the possibility of adsorption on various mediae In one experiment^ 

 stock extract was concentrated by boilings filtered^ boiled with 

 activated caibon^ filtered;, evaporated to a gumnny residue^ re-^ 

 dissolved and tested^ giving a positive response iNOo 125;)o In 

 anotherp the same procedure was followed except that after boiling 

 with activated carbon the filtr-ste was treated with activated aluminaj 

 again with a positive response (Noo 130'o Similar results were 

 obtained with anion fNoo 133) and cation (Noo 134) exchange resinso 

 Apparently the attractant was not fcompletely) adsorbed on any of 

 these media.0 



At this time (September 1952j the chemical work was 

 undertaken at the University of Hawaii rather than the Hawaii Marine 

 Laboratory; the 'estSo of courscp were continued on the fish in Pond 

 Noo 5 at Coconut Islando To obviate the need for refrigeration and to 

 facilitate condensation^ extracts were prepared with 95=percemt ethyl 

 alcohol instead of watero *The actiial strength of the alcohol in the 

 extract was less than 95='percenta depending on the quantity of water 

 in the flesh and the relative quantities of flesh and alcohol which were 

 used; but it was always strong enough to act as a preservative at room 

 temperature o Numerous e>:periment6 showed that this alcohol extract 

 promoied a strong posiSrive response fNoSo 147p lb3s I8O5 235^ etCa)^ 

 whereas there was no response to alcohol alone ilNoo 19)o However„ 

 complete extraction of the attractive substance with alcohol was 

 difficult to attairio In two experiments^ the flesh was treated with 95=' 

 percent alcohol for severad hours and the alcohol, containing some of 

 the attractantj, was filtered o£i and used for further fractionationo 

 Tlie flesh was agdin treated with alcoholg and the alcohol filtered off 

 and usedo The flesh was then treated with water and the filtrate was 

 testedo It still gave a strong response (NoSo 164 and 169) showing that 

 the attractant had not been connpletely removed by the previous two 

 alcohol extractionso To see if complete separation could be achieved 

 with alcohol extractionp flesh was extracted twice with 95-^percent 

 alcohol as fiboveo It was then further extracted with alcohol for 14 

 hours in the Soxhlet appaxatuSo The alcohol waS then removedp and 

 the flesh was extracted for ah. additional 12 hours with distilled watero 

 The cen'rlfuged water extrac*. gave only a slight response (NOo 17\ 

 indicating that almost complete extraction of the attractant had been 

 achieved by the previous alcohol treatmentSo 



Further attention was directed at fracMonjition of the 

 alcohol extract by use of various adsorption medi^. To improve the 

 technique;, columns of the adsorbing media were piep.ij-edc through 

 which the extract was allowed to filter slowly,, Two portions were 

 obtained, the filtrate or material which filtered ♦hrough the column 

 without being adsorbed^ and the eluate or portion which w^s adsorbed 



41 



