parasitic diseases represent a major global health problem . in 2010 , the global disability - adjusted life years ( dalys ) of parasitic diseases are estimated to be over 102  000 , which are greater than some well - known diseases , such as rheumatic heart disease and diabetes ( murray et  al . , 2012 ) . however , these parasitic diseases are usually  neglected  , since they are prevalent mostly in low - income developing countries and poor or marginalized communities . most of their current chemotherapeutic strategies suffer from serious deficiencies such as poor efficacy , unacceptable toxicity , high costs and resistance occurrence . malaria , caused by infection with protozoan of the genus plasmodium , is the most socioeconomically devastating parasitic disease world widely , causing an estimated one million deaths annually ( who , 2013 ) . plasmodium falciparum is the most virulent human malaria parasite , which is responsible for the vast majority of the malaria - related deaths ( nosten et  al . , 2004 ) . in the absence of effective vaccines , chemotherapy is the important pillar for malaria treatment . however , the global emergence of resistance strains results in the gradual loss of effectiveness of the marketed drugs , including chloroquine , mefloquine and artemisinins ( dondorp et  al . , 2009 ) . schistosomiasis is a chronic and debilitating parasitic disease caused by blood - flukes of the genus schistosoma . it is the second major parasitic disease in the world after malaria with more than 230 million individuals infected ( gryseels et  al . , schistosomiasis japonica is especially prevalent in lake and marshland regions in asia , where it still remains significant health concern and considerable economic burden ( garjito et  al . , 2008 , zhou et  al . , 2012 ) . current chemotherapy relies on the only drug , praziquantel , which has been widely used as an effective antischistosomal for decades . praziquantel is very potent against the adult worms , but much less effective against the juvenile worms ( schistosomula ) ( fenwick and webster , 2006 ) . in addition , the adverse effect of mass treatment and long - term medication of praziquantel has revealed available evidence for the emergence of praziquantel resistance in schistosomes ( melman et  al . , 2009 , pica - mattoccia et  al . , 2009 ) . given the lack of alternative chemotherapeutics , there is a pressing need for new chemical entities for schistosomiasis treatment . it is known that antiplasmodials have been demonstrated to be able to kill schistosomas in  vitro and/or in  vivo , such as artemisinins ( utzinger et  al . , 2007 ) , mefloquine ( keiser et  al . , 2010 ) , chloroquine ( oliveira et  al . , 2004 ) and pyronaridine ( xue et  al . , 2013 ) . the detail mechanisms of these drugs exhibiting dual anti - parasitic activity are still unclear . one of the molecular mechanisms involves hemozoin formation ( de villiers and egan , 2009 ) . hemozoin is an aggregate of hematin ( oxidized heme ) produced upon hemoglobin digestion by hematophagous organisms . it is the main mechanism of heme detoxification in several blood - feeding organisms , including plasmodium ( noland et  al . , 2003 ) , schistosoma ( oliveira et  al . , 2000 ) , haemoproteus columbae ( chen et  al . , 2001 ) and rhodnius prolixus ( stiebler et  al . , 2010 ) . free heme ( ferriprotoporphyrin ix ) is toxic to the parasites , because it can peroxidate lipids , produce oxygen radicals , inhibit enzyme activities and damage cell membranes ( aft and mueller , 1983 , aft and mueller , 1984 ) . hence , how to dispose free heme is of central importance in the physiological processes of hematophagous organisms . to detoxify the free heme , a similar process is observed in schistosomes , and hemozoins are produced and filled in the gut of the worms ( homewood et  al . , 1972 ) . since hemozoin formation is essential for the survival of these parasites , inhibiting hematin aggregation represents an attractive drug target . indeed , plenty of evidence has indicated that antiplasmodial drugs with proved hemozoin formation inhibitory activity were effective for schistosomiasis , e.g. chloroquine ( oliveira et  al . , 2004 ) , mefloquine ( xiao et  al . , 2014 ) , and pyronaridine ( auparakkitanon et  al . , 2006 ) . in our previous work , a phenotypic in  vitro screening against adult s.  japonicum was performed . among the positive test results , two hits , jfd03612sc and btb12253sc ( maybridge database , fig .  1 ) , arose our interest . both compounds have a carbazole aminoalcohol scaffold , and caused 100% mortality of adult worms at 10  g / ml . besides , further assay results indicated that jfd03612sc exhibited moderate antiplasmodial activity against p.  falciparum 3d7 strain ( ic50  =  2.671  m , btb12253sc was not tested ) . carbazole occurs in a wide - range of biologically active compounds , including antivirals ( yamada et  al . , 2012 ) , antibiotics ( hurley et  al . , 2015 ) , antiplasmodials ( molette et  al . , 2013 ) . in addition , the aminoalcohol functional group was considered as a privileged structure for antischistosomal activity ( keiser et  al . , 2009 ) . thus , we believe that the two hits are good starting points for discovering novel antiparasitic agents against p.  falciparum and s.  japonicum . in this work , sixteen carbazole aminoalcohol derivatives were synthesized to ascertain the importance of the carbazole core , the amine type and the stereochemical structure . their antiplasmodial activities against pf3d7 and pfdd2 strains and antischistosomal activities against adult and juvenile s.  japonicum were determined . additionally , -hematin formation inhibitory activities of target compounds have also been evaluated . preliminary structure - activity relationships ( sars ) were discussed . stoichiometry determination and molecular docking studies were carried out , which helped to explain the mode of action of carbazole aminoalcohols . reagents and solvents were all purchased from sigma - aldrich , and generally were used without further treatment . chemical shifts were given in ppm (  , tms ) and coupling constants in hz . high resolution mass spectra ( hrms ) were recorded on a thermo q exactive orbitrap lc - ms / ms . thin layer chromatography ( tlc ) was performed on silica gel f254 plates from merck . to a solution of 9-(oxiran-2-ylmethyl)-9h - carbazole ( 2a - c , 2  mmol ) in etoh ( 20  ml ) , corresponding amines ( 6  mmol ) were added . for amines with lower reactivity ( e.g. arylamines ) , the reaction mixtures were heated to reflux for 6  h. the progress of the reactions was monitored by tlc . the mixtures were quenched with water ( 10  ml ) and extracted with etoac ( 3    10  ml ) . the organic phases were washed with water ( 3    20  ml ) and brine ( 3    20  ml ) , dried over anhydrous na2so4 , and concentrated under reduced pressure . the obtained residues were purified by recrystallization from ethanol to afford target compounds . the characterization data of compounds 68 and 1216 were reported in our previous work ( wang et  al . , 2016b ) , and the characterization data of compounds 35 , 911 , ( r)-7 and ( s)-7 ) were given as follows . white solid ( 59% , over two steps ) , mp : 135.1137.4  c . h nmr ( 400  mhz , dmso - d6 )  8.30 ( d , j  =  2.0  hz , 2h , ar - h ) , 7.66 ( d , j  =  8.8  hz , 2h , ar - h ) , 7.48 ( dd , j  =  8.8 , 2.1  hz , 2h , ar - h ) , 4.96 ( d , j  =  4.2  hz , 1h , oh ) , 4.784.26 ( m , 2h , ch2 ) , 4.043.97 ( m , 1h , ch ) , 2.562.36 ( m , 6h , 3ch2 ) , 1.731.60 ( m , 4h , 2ch2 ) . c nmr ( 100  mhz , dmso - d6 )  139.77 , 125.94 , 123.35 , 122.37 , 120.07 , 111.83 , 68.32 , 59.47 , 54.13 , 47.58 , 23.14 . white solid ( 66% , over two steps ) , mp : 140.3141.3  c . h nmr ( 400  mhz , dmso - d6 )  8.32 ( d , j  =  2.0  hz , 2h , ar - h ) , 7.67 ( d , j  =  8.8  hz , 2h , ar - h ) , 7.48 ( dd , j  =  8.8 , 2.0  hz , 2h , ar - h ) , 4.94 ( s , 1h , oh ) , 4.484.36 ( m , 2h , ch2 ) , 4.053.98 ( m , 1h , ch ) , 2.452.24 ( m , 6h , 3ch2 ) , 1.551.34 ( m , 6h , 3ch2 ) . c nmr ( 100  mhz , dmso - d6 ) 139.73 , 125.92 , 123.34 , 122.38 , 120.12 , 111.83 , 68.77 , 59.77 , 54.76 , 47.82 , 25.61 , 23.92 . white solid ( 51% , over two steps ) , mp : 136.3137.4  c . h nmr ( 400  mhz , dmso - d6 )  8.30 ( s , 2h , ar - h ) , 7.67 ( d , j  =  8.8  hz , 2h , ar - h ) , 7.48 ( dd , j  =  8.7  hz , 2.1  hz , 2h , ar - h ) , 5.00 ( d , j  =  5.0  hz , 1h , oh ) , 4.534.21 ( m , 2h , ch2 ) , 4.05 ( m , 1h , ch ) , 3.593.53 ( m , 4h , 2ch2 ) , 2.452.25 ( m , 6h , 3ch2 ) . c nmr ( 100  mhz , dmso - d6 )  139.70 , 125.94 , 123.36 , 122.38 , 120.09 , 111.83 , 66.72 , 66.13 , 62.10 , 54.01 , 47.72 . white solid ( 43% , over two steps ) , mp : 105.1107.9  c . h nmr ( 400  mhz , dmso - d6 )  8.31 ( d , j  =  2.0  hz , 2h , ar - h ) , 7.67 ( d , j  =  8.8  hz , 2h , ar - h ) , 7.47 ( dd , j  =  8.8 , 2.1  hz , 2h , ar - h ) , 5.04 ( s , 1h , oh ) , 4.484.19 ( m , 2h , ch2 ) , 3.953.93 ( m , 1h , ch ) , 2.572.45 ( m , 4h , 2ch2 ) , 1.411.25 ( m , 8h , 4ch2 ) , 0.86 ( t , j  =  6.8  hz , 3h , ch3 ) . c nmr ( 100  mhz , dmso - d6 )  139.61 , 125.92 , 123.37 , 122.34 , 120.10 , 111.68 , 68.83 , 53.13 , 49.54 , 47.33 , 31.28 , 29.62 , 26.54 , 22.11 , 13.91 . white solid ( 63% , over two steps ) , mp : 74.578.1  c . h nmr ( 400  mhz , dmso - d6 )  8.31 ( d , j  =  2.0  hz , 2h , ar - h ) , 7.67 ( d , j  =  8.8  hz , 2h , ar - h ) , 7.47 ( dd , j  =  8.8 , 2.1  hz , 2h , ar - h ) , 5.03 ( s , 1h , oh ) , 4.484.21 ( m , 2h , ch2 ) , 3.953.92 ( m , 1h , ch ) , 2.572.44 ( m , 4h , 2ch2 ) , 1.401.25 ( m , 10h , 5ch2 ) , 0.86 ( t , j  =  6.7  hz , 3h , ch3 ) . c nmr ( 100  mhz , dmso - d6 )  139.62 , 125.93 , 123.37 , 122.34 , 120.12 , 111.71 , 68.83 , 53.13 , 49.53 , 47.34 , 31.29 , 29.65 , 28.70 , 26.82 , 22.08 , 13.93 . white solid ( 70% , over two steps ) , mp : 87.891.2  c . h nmr ( 400  mhz , dmso - d6 )  8.29 ( d , j  =  2.0  hz , 2h , ar - h ) , 7.66 ( d , j  =  8.8  hz , 2h , ar - h ) , 7.45 ( dd , j  =  8.8 , 2.0  hz , 2h , ar - h ) , 4.474.23 ( m , 2h , ch2 ) , 3.973.92 ( m , 1h , ch ) , 2.572.43 ( m , 4h , 2ch2 ) , 1.381.22 ( m , 12h , 6ch2 ) , 0.84 ( t , j  =  6.8  hz , 3h , ch3 ) . c nmr ( 100  mhz , dmso - d6 )  139.61 , 125.91 , 123.38 , 122.35 , 120.10 , 111.67 , 68.82 , 53.11 , 49.53 , 47.32 , 31.29 , 29.65 , 29.01 , 28.74 , 26.87 , 22.10 , 13.92 . white solid ( 68% , over two steps ) , mp : 73.275.4  c . h nmr ( 400  mhz , dmso - d6 )  8.31 ( d , j  =  2.1  hz , 2h , ar - h ) , 7.70 ( d , j  =  8.8  hz , 2h , ar - h ) , 7.48 ( dd , j  =  8.8 , 2.1  hz , 2h , ar - h ) , 4.754.61 ( m , 1h , oh ) , 4.484.29 ( m , 2h , ch2 ) , 4.053.98 ( m , 1h , ch ) , 2.722.51 ( m , 4h , 2ch2 ) , 1.481.29 ( m , 4h , 2ch2 ) , 0.88 ( t , j  =  7.3  hz , 3h , ch3 ) . c nmr ( 100  mhz , dmso - d6 )  139.63 , 125.96 , 123.37 , 122.33 , 120.12 , 111.74 , 68.75 , 53.08 , 49.12 , 47.34 , 31.71 , 19.93 , 13.92 . white solid ( 60% , over two steps ) , mp : 121.5125.3  c . h nmr ( 400  mhz , dmso - d6 )  8.31 ( d , j  =  2.1  hz , 2h , ar - h ) , 7.68 ( d , j  =  8.8  hz , 2h , ar - h ) , 7.47 ( dd , j  =  8.8 , 2.1  hz , 2h , ar - h ) , 4.724.63 ( m , 1h , oh ) , 4.484.26 ( m , 2h , ch2 ) , 3.983.93 ( m , 1h , ch ) , 2.592.47 ( m , 4h , 2ch2 ) , 1.431.27 ( m , 4h , 2ch2 ) , 0.88 ( t , j  =  6.9  hz , 4h , ch3 ) . c nmr ( 100  mhz , dmso - d6 )  139.61 , 125.98 , 123.41 , 122.36 , 120.14 , 111.73 , 68.26 , 52.57 , 48.76 , 47.27 , 31.02 , 19.83 , 13.84 . pf3d7 ( chloroquine - sensitive ) and pfdd2 ( chloroquine - resistant ) strains were used in an in  vitro blood stage culture to evaluate the antiplasmodial efficacy of carbazole aminoalcohols . chloroquine was dissolved in water ( milli - q grade ) to prepare stock solution , and carbazole aminoalcohols and dihydroartemisinin in dmso . all the stock solutions were diluted with 1640 incomplete medium to reach the corresponding dilutions . synchronous ring - stage parasites ( 1% parasitaemia and 2% haematocrit ) were incubated in 96-well plates with serial dilutions of test compounds or controls for 72  h  at 37  c . in all cases except chloroquine , the highest final concentration of dmso was 0.2% , which was found to be nontoxic to the parasites . the antiplasmodial effect of carbazole aminoalcohols was determined by a sybr green i fluorometric assay ( xu et  al . , 2013 ) . adult schistosomes were harvested by dissection from mesenteric veins and livers of infected mice , 3438 days post - infection . through perfusion with ice cold hanks balanced salt solution ( hbss ) containing heparin , the in  vitro culture mediums containing rpmi 1640 ( with 10% calf serum ) , 100 iu / ml streptomycin , 100 iu / ml penicillin sodium , and 0.25  g / ml amphotericin b were prepared to maintain the schistosomes . four pairs of schistosomes ( four of both sexes ) were placed in each well . the plates were incubated at 37  c in 5% co2 for 2  h. then test compounds ( final drug concentrations : 10 and 5  g / ml ) were added . phenotypes of the schistosomes , including motility , viability and morphological alterations , were monitored at 24 , 48 and 72  h post - incubation . worm death was defined as no any motor activity observed in suckers and worm bodies for 2  min . s.  japonicum cercariae were mechanically transformed to schistosomula , and stored in culture medium ( rpmi 1640 medium supplemented with 5% fetal bovine serum , 100 iu / ml penicillin and 100  g / ml streptomycin ) at 37  c in 5% co2 , as described by keiser ( 2010 ) . for drug assay , schistosomula ( 50/well ) were incubated with test compounds ( final drug concentration : 10  g / ml ) in a 40-well culture plate at 37  c in a 5% co2 incubator for 72  h. dmso was used as negative control . the activity status , survival time , mortality and body morphology of the schistosomula were evaluated microscopically at 12  h , 24  h , 48  h and 72  h post - incubation . the inhibition assay of -hematin was performed using the np-40 detergent - mediated method ( sandlin et  al . , 2011 ) . dmso solution ( 10  l ) of test compounds at various concentrations was delivered to a 96-well plate , and then 20  l of np-40 ( 30.55  mm ) and 70  l of deionised h2o were added into each well . the hematin stock solution ( 25  mm ) was prepared by dissolving hematin in dmso through sonicating , then 178  l of which was suspended in a 2  m acetate buffer ( ph 4.8 ) . for each well , 100  l of the homogenous suspension was added to reach 0.5  m final buffer and 100  mm hematin . amodiaquine ( 100  m , final concentration ) was used as a positive control . the plates were covered and incubated at 37  c for 4  h. analysis was conducted by using the pyridine - ferrichrome method . a solution of 50% ( v / v ) pyridine , 20% ( v / v ) acetone , 30% ( v / v ) h2o , and 0.2  m hepes buffer ( ph 7.4 ) was prepared , and 32  l of this was delivered to each well to reach a final pyridine concentration of 5% ( v / v ) . in order to give assistance to hematin dispersion , additional acetone ( 60  l ) was also added . the uv - vis absorbance was recorded by a bio - rad model 680 xr microplate reader at 405  nm . cells were plated in a 96-well plate at 10  000  cells per well in 180  l of dulbecco 's modified eagle medium ( dmem ) or minimal essential medium ( mem ) supplemented with 10% fetal bovine serum and 1% penicillin - streptomycin . after 12  h incubation at 37  c in 5% co2 to allow the cells to adhere , 20  l of 2-fold serial dilutions of compounds were added in the well in triplicate . the final concentrations of compounds were 50 , 25 , 12.5 , 6.25 , 3.13 and 1.57  m . . then supernatants were removed , and 90  l of fresh medium and 10  l of mtt solution were added . after 4  h incubation , the supernatants were removed again and 110  l of dmso was added in each well . the plates were swirled gently for 10  min , and then read the absorbance at 490  nm . stoichiometry determination by the continuous variation method ( job 's plot ) was carried out to study the drug - hematin interaction through determining the spectral changes . the aqueous dmso ( 40% , v / v ) solution of 10  m hematin and test compounds were prepared as previously described by auparakkitanon et  al . the combined concentration of drug and hematin was kept constant ( 10  m ) . for each test drug , 11 solutions of drug and hematin combinations in different molar ratios were prepared as follows : 0:10 , 1:9 , 1:4 , 3:7 , 2:3 , 1:1 , 3:2 , 7:3 , 4:1 , 9:1 , and 10:0 . spectra between 240 and 700  nm were read on a beckman coulter du730 spectrophotometer . docking simulation was performed by using cdocker module ( discovery studio , version 2.1 , accelrys ) . the three - dimensional structure of heme was obtained from the free heme crystal structure ( pdb : 3p5q ) . the fe atom was charged  + 3 , and the fe ( iii ) protoporphyrin ix form was used for docking simulation . hemozoin formation occurs within the plasmodium digestive food vacuoles ( ph 4.8 ) . at relevant acidic conditions , both n atoms of compound 7 are protonated ( wang et  al . , 2016a , wang et  al . , 2016b ) . accordingly , they were charged  + 1 for docking simulations . for each isomer of 7 , random conformations were generated by utilizing charmm field molecular dynamics ( 1000 steps ) , and then docked into the defined binding site with a radius set as 11   . the binding conformations of ( r)-7 and ( s)-7 with heme were determined and ranked according to the calculated cdocking energy . among the top 30 docking poses , reaction of carbazoles ( 1a - c ) and epichlorohydrin ( racemic or enantiomerically pure ) in the presence of koh afforded the epoxypropane intermediates ( 2a - c ) , which subsequently reacted with appropriate amines to obtain corresponding target compounds 316 . the in  vitro antiplasmodial activities of carbazole aminoalcohols were determined against chloroquine - sensitive pf3d7 and chloroquine - resistant pfdd2 strains . chloroquine and dihydroartemisinin were used as the positive controls , and the results are summarized in table  1 . in general , most of the carbazole aminoalcohols exhibited moderate to potent antiplasmodial activities against pf3d7 and pfdd2 strains . retaining the dichloronated carbazole core , manipulating all the compounds with alkylamine tails ( 611 ) displayed remarkable antiplasmodial activity with ic50s in the submicromolar range against pf3d7 strain and nanomolar range against pfdd2 strain . among them , n - hexylamino ( 9 , pf3d7 ic50  =  0.274  m , pfdd2 ic50  =  0.047  m ) and n - octylamino groups ( 11 , pf3d7 ic50  =  0.132  m , pfdd2 ic50  =  0.054  m ) were the preferred substituents . for the compounds with aniline substituents ( 1214 ) , a drop of potency was observed . on the other hand , dihalogenated carbazole was found to be a privileged core structure after varying the substituents of carbazole . comparing with compound 7 ( pf3d7 ic50  =  0.248  m , pfdd2 ic50  =  0.091  m ) , eliminating the chlorine atoms of carbazole core ( 15 , pf3d7 ic50  =  0.760  m , pfdd2 ic50  =  0.334  m ) resulted in 3-fold decrease in potency in both strains . in addition , in order to find out whether the stereochemistry of the secondary hydroxyl group exerted influence on potency , the r- and s- enantiomers of 7 were prepared and evaluated . both enantiomers showed similar antiplasmodial activity ( ( r)-7 , 
 pf3d7 ic50  =  0.344  m , pfdd2 ic50  =  0.172  m ; ( s)-7 , pf3d7 ic50  =  0.378  m , pfdd2 ic50  =  0.112  m ) . all prepared target compounds were further evaluated for their antischistosomal activity against adult and juvenile s.  japonicum . sixteen compounds were tested , and twelve of them killed all adult s.  japonicum at 10  g / ml after 72  h exposure . among them , seven compounds killed worms with 50100% mortality at 5  g / ml . in consistent with the tendency observed in antiplasmodial activity , most of the compounds with alkylamine tails ( 611 ) showed significant antischistosomal activity , and the most potent compound 7 , with an n - butylamino group , killed adult s.  japonicum with 100% mortality at 5  g / ml . both enantiomers of 7 , ( r)-7 and ( s)-7 , were as effective as the racemate . in accordance with the results of p. falciparum whole cell assay , compounds with arylamine substituents ( 1214 ) suffered a significant loss of potency . in addition , compared with the dichlorinated carbazole derivative 7 , the dibromo- ( 15 ) and non - substituted ( 16 ) derivatives exhibited reduced activity against adult worms . based on the adult worm killing ability , representative compounds were further evaluated for their antischistosomal activity against schistosomula ( table  1 ) . all the tested compounds ( 6 , 7 , 911 , ( r)-7 and ( s)-7 ) demonstrated significant juvenile worm killing ability , causing 100% mortality at 5  g / ml in 24  h. the who recommended activity criterion of hit compounds for schistosomiasis is 100% inhibition of motility of schistosoma adults at 5  g / ml ( nwaka and hudson , 2006 ) . based on this criterion , compound 7 can be considered for further in  vivo animal studies . according to the above results , most of the carbazole aminoalcohols showed not only antiplasmodial but also antischistosomal activities , especially those with alkylamine substituents . plasmodium and schistosome are both hematophagous organisms , and the hemozoin formation is crucial for the survival of these parasites . we speculated that inhibiting hemozoin formation was one of the mechanisms of action of these compounds , similar to the known -hematin formation inhibitors with dual antiparasitic activities ( e.g. chloroquine , mefloquine and pyronaridine ) . in order to verify our hypothesis , the known inhibitors , chloroquine and amodiaquine , were used as positive controls . as shown in table  1 , in the preliminary assay compounds with alicyclic amine groups ( 3 and 5 ) or arylamine groups ( 1214 ) were not able to inhibit -hematin formation . the removal of two halogens on the carbazole moiety ( 15 ) resulted in a dramatic decrease in potency , which demonstrated the essential role of dihalogen - substituted carbazole core . further assay results indicated that almost all the compounds possessing alkylamine groups ( 611 ) exhibited significant -hematin formation inhibitory activity . especially , compounds with n - butylamino ( 7 , ic50  =  91.839  m ) , n - hexylamino ( 9 , ic50  =  54.983  m ) , n - heptylamino ( 10 , ic50  =  91.646  m ) , and n - octylamino ( 11 , ic50  =  65.377  m ) groups displayed potent activity equivalent to that of chloroquine ( ic50  =  61.01  m ) . the antiplasmodial activity of carbazole aminoalcohols have been identified in several phenotypic screenings , but their mode of action remains unclear . recently , it has been reported that plasmodium falciparum hsp90 was a plausible target of carbazole aminoalcohols ( wang et  al . , 2016b ) , while there is still no clue of their mechanism of action on schistosomes . in this work , three kinds of biological assay revealed a similar sar pattern : ( 1 ) dichlorinated carbazole acted as a privileged core ; ( 2 ) an alkylamine tail was beneficial for activity ; conversely , arylamine substituents imparted negative effect to potency ; and ( 3 ) stereochemistry of the secondary hydroxyl group had no influence on potency . the sars indicated that -hematin formation inhibitory activity of target compounds showed correlation with their antiplasmodial and antischistosomal activities , especially the latter . although further investigations are needed , the current data basically confirmed our hypothesis that inhibiting hemozoin formation was one of the mechanisms of action of carbazole aminoalcohols . in order to assess the toxicity of carbazole aminoalcohols , representative compounds ( 611 ) were tested for their cytotoxicity against human embryonic lung fibroblast wi38 . generally , all of the tested compounds showed moderate cytotoxicity with cc50s in the micromolar range . the selectivity indices ( si ) , the ratio of cytotoxicity ( cc50 ) and antiplasmodial activity ( ic50 for pf strains ) , were also calculated ( table  2 ) , allowing to identify the potential therapeutic windows . all the compounds showed acceptable si values in chloroquine - sensitive pf3d7 strain ranging from 19 to 32 , and more satisfactory si were observed in chloroquine - resistant pfdd2 strain ranging from 45 to 126 ( table  1 ) . particularly , compound 7 was the safest molecule with a cc50 value of 7.931  m , a si [ cc50/ic50 ( pf3d7 ) ] value of 32 , and a si [ cc50/ic50 ( pfdd2 ) ] value of 87 . the binding of carbazole aminoalcohols with hematin was investigated by the continuous variation technique ( job 's plot ) . a sharp peak at 401  nm was observed in hematin solution at ph 7.4 , indicating that monomeric hematin predominated under the experimental conditions . the addition of represented carbazole aminoalcohols ( 611 ) led to the decrease of absorption in the soret band , indicating the association of compounds and hematin . when the molar ratios of hematin and compounds were 1:1 , changes in absorbance intensity reached the maximum ( see fig .  3 for representative plot of 7 ) . in contrast with 7 and 9 , compounds 6 and 8 produced minor changes in absorbance ( data not shown ) , suggesting that they interacted weakly with hematin , which was in agreement with their less potent -hematin formation inhibitory activity . molecular docking studies were performed to predict the binding modes of representative compound 7 ( r and s isomers ) with heme by utilizing the c - docker program within discovery studio 2.1 software package . as expected , the docking models ( fig . 4 ) indicated that ( r)-7 and ( s)-7 exhibited similar binding modes with heme . both isomers could form stable non - covalent complexes with heme , which consequently led to the inhibition of hemozoin formation . as observed from the axial view ( fig . 4a and b ) , the carbazole core formed a - stacking interaction with porphyrin ring . the carbazole core was not right above the iron center , but located toward one side of porphyrin ring , presumably to form preferable - interaction . similar binding modes were observed in amodiaquine- , chloroquine- , and quinine - heme complexes ( leed et  al . , 2002 , de dios et  al . , 2004 ) . the aminoalcohol functional groups ( nh and oh ) of ( r)-7 and ( s)-7 formed three hydrogen bonds with both carboxyls of heme , respectively , which probably further stabilized the complex . 4c and d ) showed that the distances between carbazole centroid and heme porphyrin planar of ( r)-7 and ( s)-7 were 3.78 and 3.73   , respectively , which satisfied the optimal interplanar distance value of approximate 3.04.0   ( de sousa et  al . , 2015 ) . in this work , novel carbazole aminoalcohols were designed and prepared , and their antiplasmodial and antischistosomal potential have been confirmed . most of the compounds displayed significant -hematin formation inhibitory ability , which showed correlation with their dual antiparasitic activities , identifying that inhibiting hemozoin formation was one of their mechanisms of action . the preliminary sars confirmed the importance of the amine type and the dichlorinated carbazole core . job 's plot revealed that carbazole aminoalcohol compound interacted with hematin through forming a 1:1 complex . it is noteworthy that compound 7 showed not only potent dual antiparasitic activities but also low cytotoxicity , which could be developed as a promising lead compound for further investigation .
malaria and schistosomiasis are two of the most socioeconomically devastating parasitic diseases in tropical and subtropical countries . 
 since current chemotherapeutic options are limited and defective , there is an urgent need to develop novel antiplasmodials and antischistosomals . 
 hemozoin is a disposal product formed from the hemoglobin digestion by some blood - feeding parasites . 
 hemozoin formation is an essential process for the parasites to detoxify free heme , which is a reliable therapeutic target for identifying novel antiparasitic agents . 
 a series of novel carbazole aminoalcohols were designed and synthesized as potential antiplasmodial and antischistosomal agents , and several compounds showed potent in  vitro activities against plasmodium falciparum 3d7 and dd2 strains and adult and juvenile schistosoma japonicum . 
 investigations on the dual antiparasitic mechanisms showed the correlation between inhibitory activity of -hematin formation and antiparasitic activity . inhibiting hemozoin formation 
 was identified as one of the mechanisms of action of carbazole aminoalcohols . 
 compound 7 displayed potent antiplasmodial ( pf3d7 ic50  =  0.248  m , pfdd2 ic50  =  0.091  m ) and antischistosomal activities ( 100% mortality of adult and juvenile schistosomes at 5 and 10  g / ml , respectively ) and exhibited low cytotoxicity ( cc50  =  7.931  m ) , which could be considered as a promising lead for further investigation . 
 stoichiometry determination and molecular docking studies were also performed to explain the mode of action of compound 7 .