Program Version: 2.1 (27-Mar-2022) Program (Re)Started: 04/01/2022 12:44:01, time = 0.53057 d Sample loop purge time for Air (min): 2.00; On port: 10 Sample loop purge time for MC1 (min): 2.00; On port: 1 Sample loop purge time for MC2 (min): 2.00; On port: 2 Sample loop purge time for MC3 (min): 2.00; On port: 3 Sample loop purge time for MC4 (min): 2.00; On port: 4 Sample loop purge time for MC5 (min): 2.00; On port: 5 Sample loop purge time for MC6 (min): 2.00; On port: 6 Loop time (hr): 1.00 --------------------------- Experiment Description ---------------------------- Starting Exp 4 with inoculum from Siders pond 600 mL per reactory Pond water was incubated at 20 C for 36 hrs. Salinity was 3.8 and pH 7.7 and temp was 8.39 ------------------------------------------------------------------------------- Note Number: 1 (04/06/2022 09:18:44 time = 5.38801 d) pH probe in MC 5 got exposed to air from t = 4.55 to 5.37 d ------------------------------------------------------------------------------- Note Number: 2 (04/06/2022 10:12:51 time = 5.42559 d) Note, current volume of reactors is about 1 L. One more sample before the start of chemostat mode today ------------------------------------------------------------------------------- Note Number: 3 (04/06/2022 10:38:38 time = 5.44350 d) Reactor volumes differ. Currently they are appoximiately MC1: 1300 mL MC2: 1300 mL (actually, MC1 is closer to 1400 mL) MC3: 1300 mL MC4: 1200 mL MC5: 1000 mL MC6: 1200 mL ------------------------------------------------------------------------------- Note Number: 4 (04/06/2022 17:41:36 time = 5.73722 d) Pumps turned on at a dilution rate of 1 per day ------------------------------------------------------------------------------- Note Number: 5 (04/07/2022 08:43:57 time = 6.36385 d) An addition 1 mL of ~1 N NaOH was added to the barrel to raise pH in the microcosms ------------------------------------------------------------------------------- Note Number: 6 (04/07/2022 11:33:12 time = 6.48139 d) Added 5 mL of 1M NaOH to 200L barrel at ~715 AM and 1 mL of the 1M NaOH at 1132 AM ------------------------------------------------------------------------------- Note Number: 7 (04/07/2022 11:38:29 time = 6.48506 d) ------------------------------------------------------------------------------- Note Number: 8 (04/07/2022 11:39:00 time = 6.48542 d) Adjusted heights of chemostats to ~3L mark at 1139AM ------------------------------------------------------------------------------- Note Number: 9 (04/08/2022 10:25:38 time = 7.43447 d) Checked on chemostats. Look good. I did some small adjustments to exit tube to increase chemostat volume/water level in a few, but minor. Also notice some bunching of tubintg on the cartrige pump heads for both media. Pulled the tubing out some to avoid failure. We will need to monitor this. ------------------------------------------------------------------------------- Note Number: 10 (04/11/2022 11:46:51 time = 10.49087 d) Began sampling for TCF5 - Chemostat flush T5, POC and Feed ------------------------------------------------------------------------------- Program Version: 2.1 (27-Mar-2022) Program (Re)Started: 04/11/2022 11:55:56, time = 10.49718 d Sample loop purge time for Air (min): 2.00; On port: 10 Sample loop purge time for MC1 (min): 2.00; On port: 1 Sample loop purge time for MC2 (min): 2.00; On port: 2 Sample loop purge time for MC3 (min): 2.00; On port: 3 Sample loop purge time for MC4 (min): 2.00; On port: 4 Sample loop purge time for MC5 (min): 2.00; On port: 5 Sample loop purge time for MC6 (min): 2.00; On port: 6 Loop time (hr): 1.00 --------------------------- Experiment Description ---------------------------- Restarting program after crash ------------------------------------------------------------------------------- Note Number: 4 (04/11/2022 14:33:14 time = 10.60641 d) Some problems with the ASW pump for MCs 5 and 6, and a but of MC 4. Olivia is working on fixing the problem ------------------------------------------------------------------------------- Note Number: 5 (04/12/2022 14:18:31 time = 11.59619 d) Still having issues with the ASW pump. There appears to be a leak in the tubing feeding the manifold. I think we fixed it though. ------------------------------------------------------------------------------- Note Number: 6 (04/12/2022 14:19:30 time = 11.59688 d) Did a minor adjustment on the occulsion setting on the C+TE pump #3. Turned out (counter clockwise) the occlusion screw 1/2 to 1 turn. See if that helps the lower respiration in MC 3 ------------------------------------------------------------------------------- Note Number: 7 (04/12/2022 14:23:52 time = 11.59991 d) Calibrating ASW pump after tubing was changed this morning ------------------------------------------------------------------------------- Note Number: 8 (04/13/2022 13:17:41 time = 12.55395 d) began sampling TCF7 - feed and POC samples at chemostat flush day 7 ------------------------------------------------------------------------------- Note Number: 9 (04/14/2022 10:18:36 time = 13.42958 d) checking air flow ------------------------------------------------------------------------------- Note Number: 10 (04/14/2022 11:08:24 time = 13.46417 d) Put bubble catcher on-line to help keep the ASW feed at the right flow rate ------------------------------------------------------------------------------- Note Number: 11 (04/14/2022 11:23:09 time = 13.47441 d) Leur lok connector to Oxigraf was a bit lose! ------------------------------------------------------------------------------- Note Number: 12 (04/14/2022 11:55:49 time = 13.49710 d) Collect waste water and measured flow rate, which came to 17.1 L/d, which is a bit lower that the desired 18 L/d ------------------------------------------------------------------------------- Note Number: 13 (04/15/2022 13:03:01 time = 14.54376 d) Exit flow rate was measured over an hour and was 16.1 L/d, so it's dropped a bit more from yesterday ------------------------------------------------------------------------------- Note Number: 14 (04/15/2022 13:52:01 time = 14.57779 d) Changing 12C Carbon + Trace Element media and checking calibrations for the C+TE pump. Will turn off the C+TE pump to all chemostats for a short period of time ------------------------------------------------------------------------------- Note Number: 15 (04/15/2022 14:01:08 time = 14.58412 d) Increased ASW pump rate from 14.85 mL/min to 16.6 mL/min to increase flow rate to the desired 18 L/d. Note, this pump has a 5:1 gear reduction so set flow rate on pump is 5 times greater that what it pumps at. ------------------------------------------------------------------------------- Note Number: 16 (04/15/2022 15:03:22 time = 14.62734 d) New N2 tank on line. 2590 psig. The previous tank still had 480 psig. So N2 consumption is still 80 psig per day or so. ------------------------------------------------------------------------------- Note Number: 17 (04/15/2022 15:07:06 time = 14.62993 d) C+TE pump was calibrated. It was pretty close. Only the occlusion of some of the cartridges were adusted to get slightly higher flow rate ------------------------------------------------------------------------------- Note Number: 18 (04/16/2022 11:10:26 time = 15.46558 d) Note, after calibration of C+TE pump, the pump stayed at high flow (perhaps 10x higher) for about an hour before it was noticed and fixed. ------------------------------------------------------------------------------- Note Number: 19 (04/16/2022 13:31:37 time = 15.56362 d) Changing ASW+Nutrients barrel. Pausing ASW feed while switching to 21L ------------------------------------------------------------------------------- Note Number: 20 (04/16/2022 13:44:44 time = 15.57273 d) Changed to 21L while OAhern refills 200L barrel ------------------------------------------------------------------------------- Note Number: 21 (04/16/2022 14:14:33 time = 15.59344 d) Added 1L of bleach to output garbage can. Will let set for ~1hr and drain ------------------------------------------------------------------------------- Note Number: 22 (04/16/2022 14:23:19 time = 15.59953 d) began sampling for POC while 200L barrel sits and salts dissolve ------------------------------------------------------------------------------- Note Number: 23 (04/16/2022 15:28:04 time = 15.64449 d) Turning off ASW pump for formal pump calibration and switch from 21L to 200L barrel ------------------------------------------------------------------------------- Note Number: 24 (04/16/2022 15:58:47 time = 15.66582 d) Finished calibrations and started ASW pump to 200 L barrel ------------------------------------------------------------------------------- Note Number: 25 (04/19/2022 11:51:38 time = 18.49419 d) turned off C feed to calibrate for 13C chemostat start ------------------------------------------------------------------------------- Note Number: 26 (04/19/2022 13:50:58 time = 18.57706 d) Started 13C Feeds on all chemostats (except MC1). Start of Chemostat phase ------------------------------------------------------------------------------- Note Number: 27 (04/19/2022 19:06:20 time = 18.79606 d) Begin sampling TC0.3 - POC and C Substrates about 6 hours after start ------------------------------------------------------------------------------- Note Number: 28 (04/19/2022 19:40:02 time = 18.81947 d) Noticed C+TE pump had a failure (watchdog #18) and stopped pumping. Pump was turned off then on, and is now working ------------------------------------------------------------------------------- Note Number: 29 (04/20/2022 08:20:37 time = 19.34765 d) Starting TC0.75 - POC and C substrates. Using TC0.5 labelled sample tubes ------------------------------------------------------------------------------- Note Number: 30 (04/20/2022 12:13:28 time = 19.50935 d) Turning off C+TE pump to change out to new pump. No C flow while changing the pumps out ------------------------------------------------------------------------------- Note Number: 31 (04/20/2022 12:35:04 time = 19.52435 d) Took out C lines to calibrate new C+TE pump ------------------------------------------------------------------------------- Note Number: 32 (04/20/2022 14:57:38 time = 19.62336 d) Due to large 13C perturbation, we decided to restart the experiment So for the next 5 or so days we will be switching back to 12C and then back again to 13C ------------------------------------------------------------------------------- Note Number: 33 (04/20/2022 15:17:56 time = 19.63745 d) Switch back to 12C carbon. Letting the label flush out again in chemostat mode until back to 13C labelled chemostat. ------------------------------------------------------------------------------- Note Number: 34 (04/21/2022 09:56:20 time = 20.41412 d) Turning off ASW and C+TE pumps to switch. New C+TE pump is running a little high because its model is for higher flow rates. Pump switch will be followed by calibrations ------------------------------------------------------------------------------- Note Number: 35 (04/21/2022 10:55:39 time = 20.45531 d) ASW pump calibrated and back online. Variance between flow rates about 0.2 mL ------------------------------------------------------------------------------- Note Number: 36 (04/21/2022 12:52:07 time = 20.53619 d) C+TE pump back online and calibrated ------------------------------------------------------------------------------- Note Number: 37 (04/24/2022 15:49:21 time = 23.65927 d) Noticed C flow was too high for MC6 - tighted the occlusion screw a bit. C flow for MC2 was low and loosened occlusion screw ------------------------------------------------------------------------------- Note Number: 38 (04/25/2022 16:30:37 time = 24.68793 d) Turning off C+TE feed to change C+TE bottles ------------------------------------------------------------------------------- Note Number: 39 (04/25/2022 16:31:44 time = 24.68870 d) CHanged 12C C+TE bottle. Returned to normal pump operations ------------------------------------------------------------------------------- Note Number: 40 (04/25/2022 18:33:22 time = 24.77317 d) pausing C+TE feed to make sure flow rate on MC2 is the same ------------------------------------------------------------------------------- Note Number: 41 (04/25/2022 18:42:35 time = 24.77957 d) Flow rate the same for all MCs ------------------------------------------------------------------------------- Note Number: 42 (04/26/2022 11:42:01 time = 25.48751 d) Pausing ASW and C+TE Lines. Dip in respiration around T 24.83. C+TE pump failure - Digitial display indicated no error but was not spinning. Will switch around pumps and replace C+TE pump. ------------------------------------------------------------------------------- Note Number: 43 (04/26/2022 13:49:24 time = 25.57597 d) Changed pumps and recalibrated both C+TE and ASW pumps. All lines hooked back up and running again. ------------------------------------------------------------------------------- Note Number: 44 (04/28/2022 10:18:29 time = 27.42950 d) pausing ASW to switch to 21L ------------------------------------------------------------------------------- Note Number: 45 (04/28/2022 11:29:42 time = 27.47896 d) ASW back on. Bubbler broke. Starting chemostat switch ------------------------------------------------------------------------------- Note Number: 46 (04/28/2022 12:05:26 time = 27.50377 d) Did not do switch yet, just took community samples ------------------------------------------------------------------------------- Note Number: 47 (04/28/2022 12:53:27 time = 27.53712 d) Tightened occlusion screw for MC6 C+TE maybe 1/16". Starting chemostat-go-round with MC2, 180 mL into MC1, MC3-6. Followed by Siders reinoculation. ------------------------------------------------------------------------------- Note Number: 48 (04/28/2022 15:05:04 time = 27.62852 d) Cross transfers of chemostats and 600 mL of Siders inoculation complete. ASW still in 21 L barrel ------------------------------------------------------------------------------- Note Number: 49 (04/28/2022 15:45:43 time = 27.65675 d) Pause ASW to switch back to 200L barrel ------------------------------------------------------------------------------- Note Number: 50 (04/28/2022 15:51:01 time = 27.66043 d) ASW back online, with 200L barrel ------------------------------------------------------------------------------- Note Number: 51 (04/28/2022 16:53:49 time = 27.70404 d) Pausing ASW temporarily to replace crazy glued bubble catcher that had a hairline fracture ------------------------------------------------------------------------------- Note Number: 52 (04/29/2022 11:29:23 time = 28.47874 d) Replaced bubble catcher with new one. Both pumps off for less than 5 min. ------------------------------------------------------------------------------- Note Number: 53 (04/29/2022 15:58:30 time = 28.66562 d) Around noon today it was noticed that MC2 was about 250 mL above the 3 L mark (oops, that's MC1), and MC2 was perhaps ~100 mL above its 3 L mark. The exit flow on MC1-3 was increased from 3.5 to 3.8 L/d. Checking levels now show that they are dropping, but only perhaps 50-100 mL. Consequently, exit flwo was increased to 4.0 L/d just to be sure. ------------------------------------------------------------------------------- Note Number: 54 (05/02/2022 12:02:43 time = 31.50189 d) Exit flow measured at 15.24 L/d, but should be 18.00 L/d ------------------------------------------------------------------------------- Note Number: 55 (05/02/2022 12:11:59 time = 31.50832 d) Added the 21 L of extra ASW media to the barrel. ------------------------------------------------------------------------------- Note Number: 56 (05/02/2022 16:58:09 time = 31.70705 d) Exit flow rate measured at 15.74 L/d Increased exit flow pump from 14.85 L/d to 16.00 L/d as a results ------------------------------------------------------------------------------- Note Number: 57 (05/04/2022 09:16:56 time = 33.38676 d) Pausing C+TE to switch to 13C ------------------------------------------------------------------------------- Note Number: 58 (05/04/2022 09:29:58 time = 33.39581 d) pausing asw to calibrate ------------------------------------------------------------------------------- Note Number: 59 (05/04/2022 10:01:24 time = 33.41764 d) ASW calibrated and back on ------------------------------------------------------------------------------- Note Number: 60 (05/04/2022 10:12:29 time = 33.42533 d) C+TE back on ------------------------------------------------------------------------------- Note Number: 61 (05/04/2022 10:22:17 time = 33.43214 d) Chemostat 13C labelling TO. removing C+TE and ASW lines for feed samples ------------------------------------------------------------------------------- Note Number: 62 (05/04/2022 10:58:18 time = 33.45715 d) ASW and C+TE lines back in after collecting feed samples ------------------------------------------------------------------------------- Note Number: 63 (05/04/2022 11:52:38 time = 33.49488 d) Pausing ASW to double check calibration ------------------------------------------------------------------------------- Note Number: 64 (05/04/2022 12:11:48 time = 33.50819 d) Quickly checked ASW for ~5 min. Flow rate was fine. It just looks faster because the barrel is lower. Done Sampling TC0 and Everything is tucked away for now ------------------------------------------------------------------------------- Note Number: 65 (05/04/2022 16:08:15 time = 33.67240 d) Begin sampling for TC0.25*. Carbon substrates and PO13C 6 hours after 13C addition ------------------------------------------------------------------------------- Note Number: 66 (05/04/2022 21:39:34 time = 33.90248 d) Beginning to sample TC0.5* - PO13C and C substrates (GFF filtered) 12 hours after addition of 13C substrates. Using TC0.75* labelled sampling tubes ------------------------------------------------------------------------------- Note Number: 67 (05/05/2022 09:48:14 time = 34.40850 d) Begin sampling TC1 - full sampling 1 day after 13C addition ------------------------------------------------------------------------------- Note Number: 68 (05/06/2022 10:56:34 time = 35.45595 d) Begin Sampling TC2 - Full sampling from chemostats ------------------------------------------------------------------------------- Note Number: 69 (05/06/2022 11:13:00 time = 35.46736 d) taking out lines for feed samples ------------------------------------------------------------------------------- Note Number: 70 (05/06/2022 11:46:59 time = 35.49096 d) Collected ~70 mL for feed from each MC, putting C+TE and ASW lines back in ------------------------------------------------------------------------------- Note Number: 71 (05/06/2022 11:56:50 time = 35.49780 d) All MCs have lines returned and excess feed sample returned to their respective MCs. Flow rates as follows: MC1 69 mL MC2 70 mL MC3 65 mL MC4 68 mL MC5 64 mL MC6 66 mL No exact timer was on each of them. approximately (+/- 1 min) same amount of time for each MC feed ------------------------------------------------------------------------------- Note Number: 72 (05/06/2022 12:18:26 time = 35.51280 d) In regards to Note 71 - I had only sampled MC1-3, taken ~100 mL from MC4, and note sampled MC5-6 inbetween the O2/CO2 sampling at 11 - 12 ------------------------------------------------------------------------------- Note Number: 73 (05/07/2022 09:30:37 time = 36.39626 d) Start sampling TC3 ------------------------------------------------------------------------------- Note Number: 74 (05/08/2022 11:14:00 time = 37.46806 d) Start sampling TC4, no feed samples ------------------------------------------------------------------------------- Note Number: 75 (05/09/2022 11:39:43 time = 38.48592 d) Begin sampling TC5 ------------------------------------------------------------------------------- Note Number: 76 (05/10/2022 10:03:54 time = 39.41938 d) Changed back to original N2 tank which has about 450 psig left ------------------------------------------------------------------------------- Note Number: 77 (05/10/2022 10:52:06 time = 39.45285 d) Pausing ASW temporarily to switch to 2L for refilling 200 L ------------------------------------------------------------------------------- Note Number: 78 (05/10/2022 11:59:15 time = 39.49948 d) Pausing pump again for <5 min to switch to 200 L barrel ------------------------------------------------------------------------------- Note Number: 79 (05/10/2022 12:03:41 time = 39.50256 d) ASW 200L refilled and online ------------------------------------------------------------------------------- Note Number: 80 (05/11/2022 10:14:14 time = 40.42655 d) Start sampling TC7 ------------------------------------------------------------------------------- Note Number: 81 (05/13/2022 09:19:21 time = 42.38844 d) Increased the occlusion on MC5 by perhaps 1/16". In theory this will reduce the ASW flow rate to MC5 and may increase it's observed respiration, as that is a bit low compared to the other 5 MCs ------------------------------------------------------------------------------- Note Number: 82 (05/13/2022 13:50:18 time = 42.57660 d) Decreased the occlusion (less) on C+TE pump for MC5. Very tiny change. ------------------------------------------------------------------------------- Note Number: 83 (05/13/2022 15:27:11 time = 42.64388 d) Switch N2 tank to Air because it only had ~100 psig left. Breathing air should work the same for the dryer, but we'll see ------------------------------------------------------------------------------- Note Number: 84 (05/13/2022 15:28:25 time = 42.64473 d) Sampling after changing to Air instead of N2 for gas dryer ------------------------------------------------------------------------------- Note Number: 85 (05/14/2022 10:06:36 time = 43.42125 d) Start sampling TC10. Taking lines out for feed samples ------------------------------------------------------------------------------- Note Number: 86 (05/14/2022 11:54:33 time = 43.49622 d) Finished sampling TC10. Flow rates for feed are as follows ------------------------------------------------------------------------------- Note Number: 87 (05/14/2022 11:55:11 time = 43.49665 d) Flow rates for feed: MC1-C12 71 mL MC2-Met 72 mL MC3-Eth 67 mL MC4-Ace 69 mL MC5-Glu 65 mL MC6-Xyl 67 mL ------------------------------------------------------------------------------- Note Number: 88 (05/18/2022 10:29:09 time = 47.43691 d) Start sampling TC14 ------------------------------------------------------------------------------- Note Number: 89 (05/18/2022 12:00:46 time = 47.50053 d) Lines out of MC1 and MC2 for feed for ~25 min. Taking lines out MC4 and MC5 ------------------------------------------------------------------------------- Note Number: 90 (05/18/2022 12:26:17 time = 47.51825 d) All lines out for feed samples ------------------------------------------------------------------------------- Note Number: 91 (05/18/2022 13:34:23 time = 47.56554 d) Feed volumes for each MC as follows: MC# Time Volume (mL) MC1-C12 1:26:05 172 MC2-Met 1:26:05 176 MC3-Eth 1:11:05 124 MC4-Ace 1:04:58 123 MC5-Glu 1:04:58 120 MC6-Xyl 1:07:26 125 ------------------------------------------------------------------------------- Note Number: 92 (05/18/2022 13:37:29 time = 47.56770 d) Lines out for ASW calibration check ------------------------------------------------------------------------------- Note Number: 93 (05/18/2022 13:48:50 time = 47.57558 d) ASW offline for end of experiment. Calibration was done to get flow rates from individual ASW lines. Target was 2.8 mL: MC# Volume MC1-C12 2.9 MC2-Met 2.95 MC3-Eth 2.8 MC4-Ace 2.8 MC5-Glu 2.7 MC6-Xyl 2.6 Average: 2.792 Target 14.85 L/day Actual 13.958 L/day Note: this one was on a 5:1 split pump head ------------------------------------------------------------------------------- Note Number: 94 (05/18/2022 14:09:52 time = 47.59019 d) C+TE pump offline end of experiment. Calibration was done to see flow rates of C+TE in all MCs: MC# Volume (mL) MC1-C12 3.8 MC2-Met 3.8 MC3-Eth 3.75 MC4-Ace 3.9 MC5-Glu 4 MC6-Xyl 3.7 Target was 4.0 mL ------------------------------------------------------------------------------- Note Number: 95 (05/18/2022 14:12:11 time = 47.59179 d) Begin terminal batch phase -------------------------------------------------------------------------------