proportion. The different molar ratio between the synthesesof 1 and 2 and complexes 3–6 could lie in the different coordination modes of the ligands(monodentate in 1 and 2 and chelate in 3–6). The synthesis of 1, 2, and 6 was done in MeOH at room temperature(RT), while the synthesis of 3–5 inEtOH was done under reflux conditions. Therefore, the synthesis ofthe complexes depends mainly on the molar ratio and on the reactionconditions. The corresponding crystals suitable for X-ray crystallographicanalysis were grown via recrystallization in MeOH for 1–3 and 6, in fluorobenzene for 4, or by slow evaporation of mother liquors for 5. Complexes 4 and 6 crystallized occludingsolvent molecules, yielding 4a and 6a. Asaforementioned, Hg(II) ions having a d10 electronic configurationentail a zero crystal field stabilization energy and enable an easierligand rearrangement. Thus, the incorporation of the dPy ligands hasled to different arrangements, coordination numbers, and geometriesand the concomitant variation of their photophysical properties.Complexes 1H and 13C{1H}, and DEPT-135 NMR spectroscopiesand single crystal X-ray diffraction method. Compounds 1 and 2 were also characterized by HR-ESI+-MS in MeOH solution. In addition, the thermal stability of complex 5 was studied via TG/DTA determinations. Finally, the UV–visand photoluminescence spectra of 1–6 were recorded in MeOH solution and their quantum yields calculated.The photophysical properties of 1–5 were analyzed using TD-DFT calculations via 2D color mapping ofTDM supported by NTO analysis.All of the six compounds were characterized by EA, FTIR-ATR, +-MS of complexes 1–6 have been recorded. For complexes 1 and 2, the weaker monodentate Hg–N coordinationbond of 3-phpy (1) or 4-phpy (2) is brokenduring the ESI+ fragmentation, resulting in the formationof fragments with m/z 156.0813 (100%)[3-phpy + H]+ and 156.0822 (100%) [4-phpy + H]+, respectively. As a consequence, the formation of {[Hg(Pip)2] + Na}+ species is observed at m/z 554.9998  and m/z 554.9990 . Besides, {[Hg2(Pip)4] + Na}+ species are also identifiedat m/z 1087.0053  and m/z 1087.0057  . On the other hand, for complexes 3–6, the bidentate and tridentate dPy ligands are kept during ESI+ and a Pip linker is removed displaying values of m/z 523.0570 , m/z 547.0576 , m/z 600.0837 , and m/z 566.0994  correspondingto {[Hg(Pip)(dPy)]}+ fragments .EA of all of the compounds agreewith the proposal formula. Besides,ESI1–6, showingdecomposition between 171and 197 °C . The measurement was performed using 46.0 mg ofsample. The complex starts to lose the occluded ethanol molecule at90 °C  until 140 °C,followed up by decomposition at about 200 °C by the loss of theterpy ligand  and one Pip unit.The thermal decomposition temperaturewas recorded for complexes  see the . In addi–1 attributable to hydrogen bonded ν(O—H)HPip and at 1667 cm–1 from ν(COOH)indicates that the HPip is deprotonated in the six complexes. Thespectra of 1–6 display the characteristiccarboxylate bands in the range 1626–1530 cm–1 for νas(COO) and 1436–1415 cm–1 for νs(COO) . The difference between these bands (Δ= νas(COO) – νs(COO)) revealsthe coordination modes of the carboxylate linkers.45 Compounds 1, 2, and 4 display two different Δ values, 168 and 120, 179 and 114,and 188 and 101 cm–1, suggesting bidentate bridgedand bidentate chelate (μ2-η2:η1) and chelate (μ1-η2) coordinationmodes of the carboxylate groups. Compounds 3, 5, and 6 have a Δ value of 112, 102, and 102 cm–1, indicating a bidentate chelate (μ1-η2) coordination mode. All of these values arein line with the data from the X-ray analysis. Moreover, in 6, the peak corresponding to the ν(N—H) of thedpa is sharper compared to the free ligand, suggesting the N-amine coordination to the Hg(II) center. Further bandsassigned to ν(C=C)/(C=N), δ(C=C)/(C=N),δip(C—H), and δoop(C—H)from the aromatic rings have also been identified.46In the FTIR-ATR spectra,the absence of bands between 2630 and2518 cm1H NMR spectra of complexes 1, 2, 4, and 5 wererecorded in CDCl3 solvent, while those of 3 and 6 were recorded in dmso-d6 due to theirlower solubility . The spectra of all of the complexes show threesignals assigned to the aromatic protons of the Pip ligand between7.77 and 6.71 ppm and to the −CH2– between 6.07 and 5.94 ppm. The bands of the correspondingdPy are located between 9.43 and 7.37 ppm, with the chemical shiftsof o-Hpy being consistentwith the presence of N-coordinated ligands (8.91(1), 8.76 (2), 8.88 (3), 9.43(4), 9.23 (5), and 8.75 (6)ppm).46The 13C{1H} NMR spectra of complexes 1–6 have been recorded in dmso-d6 solution as well as DEPT-135 experiments,which were performed only for those complexes required to ensure thecorrect assignation of the carbon atoms from the aromatic rings . The spectra of all of the complexes display the band assignableto the carbon atom of the carboxylate group at 169.20 (1), 168.97 (2), 169.01 (3), 169.16 (4), 169.38 (5), and 169.47 (6) ppm.The position of the pyridyl o-C atomssorted in pairs of N-donors , and 150.26 (2)), N^N-donors (149.73 (3) and 150.37 (4)), and N^N^N-donors(150.65 (5) and 149.42 ppm (6)) enable usto better understand the electron donating ability of those ligands.47 These differences with the consequent upfieldshift of those complexes containing 4-phpy (2), 1,10-phen(4), and terpy (5) are essentially causedeither by resonance stabilization, inductive effects (−I), or both. In the case of 3-phpy (1) and4-phpy (2), the better resonance stabilization, the more−I effect , and the less steric hindranceconfer to 4-Phpy a better donor character. Regarding the N^N-donors, the additional phenyl ring of 1,10-phen(4) with respect to 2,2′-bipy (3)results in a −I effect. Finally, the pyridylsubstituents have a −I effect in terpy (5) compared to that of the −CH2– substituents in dpa (6), which is reflectedin the upfield shift.The P̅1 (1) and monoclinic C2/c (2) space groups, withboth being linear coordinationpolymers which expand through the a or c axis, respectively. All of the distances of less than the sum ofthe van der Waals radii have been considered to define the coordinationsphere49  or 4-phpy(2) ligand. The Pip ligands define the formation of theedge-sharing polymeric array acting at the same time as bidentatebridged and chelate (μ2-η2:η1) linkers (N bond lengths withinthe range of other reported coordination polymers.23They belongto the triclinic sphere49 3. The Hg linkers 1. Both h2 (4.1780(1)Å) with respect to 1 (4.4091(3) and 4.4763(3) Å).In 1, the polymeric chains are stacked throughout the b axis by weak π···π interactions···Cg(2)′4.679 Å) as well as the phenyl groups of the 4-Phpy ligands (Cg(3)···Cg(3)′4.274 Å) to be considered as an effective π···πinteraction  monomerwith a [HgO4N2] core (1-η2), two Pip, and one 2,2′-bipy Å). These Pip ligands stack by π···πinteractions (Cg(1)···Cg(1)′ 3.530 Å) anda double C–H···O association between one hydrogenatom of each dioxole group and the remaining carboxylate oxygen atom,which expands the crystal packing into 2D layers along the (1̅ 0 1)plane. Finally, a π···π interaction betweenthe 2,2′-bipy (Cg(2)···Cg(2)′ 3.886 Å)and a C–H···O interaction between the p-H of the 2,2′-bipy and the remaining carboxylate O atom (3.288(3) Å) define the 3D supramolecular net has a [HgO5N2] core (1-η2) coordinationmode, occupying four coordination positions, while the remaining Piplinker joins the two Hg(II) centers through both bidentate chelateand bridged (μ2-η2:η1) coordination modes. In addition, there is an occluded fluorobenzene(Fbz) molecule ···Cg(6), 3.603 Å) whichdrive the formation of the 3D net (The occluded Fbz molecules are π···πstacked (Cg(2)···Cg(2)′, 1.424 Å) in pairsand joined to the dimeric units by π···πwith the Pip rings (Cg(1)···Cg(2), 3.918 Å) anda C–H···F interaction 5b. Beside 3D net 6.C2/c space group. It is a Hg(II) monomerwith a [HgO4N3] core (1-η2) and a tridentate chelate terpy (μ1-η3) ligand (53 (It belongs to the monoclinic N3] core 7a and a ) ligand 7b with Hgand (53 9.b axis by two different C–H···Ointeractions (terpy···terpy and terpy···Pip)···Cg(2),3.601 Å) along the [1̅01] direction. These chains are joinedthrough the y···Pip)10. Both  the m-H 8.P̅1 space group,having two crystallographically independent monomeric units (A and B) and sharing the same [HgO4N3] core (1-η2) linkers and one mer-tridentate chelate dpa (μ1-η3) ligand.54 The twisting of thepyridyl rings  is favored by thefree rotation of the −CH2–···Cg(4),3.751 Å; A–A = Cg(1)···Cg(2),4.203 Å).Both monomers are associated by reciprocal N–H···Ointeractions between the amino group and a carboxylate O atom through O–H···O interactions  and a carboxylateO atom (B) (The occluded water molecule acts simultaneouslyas an acceptorwith a MeOH molecule and as a donor with the carboxylate ractions 10. Both ractions 11a assocatom (B) 11b.N^N′-donor  and tridentate N^N^N-donor (terpy and dpa) ligands formed five-memberedrings after chelation with the Hg(II) centers. Since Hg(II) has abig van der Waals radius (1.7–2.0 Å)55 allowing high coordination numbers (6 and 7), the stericeffects of dPy ligands guide the formation of complexes with diversestructural arrangements, varying from monomers to coordination polymers.Bidentate N–M–N) measurements were an initial tool to evaluate stericeffects of chelate ligands, but as only the donor atoms are considered,the bulkiness of the ligand tends to be underestimated. Further stericparameters, such as the inter alia tolman angle,56exact ligand cone angle,57 and solid angle,58 better define free rotation ligands. For thisreason, we have defined a straightforward angle measurement (outer atoms angle) that was done considering the planarityof the linkers and, therefore, assuming that the steric effects arepredominantly generated by two hydrogen outer atoms placed at thesides of the dPy ligands, especially remarkable for those having achelate effect and free rotation restrictions. All of the angles formedby two outer hydrogen atoms of each ligand and its metal center havebeen examined from the crystallographic data using Mercury softwareversion 4.3.1,32 and only the bigger ones are listed in Bite angle . Besides,the 1,10-phen linker being almost 5° less bulky than 2,2′-bipypromotes the formation of a dimeric structure instead of the monomericarray promoted by the ligands having bigger angles .60 additive UV–vismeasurements were performed within a concentration range from ∼1× 10–9 to 1 × 10–4 Mto ensure the nonaggregation of the ligands and complexes 1–6 at the selected concentration for the photoluminescenceexperiments and avoid the aggregation-caused emission quenching (ACQ)effect61 . Complexes start to aggregateat 7.67 × 10–6 M (1), 8.41 ×10–6 M (2), 1.89 × 10–7 M (4), 4.44 × 10–7 M (5), and 1.63 × 10–5 M (6). Within this tendency, complex 3 which presented weakerinteractions seems to avoid aggregation effects at higher concentrations.Those which exhibited stronger interactions in the solid state identifiedin the structural analysis tend to easily aggregate in solution andpromote a bathochromic shift emphasized at higher wavelength absorptionpeaks.62 Especially, complexes 5 and 6 exhibited bathochromic aggregation effects reflectedby the shift of absorption bands at 333 and 302 nm, respectively.Likewise, after aggregation of 1, a band at 293 nm emerged.Hereafter, the UV–vis measurements of complexes 1–6 have been done using a concentration around∼1.00 × 10–7 M . The UV–vis spectraof the ligands and l-tyrosine (l-tyr) have beenrecorded using the same concentration (Supporting Information: Table S1). The absorption maxima (λmax) of the complexes and the ligands have been identifiedand their molar absorptivity values (ε) calculated  for complexes 1–6 at the selected λexc is blue, as displayedin CIE1931 chromaticity diagrams.63 Theeffect on the emission spectrum after coordination of the ligandsto the Hg(II) center has been analyzed by comparing the emission ofthe ligands with the corresponding complex being sorted in groupsof three including the pair of ligands (Pip + dPy) with their correspondingcomplex (1–6), all being excitedat the emission maxima of the complexes. These spectra are depictedin Figure S25 of the Supporting Information and show the quenching or enhancementeffect after complexation of the ligands. Complex 6 is the only keeping the emission intensity.On the other hand, complexes 3 and 5 exhibitan emission enhancement effect , while complexes 1, 2, and 4 display a quenching effect .All of the measurements were performedat 298 K, in MeOH as solvent, and using the concentration extractedfrom the UV–vis data. The emission spectra of complexes  < 6 < 5 12. The s64 By comparisonwith a reference (standard), the relative quantum yield (ϕs) of the selected product can be obtained.65The fluorescence quantum yield(ϕ) is defined as the ratiobetween the number of photons emitted and the number of photons absorbedand describes how efficient the sample is in converting the excitationlight into photon emission.1–6 is calculatedusing eq I is the area under the curve of the emission spectra, and n is the refractive index of the solvent. In this study, l-tyrosine has been used as the standard (φref = 0.14)66 and ODref and Iref values have been obtained from a solutionat a concentration of 1.01 × 10–4 M in Milli-Qwater as solvent (nref = 1.3325)67 at 298 K. The values of As and Is of HPip and dPy ligandsas well as those of 1–6 have beenmeasured from solutions of 9.95 × 10–7 M (HPip)and ∼6.16 × 10–6 M (dPy and complexes 1–6) in MeOH as solvent (ns = 1.3314)68 at 298 K. Thevalues of ϕs obtained for compounds 1–6 are 0.027 (1), 0.011 (2), 0.968 (3), 0.051 (4), 0.370 (5), and 0.047 (6) (The quantum yield of .047 (6) 14.s values sorted by the number ofdPy N-donor sites, 1 has a slightlyhigher efficiency than 2 but is constant in the sameorder of magnitude. In the complexes with chelating ligands (3–6), even if chelation enhanced fluorescence(CHEF) competes with steric crowding, the bigger size of the Hg(II)center enables geometrical distortions without severe alterationsof the Hg–N bond lengths.69 In this sense, the non-negligible better ϕs of complex 5 with respect to 6 is probablycaused by the greater symmetry and rigidity of the terpy linker comparedto dpa, in which the aliphatic −(CH2)– groupsallowed severe bond length distortion of the [HgO4N3] core and, thus, facilitated a nonradiativedecay. Finally, the largest difference in ϕs is reflectedin complexes 3 and 4, with 3 having a significantly larger value. Despite that the 1,10-phenligand has more rigidity than 2,2′-bipy, it is known that ithas close-lying ππ* and nπ* singlet excited states,with nπ* excited states promoting its nonradiative decay andconsequently vanishingly low emission ϕs.70 On the other hand, the 2,2′-bipy ligandexhibited ππ* relaxation in polar solvents as MeOH.71 This is maintained after complexation with d10 metal ions, which do not have low-lying MLCT electroniclevels and, thus, keep ligand centered transitions. Only 1,10-phenfunctionalization, especially in 2,9-positions, led to significantlybetter ϕs values.70Comparing the ϕ1–5 agree reasonablywell with the experimental profiles. The shift in the theoreticalabsorption spectra with respect to the experimental spectra lies withinthe range of typical TD-DFT calculations (∼0.3 eV) and is causedby computing absorptions as vertical transitions.72 Only transitions with higher f valueshave been selected for the 2D color filled mapping of TDM and NTOanalysis. The HOMO and LUMO outline as well as energy band gaps canbe found in Figure S26 in the Supporting Information. Subsequently, the maincontributor transitions, being either Hg, Pip, dPy, or a combinationof them, have been analyzed for each absorption band to identify boththe character and the regions involved in the electronic transition.Each set of transitions has been represented as a 2D color filledmap of the TDM, and these results were confirmed by NTO analysis.All of the calculated UV–visspectra of complexes 1–5 have π symmetry with the HOMO being distributed among thePip ligand and the LUMO being uniformly localized over the dPy linkers.It has been previously demonstrated for Pt complexes bearing N,N- and N,N,N-donor ligands that this MO seclusion allowedthe HOMO and LUMO energies to be separately modified by functionalization,and thus, introduction of substituents on the dPy or Pip ligands couldselectively tune the band gap.74The HOMO and LUMO orbitals of 3 has been provided in 3 with the corresponding atom labeling scheme is displayed in Supporting Information: Figure S34). Both the LMCT and MLCT transitionsare predominantly responsible for the higher energetic transitionsbeing located around 200 nm. At lower energies, the more red-shiftedbands in the spectra arise from Pip in complexes 1, 2, and 3, while, in those containing more pielectron rich ligands , dPy are responsible forsuch transitions (4 and 5). Regarding metalcentered transitions, these results display that MLCT transitionsrise from Hg → Pip in complexes 1–4 (identified between 203 and 212 nm), while in 5 this contribution is not observed. The role of dPy on MLCT transitionsfor 1–4 is neglectable, since dPyare not involved in them in complexes 1 and 2 and only minor contributions from 2,2′-bipy and 1,10-phencan be observed in 3 and 4. What shouldbe noted is the different behavior displayed in complex 5, in which the terpy ligand facilitates the MLCT to be from Hg →terpy, causing a remarkable bathochromic shift on absorption (253nm). This difference is probably originated from the better π-acceptorcharacter of the terpy ligand with respect to the remaining dPy. The Supporting Information displays the completeelectronic analysis of the remaining complexes with additional 2Dplots  and NTO analysis .Analysis of complex N-, N^N-, and N^N^N-donor ligands, revealingthat the use of the Hg(II) metal node allowed the primacy of the geometricdemands of the linkers with coordination numbers 6 and 7 to be accommodated.Despite that the Pip linker demonstrated its ability to form polymericstructures by μ2-η2:η1 coordination modes (1 and 2), boththe increasing bulkiness of the dPy ligands and their chelate effectafforded the predominance of the dPy over the Pip ligand (3–6) and drove the formation of discrete structuresbeing two monomers and a dimer. A comparison of the steric effectsevinced that a slight increase in the bulkiness of dPy is sufficientto promote the formation of a monomer instead of a dimer, along withbulkier ligands inter alia terpy and dpa, which unavoidablyformed monomers. These complexes were associated by weak C–H···Oand π···π interactions, which is reflectedin the aggregation patterns, except for complex 3 thatavoided aggregation up to ∼1 × 10–4 Mand presented less and weaker interactions. Furthermore, aggregationdrove significant shifts on absorption spectra, especially remarkablefor 5 and 6. The experimental and theoretical photophysical study revealed that 1–5 have a localized HOMO on Pip whilethe LUMO is over the dPy emerging as appropriate candidates for bandgap modulation. The 2D and NTO analysis evidences that less energeticabsorptions are promoted by Pip in the presence of 3-phpy and 4-phpy,while, from 2,2′-bipy and following, dPy linkers are responsiblefor such absorptions. Finally, the LMCT transitions lie between 203and 212 nm originated from Hg → Pip in 1–4, while in complex 5 such a transition arisesfrom Hg → terpy, causing a bathochromic shift up to 253 nm.We synthesized and characterizeda series of Hg(II) coordination complexes bearing HPip and