WEBVTT 1 00:00:00.120 --> 00:00:02.600 You know, in most areas of life, hitting like ninety 2 00:00:02.680 --> 00:00:06.719 nine percent on a test is it's cause for celebration, Right. 3 00:00:06.599 --> 00:00:08.279 You get an A plus, you frame it, you. 4 00:00:08.199 --> 00:00:11.160 Move on, exactly, you just move on. But if you're 5 00:00:11.160 --> 00:00:14.400 a pharmacy student, or maybe a young professional drug manufacturing, 6 00:00:14.519 --> 00:00:17.320 or really just someone fascinated by how medicines actually work, 7 00:00:18.120 --> 00:00:21.719 you probably already know that ninety nine percent pure isn't 8 00:00:21.760 --> 00:00:22.559 just a grade. 9 00:00:22.679 --> 00:00:24.120 No, it is absolutely not. 10 00:00:24.039 --> 00:00:27.280 Because that one percent difference that is where arsenic hides. 11 00:00:27.600 --> 00:00:31.879 That is where heavy metals can literally leach into the 12 00:00:31.920 --> 00:00:34.799 medicine from the glass bottle. It's started. 13 00:00:34.920 --> 00:00:37.679 Yeah, it's the gap between a therapeutic cure and a 14 00:00:37.679 --> 00:00:41.320 potential hazard. I mean, in the pharmaceutical world, we don't 15 00:00:41.359 --> 00:00:44.479 treat that remaining one percent is just some rounding error. 16 00:00:45.000 --> 00:00:47.960 It represents a direct threat to patient safety, to drug 17 00:00:48.000 --> 00:00:53.439 potency and therapeutic efficacy, and neutralizing that threat requires a massive, 18 00:00:53.880 --> 00:00:57.320 unbroken chain of quality controlled checkpoints. 19 00:00:57.039 --> 00:01:00.399 Which is exactly why we are dedicating today's deep to 20 00:01:00.439 --> 00:01:04.400 the absolute foundational bedrock of drug safety. We are exploring 21 00:01:04.439 --> 00:01:08.159 the core concepts of pharmaceutical analysis based on some incredible 22 00:01:08.200 --> 00:01:13.400 sorts material, specifically the foundational text Pharmaceutical Analysis, Volume one. 23 00:01:13.560 --> 00:01:15.480 It's essentially the bible for this stuff. 24 00:01:15.640 --> 00:01:17.920 It really is. Our mission today is to trace this 25 00:01:18.120 --> 00:01:23.040 highly precise, meticulously observed chemistry that ensures the pill in 26 00:01:23.040 --> 00:01:25.680 the bottle does exactly what it's supposed to do. 27 00:01:25.920 --> 00:01:29.000 And this isn't just running a single swab under a microscope. 28 00:01:29.040 --> 00:01:32.640 You know, this is a rigorous gauntlet starting the moment 29 00:01:32.719 --> 00:01:34.280 raw materials enter a facility. 30 00:01:34.319 --> 00:01:36.799 So set the stage for us here, what does this 31 00:01:36.959 --> 00:01:38.439 gauntlet actually look like? 32 00:01:38.640 --> 00:01:41.439 Well, the official pharmacopeia is layout a whole series of tests. 33 00:01:41.599 --> 00:01:46.599 You have chemical methods, physical chemical methods, microbiological assays to 34 00:01:46.640 --> 00:01:51.439 test antibiotic potency, and biological methods to measure responses in 35 00:01:51.519 --> 00:01:52.120 living matter. 36 00:01:52.239 --> 00:01:54.640 Wow. So it's a multi layered defense system. 37 00:01:54.480 --> 00:01:57.120 Exactly, because before you can prove a drug is effective, 38 00:01:57.159 --> 00:01:59.799 before you can even measure its potency, you have to 39 00:01:59.799 --> 00:02:03.159 prove it isn't dangerous. You have to hunt down the intruders. 40 00:02:02.840 --> 00:02:06.599 The impurities. And it completely blew my mind to realize 41 00:02:06.599 --> 00:02:10.280 that these impurities, they aren't usually the result of some 42 00:02:10.400 --> 00:02:12.479 sloppy manufacturing error, right. 43 00:02:12.400 --> 00:02:13.280 No, not at all. 44 00:02:13.479 --> 00:02:16.400 They're often an unavoidable consequence of the chemistry. 45 00:02:16.000 --> 00:02:19.080 Itself, they absolutely are. I mean, in organic chemistry, you 46 00:02:19.159 --> 00:02:22.000 almost never get a one hundre yield of just your 47 00:02:22.039 --> 00:02:25.400 target molecule. You're running multi step synthesis. 48 00:02:25.120 --> 00:02:26.000 Right, it's complex. 49 00:02:26.080 --> 00:02:28.639 Yeah, so that means you get side reactions, you get 50 00:02:28.759 --> 00:02:32.159 unreacted intermediate chemicals just left over in the vat. All 51 00:02:32.159 --> 00:02:33.439 of those become impurities. 52 00:02:33.520 --> 00:02:36.159 It's like baking a cake and having random flour and 53 00:02:36.159 --> 00:02:37.400 eggshells left in the pan. 54 00:02:37.639 --> 00:02:41.120 Exactly. And even if your synthesis is perfect, your raw 55 00:02:41.159 --> 00:02:44.159 starting materials might have carried trace amounts of heavy metals 56 00:02:44.319 --> 00:02:45.240 straight out of the ground. 57 00:02:45.360 --> 00:02:47.599 And even if you manage to filter all of that out, 58 00:02:48.240 --> 00:02:52.360 the actual storage containers can sabotage the drugs. Oh absolutely, 59 00:02:52.479 --> 00:02:55.719 Like I read that if a manufacturer stores strong alkalis 60 00:02:55.840 --> 00:02:59.680 in a standard glass container, the alkali literally attacks the 61 00:02:59.680 --> 00:03:02.639 glass and leeches lead right out of it into the 62 00:03:02.680 --> 00:03:03.439 final product. 63 00:03:03.680 --> 00:03:06.879 It's wild, right, or strong chemicals might react with iron 64 00:03:06.960 --> 00:03:11.120 vats extracting iron. Add to that the constant threat of 65 00:03:11.159 --> 00:03:15.800 decomposition from light, oxygen, or moisture over months of storage. 66 00:03:16.120 --> 00:03:17.240 It's a never ending battle. 67 00:03:17.280 --> 00:03:19.560 But wait, hold on, let's unpack this for a second. 68 00:03:19.919 --> 00:03:22.439 If we know about all these contamination risks, and we 69 00:03:22.520 --> 00:03:27.159 have incredible modern technology. Why don't we just purify every 70 00:03:27.159 --> 00:03:29.000 single batch to one hundred percent? 71 00:03:29.080 --> 00:03:30.039 That's the logical question. 72 00:03:30.199 --> 00:03:34.800 Yeah, why do the pharmacopeias set what they call permissive 73 00:03:34.840 --> 00:03:38.439 limits of tolerance? Why accept any impurities at all? 74 00:03:38.719 --> 00:03:40.479 Because of basic healthcare economics? 75 00:03:40.520 --> 00:03:41.879 Okay, meaning what well? 76 00:03:42.400 --> 00:03:48.159 Achieving absolute zero impurity perfection requires endless successive steps of purification. 77 00:03:48.719 --> 00:03:52.439 Every single step costs money, requires energy, and here's the kicker, 78 00:03:52.520 --> 00:03:54.759 it actually loses a bit of the active drug in 79 00:03:54.800 --> 00:03:55.280 the process. 80 00:03:55.319 --> 00:03:57.039 Oh so you're destroying the product to save it. 81 00:03:57.240 --> 00:04:01.400 Exactly. If a manufacturer push for absolute zero, a standard 82 00:04:01.439 --> 00:04:04.599 bottle of everyday ashburd might cost hundreds of dollars. It 83 00:04:04.599 --> 00:04:06.800 would just bankrupt the healthcare system and make the drug 84 00:04:06.840 --> 00:04:09.879 economically unviable for the patients who actually need it. 85 00:04:10.120 --> 00:04:13.759 Okay, So the limit tests are a compromise. We accept 86 00:04:13.800 --> 00:04:17.360 harmless impurities within very strict limits, and we tightly control 87 00:04:17.399 --> 00:04:18.279 the dangerous ones. 88 00:04:18.319 --> 00:04:20.759 That's the reality of the lab, and the tests for 89 00:04:20.879 --> 00:04:25.639 those dangerous impurities are just fascinating pieces of classical chemistry. 90 00:04:26.120 --> 00:04:28.879 Take the limit test for arsenic It utilizes the gut 91 00:04:28.879 --> 00:04:29.360 site test. 92 00:04:29.560 --> 00:04:31.680 Yes, I was looking at the apparatus for this and 93 00:04:31.720 --> 00:04:34.560 it looks like something straight out of a Victorian alchemy lab. 94 00:04:34.800 --> 00:04:38.160 Just this like wide mouthed bottle fitted with a glass tube. 95 00:04:38.279 --> 00:04:42.360 It is brilliantly simple but incredibly sensitive. So you place 96 00:04:42.399 --> 00:04:45.399 your drug sample into the bottle along with zinc and 97 00:04:45.519 --> 00:04:47.439 stan aid hydrochloric. 98 00:04:46.839 --> 00:04:49.560 Acid standated, meaning it's been treated with tin, right. 99 00:04:49.560 --> 00:04:54.279 Got it. This specific mixture is a highly aggressive reducing agent. 100 00:04:54.759 --> 00:04:58.639 It essentially forces electrons onto any arsenic present in the sample, 101 00:04:58.920 --> 00:05:01.040 reducing it into a ga ass called arcine. 102 00:05:01.079 --> 00:05:03.279 And because it's a gas, it travels straight up the 103 00:05:03.319 --> 00:05:04.480 glass tube exactly. 104 00:05:04.720 --> 00:05:07.480 But there's a cotton plug in the way. One soaked 105 00:05:07.480 --> 00:05:10.680 and lead acetate. Why lead acetate because if there's any 106 00:05:10.759 --> 00:05:14.160 hydrogen sulfide gas in the mix, which happens often, it'll 107 00:05:14.240 --> 00:05:18.160 ruin the final reading. The lead acetate cotton physically traps 108 00:05:18.160 --> 00:05:21.399 the hydrogen sulfide. It acts like a chemical filter while 109 00:05:21.439 --> 00:05:23.720 letting the rsale gas pass right through. 110 00:05:23.920 --> 00:05:26.040 Up to the very top of the tube where there's 111 00:05:26.079 --> 00:05:29.560 a piece of mercuric chloride paper, And when that arcine 112 00:05:29.600 --> 00:05:33.319 gas hits the paper, it triggers a reaction that produces 113 00:05:33.360 --> 00:05:37.480 this highly distinct yellow stain, and the analyst literally just 114 00:05:37.639 --> 00:05:40.519 looks at it and compares the intensity of that yellow 115 00:05:41.000 --> 00:05:44.480 against a standard stain produced from a known safe limit 116 00:05:44.560 --> 00:05:45.279 of arsenic. 117 00:05:45.360 --> 00:05:49.279 It's completely visual. If your sample stain is lighter, it passes. 118 00:05:49.879 --> 00:05:52.120 And the limit test for lead operates on a very 119 00:05:52.120 --> 00:05:55.480 similar visual principle. You use a region called dithozone dissolved 120 00:05:55.519 --> 00:05:59.079 in chlorophyll and fizzo. Yeah, when that region extracts lead 121 00:05:59.120 --> 00:06:02.120 from an alkaline solution, it binds to it and forms 122 00:06:02.120 --> 00:06:04.720 this vibrant, unmistakable red complex. 123 00:06:04.759 --> 00:06:07.120 And for general heavy metals, they use hydrogen sulfide. 124 00:06:07.120 --> 00:06:09.800 Ready to do. When the metal ions in the drug 125 00:06:09.879 --> 00:06:13.240 react with the hydrogen sulfide, they form metal sulfides. But 126 00:06:13.279 --> 00:06:14.399 they don't drop to the bottom. 127 00:06:14.399 --> 00:06:14.920 Oh they don't. 128 00:06:15.160 --> 00:06:18.759 No, they stay suspended in a colloidal state, which just 129 00:06:18.800 --> 00:06:21.839 makes the whole liquid look brownish. And again you compare 130 00:06:21.879 --> 00:06:26.279 that brown tint against a standard lid solution, usually capped 131 00:06:26.279 --> 00:06:28.040 to a strict twenty parts per million. 132 00:06:28.480 --> 00:06:30.800 But I mean, I'm stuck on something here. We are 133 00:06:30.839 --> 00:06:34.480 looking for impurities at twenty parts per million. That is 134 00:06:34.519 --> 00:06:38.839 an incredibly tiny needle in a massive vat of chemicals, 135 00:06:38.920 --> 00:06:40.839 it really is, So how do you even grab a 136 00:06:40.920 --> 00:06:43.920 sample to test? You can't just scoop a handful of 137 00:06:43.920 --> 00:06:45.920 pills off the top of a one ton batch and 138 00:06:46.000 --> 00:06:47.639 assume it represents the whole thing. 139 00:06:47.800 --> 00:06:51.040 You've hit on the absolute foundation of all measurement. If 140 00:06:51.079 --> 00:06:54.680 your physical sample doesn't perfectly represent the bulk material, every 141 00:06:54.720 --> 00:06:57.199 single mathematical calculation you run after that. 142 00:06:57.360 --> 00:06:59.720 Is useless, right, garbage in, garbage out. 143 00:06:59.680 --> 00:07:03.600 Exactly, So analyts have to use highly specific sampling techniques. 144 00:07:03.879 --> 00:07:06.160 For powders to use a method called tabling. 145 00:07:06.439 --> 00:07:09.560 Tabling that's where you take the finely divided powder pilot 146 00:07:09.639 --> 00:07:11.639 in the center of a large sheet of oil cloth 147 00:07:12.000 --> 00:07:14.720 and then pull each corner of the cloth one by one. 148 00:07:14.879 --> 00:07:17.759 Yep, it rolls the particles over and over on themselves, 149 00:07:18.079 --> 00:07:20.759 constantly folding the powder back into the center until it 150 00:07:20.800 --> 00:07:22.079 is perfectly homogeneous. 151 00:07:22.199 --> 00:07:25.040 So it's basically like kneading dough, but with chemistry. 152 00:07:25.120 --> 00:07:28.240 Yeah, and it prevents heavier particles from settling at the bottom, 153 00:07:28.480 --> 00:07:32.279 which would totally skew your sample. For compact solids like 154 00:07:32.360 --> 00:07:35.600 chunks of metal, analysts will actually take a drill bore 155 00:07:35.639 --> 00:07:39.079 holes into the material at several selected points collect those 156 00:07:39.079 --> 00:07:41.120 physical drillings and mix them together. 157 00:07:41.480 --> 00:07:45.720 Pharmaceuticals require even more obsession, though if you are testing capsules, 158 00:07:46.199 --> 00:07:49.480 you don't weigh the whole capsule. The gelatin shell isn't the. 159 00:07:49.519 --> 00:07:52.160 Drug, No, it's just the packaging, right, So. 160 00:07:52.120 --> 00:07:54.519 You have to empty it, weigh the powder, and then 161 00:07:54.560 --> 00:07:57.399 you actually have to wash the empty capsule shell using 162 00:07:57.480 --> 00:08:00.399 alcohol and ether. And usually you add a few drops 163 00:08:00.399 --> 00:08:03.720 of acetic acid to the wash. But why the acid to. 164 00:08:03.680 --> 00:08:06.560 Cut through the gelatinous residue. You're trying to dissolve and 165 00:08:06.639 --> 00:08:10.240 capture every last microscopic trace of the active drug that 166 00:08:10.399 --> 00:08:13.319 might be clinging to the inside of that shell. Only 167 00:08:13.399 --> 00:08:16.959 once you have that perfectly representative sample can you dissolve 168 00:08:17.000 --> 00:08:18.600 it and calculate its concentration. 169 00:08:18.920 --> 00:08:22.560 And the math required for standard solutions gets very specific, 170 00:08:22.720 --> 00:08:26.079 very fast. We have molarity, which most people might remember 171 00:08:26.120 --> 00:08:29.319 from high school chemistry, moles of salute per liter of solution. 172 00:08:30.120 --> 00:08:32.440 But then there's molality with an L, which is mules 173 00:08:32.440 --> 00:08:35.519 per one thousand grams of solvent. Why do we need both? 174 00:08:36.200 --> 00:08:38.879 Why introduce grams into a liquid measurement? 175 00:08:38.960 --> 00:08:42.399 Well, think about the physical properties of a liquid. Malarity 176 00:08:42.399 --> 00:08:46.399 relies on volume, right, leaders, Okay, sure, So if the 177 00:08:46.440 --> 00:08:49.320 temperature of the laboratory goes up on a hot summer day, 178 00:08:49.960 --> 00:08:54.519 the liquid in your flask literally expands. The volume changes. 179 00:08:54.600 --> 00:08:55.799 Oh wow, I never thind of that. 180 00:08:55.879 --> 00:08:58.840 Yeah, And if the volume changes, your molarity changes, and 181 00:08:58.879 --> 00:09:01.240 suddenly your concentration calculation is wrong. 182 00:09:01.679 --> 00:09:04.399 So malarity is basically hostage to the room temperature. 183 00:09:04.559 --> 00:09:07.799 Yes, it is, But molality relies entirely on mass weight, 184 00:09:08.159 --> 00:09:10.480 and the weight of a substance doesn't change just because 185 00:09:10.519 --> 00:09:14.559 the room gets warmer. Molality is perfectly temperature independent, which 186 00:09:14.600 --> 00:09:17.360 is crucial for certain highly sensitive analyses. 187 00:09:17.639 --> 00:09:20.440 That makes total sense. Now Here is a concept where 188 00:09:20.440 --> 00:09:21.960 I think a lot of people will get tripped up. 189 00:09:22.480 --> 00:09:27.200 Equivalent weight. A molecule's molecular weight is a fixed physical constant, 190 00:09:27.399 --> 00:09:29.519 like H two O is always H two O, but 191 00:09:29.600 --> 00:09:32.320 its equivalent weight can change depending on what it's doing. 192 00:09:32.480 --> 00:09:35.440 It's a functional value, not a static one. You have 193 00:09:35.480 --> 00:09:37.960 to look at how the molecule is behaving in a 194 00:09:38.000 --> 00:09:38.919 specific reaction. 195 00:09:39.200 --> 00:09:40.720 I was trying to wrap my head around this, and 196 00:09:40.759 --> 00:09:43.480 I keep coming back to like a currency exchange rate, 197 00:09:44.120 --> 00:09:47.480 the value of a chemical its equivalent weight depends entirely 198 00:09:47.519 --> 00:09:49.759 on the specific transaction it's participating in. 199 00:09:49.960 --> 00:09:52.600 That analogy works perfectly. Actually, if you're running an acid 200 00:09:52.639 --> 00:09:56.799 based neutralization, the currency of that transaction is hydrogen atoms. 201 00:09:57.279 --> 00:10:00.440 So the equivalent weight of an acid is is its 202 00:10:00.480 --> 00:10:05.080 molecular weight divided by its basicity basically the number of 203 00:10:05.080 --> 00:10:07.039 replaceable hydrogen atoms it can donate. 204 00:10:07.120 --> 00:10:10.200 So for hydrochloric acid HCl, it only has one hydrogen 205 00:10:10.240 --> 00:10:12.480 to give, so you just divide its molecular weight by one. 206 00:10:12.519 --> 00:10:15.240 The molecular weight and equivalent weight are the same. But 207 00:10:15.399 --> 00:10:20.000 sulfuric acid H two so four has two replaceable hydrogens. 208 00:10:20.559 --> 00:10:23.639 In that transaction, it's paying out double, so you have 209 00:10:23.679 --> 00:10:26.240 to divide its molecular weight by two to find its 210 00:10:26.279 --> 00:10:26.960 equivalent weight. 211 00:10:27.000 --> 00:10:29.320 You've got it. But what happens if you aren't doing 212 00:10:29.320 --> 00:10:32.120 an acid base reaction. What if you are doing an 213 00:10:32.159 --> 00:10:37.000 oxidation reduction reaction or redox titration, The currency isn't hydrogen 214 00:10:37.000 --> 00:10:39.759 atoms anymore. The currency is electrons. 215 00:10:39.879 --> 00:10:43.399 Like the reaction between potassium permanganate and ferrosulfate. 216 00:10:43.600 --> 00:10:47.759 Yes, in that specific acidic environment, the manganese atom inside 217 00:10:47.759 --> 00:10:52.000 the potassium permanganate changes its oxidation state from plus seven. 218 00:10:52.320 --> 00:10:54.919 All the way down to plus two. It is physically 219 00:10:54.919 --> 00:10:58.600 transferred five electrons to make that reaction happen. Oh five, Yeah, 220 00:10:58.639 --> 00:11:02.000 So therefore it's equivalvllant weight for that specific test is 221 00:11:02.039 --> 00:11:05.559 its molecular weight divided by five. If you use the 222 00:11:05.639 --> 00:11:09.120 full molecular weight, your final calculation of the drugs purity 223 00:11:09.159 --> 00:11:10.360 would be wildly incorrect. 224 00:11:10.720 --> 00:11:12.720 And I mean, none of this perfect math matters if 225 00:11:12.759 --> 00:11:15.039 your glassware is lying to you. You can't just trust 226 00:11:15.039 --> 00:11:17.720 the little painted line on the side of a volumetric. 227 00:11:17.200 --> 00:11:21.399 Flask, definitely not. To achieve high accuracy, analysts have to 228 00:11:21.440 --> 00:11:25.720 calibrate their volumetric glassware at exactly twenty seven degrees celsius. 229 00:11:26.559 --> 00:11:29.279 They fill the flask with distilled water up to the mark, 230 00:11:29.720 --> 00:11:32.320 and then they physically weigh the water it holds on 231 00:11:32.360 --> 00:11:34.039 a highly sensitive balance. 232 00:11:33.960 --> 00:11:36.759 Because the density of water at that specific temperature is 233 00:11:36.799 --> 00:11:38.960 a known absolute constant. 234 00:11:39.200 --> 00:11:42.960 Right, you are establishing an unbroken chain of trust from 235 00:11:43.000 --> 00:11:45.879 the careful tabling of the powder, to the temperature independent 236 00:11:45.960 --> 00:11:48.879 math to the physical glass holding the liquid. And once 237 00:11:48.919 --> 00:11:51.480 that chain is secure, you can finally run the core 238 00:11:51.600 --> 00:11:55.200 tests of the lab Volumetric analysis titrations. 239 00:11:55.799 --> 00:11:59.200 The classic image of chemistry right, a buret dripping liquid 240 00:11:59.240 --> 00:12:01.960 into a flask until the whole thing suddenly changes color. 241 00:12:02.320 --> 00:12:05.000 You're adding a standard solution of a non concentration the 242 00:12:05.080 --> 00:12:08.120 titrant into your unknown drug sample until the chemical reaction 243 00:12:08.240 --> 00:12:11.080 is perfectly complete. The equivalence point. 244 00:12:10.840 --> 00:12:14.720 And finding that exact equivalence point relies on chemical indicators. 245 00:12:14.879 --> 00:12:17.399 Let's look at aqueous acid based titrations. You mentioned the 246 00:12:17.440 --> 00:12:20.720 color change. Do you actually know what is happening physically 247 00:12:20.799 --> 00:12:22.840 to the indicator molecule to make it change color? 248 00:12:22.960 --> 00:12:25.960 Well, the text mentioned the Henderson Hasselbulch equation for calculating 249 00:12:25.960 --> 00:12:29.000 the pH of the buffer solutions, but the actual color 250 00:12:29.080 --> 00:12:33.159 change mechanism is wild. It's a literal shape shifting at 251 00:12:33.159 --> 00:12:37.279 the molecular level. Like take methyl orange in an acidic solution, 252 00:12:37.440 --> 00:12:40.679 it looks red. In an alkaline solution, it turns yellow. 253 00:12:40.960 --> 00:12:45.360 It's an internal structural rearrangement. The methyl orange molecule contains 254 00:12:45.360 --> 00:12:46.759 an azolink. 255 00:12:46.399 --> 00:12:49.200 And a hassolink that's a specific double bond connecting to 256 00:12:49.399 --> 00:12:50.159 nitrogen atoms. 257 00:12:50.200 --> 00:12:53.919 Right, exactly as the pH of the solution changes, hydrogen 258 00:12:53.960 --> 00:12:56.440 ions from the liquid either crash into the molecule and 259 00:12:56.519 --> 00:13:00.879 attach or detach from it. This causes the internal electrons 260 00:13:00.919 --> 00:13:04.279 to shift and the entire physical structure of the molecule 261 00:13:04.559 --> 00:13:05.480 rearranges itself. 262 00:13:05.519 --> 00:13:07.559 It's like a molecular transformer, it really is. 263 00:13:07.960 --> 00:13:11.120 That new shape fundamentally changes how the molecule absorbs and 264 00:13:11.159 --> 00:13:11.840 reflects light. 265 00:13:12.360 --> 00:13:14.919 And if an analyst needs a razor sharp color change, 266 00:13:15.000 --> 00:13:18.679 they use a mixed indicator, like combining phenol thalin with 267 00:13:18.759 --> 00:13:22.519 alphanafthol thalin. Instead of a gradual fade, it snaps from 268 00:13:22.559 --> 00:13:25.240 a pale rose street to violet at a very specific 269 00:13:25.240 --> 00:13:26.360 pH of eight point. 270 00:13:26.240 --> 00:13:28.039 Nine right, which is incredibly useful. 271 00:13:28.200 --> 00:13:30.840 But wait, I have a question about these equeous titrations. 272 00:13:30.960 --> 00:13:33.919 I always thought we wanted drugs to be soluble in water. 273 00:13:34.279 --> 00:13:36.919 Why is water suddenly a problem for some of these tests? 274 00:13:37.559 --> 00:13:40.559 Because water is almost too good at its job. Water 275 00:13:40.600 --> 00:13:44.039 has a very high dielectric constant of seventy eight point. 276 00:13:43.799 --> 00:13:45.960 Five, meaning what practically it means. 277 00:13:46.000 --> 00:13:49.200 It's a fantastic insulator that easily wedges itself between charged 278 00:13:49.240 --> 00:13:53.559 particles and pulls them apart. It causes strong ionization. But 279 00:13:53.720 --> 00:13:57.960 many vital drugs are very weak acids or very weak bass. Okay, 280 00:13:58.080 --> 00:14:00.360 so if you put them in water. The water dominates 281 00:14:00.399 --> 00:14:04.519 the environment. The drug either dissolves poorly or its endpoint 282 00:14:04.559 --> 00:14:06.919 becomes so stretched out and blurry that the color change 283 00:14:07.000 --> 00:14:08.919 is just completely unreadable. 284 00:14:08.399 --> 00:14:11.480 So the analyst has to remove water from the equation entirely. 285 00:14:11.639 --> 00:14:14.120 Yes, they use non aqueous titrations. 286 00:14:14.120 --> 00:14:17.639 They swap water out for specialized solvents, protogenic solvents that 287 00:14:17.679 --> 00:14:20.240 act as heavy duty acids to force a weak base 288 00:14:20.279 --> 00:14:23.200 to react, or protophilic solvents that act as bases. 289 00:14:23.559 --> 00:14:26.799 You tailor the environment to force the drug to reveal itself, 290 00:14:27.039 --> 00:14:29.080 and you have to tailor the titrant you drip into it. 291 00:14:29.759 --> 00:14:33.799 For determining acidic substances in a non aqueous environment, analysts 292 00:14:33.799 --> 00:14:37.039 often use an alkali methoxide, but if you walk into 293 00:14:37.080 --> 00:14:40.879 a lab today, you'll see them using lithium methoxide instead 294 00:14:40.879 --> 00:14:42.679 of the more common sodium methoxide. 295 00:14:42.720 --> 00:14:46.759 Because sodium methoxide creates a gelatinous precipitate during the titration. 296 00:14:46.879 --> 00:14:49.480 As it's a mass, it literally turns the solution in 297 00:14:49.559 --> 00:14:52.600 the flag into a giant glob of jelly, which traps 298 00:14:52.600 --> 00:14:55.360 the indicator and makes it impossible to see the color 299 00:14:55.440 --> 00:15:00.000 change or even mix the liquid properly lithium methoxide completely avoid. 300 00:15:00.000 --> 00:15:00.720 It's that trap. 301 00:15:00.879 --> 00:15:05.000 It's those hands on physical realities that define analytical chemistry. 302 00:15:05.840 --> 00:15:09.200 Beyond acid base, we have complexometric titrations which use a 303 00:15:09.240 --> 00:15:13.799 chelating agent called edta. Think of Egta as a molecular claw. 304 00:15:14.000 --> 00:15:17.759 It aggressively seeks out metal ions and wraps itself around them. 305 00:15:17.559 --> 00:15:22.159 And the indicators for these complexometric tests are fascinating more 306 00:15:22.200 --> 00:15:25.320 than black two. For example, when the indicator is bound 307 00:15:25.320 --> 00:15:27.559 to a metal ion in the solution, it shows up 308 00:15:27.559 --> 00:15:30.960 as red, but as you drip the edta into the flask, 309 00:15:31.200 --> 00:15:34.559 the edta claw comes in, rips the metal ion away 310 00:15:34.559 --> 00:15:37.519 from the indicator and bonds with it. Right the mordant 311 00:15:37.519 --> 00:15:40.600 black indicator is left floating free in the solution, and 312 00:15:40.679 --> 00:15:43.200 the moment it is free, it immediately turns blue. 313 00:15:43.600 --> 00:15:48.399 It's incredibly certifying to watch. But sometimes a drug resists 314 00:15:48.440 --> 00:15:52.559 the direct approach. Calcium, for example, is notoriously stubborn and 315 00:15:52.639 --> 00:15:56.120 doesn't give a very sharp, readable endpoint with standard indicators, 316 00:15:56.919 --> 00:16:00.360 so analysts use a technique called a replacement titration. It's 317 00:16:00.440 --> 00:16:01.759 essentially analytical judo. 318 00:16:01.960 --> 00:16:04.320 They add a known amount of a magnesium complex to 319 00:16:04.320 --> 00:16:05.519 the calcium solution. 320 00:16:05.399 --> 00:16:08.080 Rice because calcium happens to form a much more stable 321 00:16:08.120 --> 00:16:11.399 complex with eedta than magnesium doest so, as you add 322 00:16:11.399 --> 00:16:14.159 the edta, it ignores the magnesium and eagerly binds to 323 00:16:14.200 --> 00:16:15.120 the calcium first. 324 00:16:15.000 --> 00:16:15.519 Oh I see. 325 00:16:15.679 --> 00:16:18.879 In doing so, it physically kicks the magnesium out into 326 00:16:18.879 --> 00:16:23.360 the solution. The free magnesium then reacts beautifully and sharply 327 00:16:23.399 --> 00:16:27.120 with the indicator. You're indirectly measuring the calcium by measuring 328 00:16:27.120 --> 00:16:28.480 the magnesium it displaced. 329 00:16:28.679 --> 00:16:31.480 That is brilliant And just to round up the volumetric toolkit, 330 00:16:31.519 --> 00:16:35.279 there are precipitation titrations where the reaction forms a physical solid. 331 00:16:35.559 --> 00:16:39.679 Like mercurometric titrations, Analysts use mercuric nitrate to test for 332 00:16:39.799 --> 00:16:43.639 chlorides or bromides, relying on a diffinal carbozone indicator that 333 00:16:43.679 --> 00:16:46.639 turns a brilliant blue violet when the solid finishes forming. 334 00:16:46.799 --> 00:16:50.600 It is an incredible toolkit. Whether you're dealing with acids, bases, 335 00:16:50.679 --> 00:16:53.799 metal ions, or halogens. There is a tailored lock and 336 00:16:53.879 --> 00:16:56.679 key titration for it. You just have to deeply understand 337 00:16:56.679 --> 00:16:59.320 the molecule to choose the right way to dismantle it. 338 00:17:00.039 --> 00:17:03.120 What if your drug sample isn't a neat clear liquid. 339 00:17:03.360 --> 00:17:05.720 What if it's a cloudy, murky mess where you could 340 00:17:05.759 --> 00:17:09.279 never see a color change anyway. I mean titrations have limits, 341 00:17:09.680 --> 00:17:13.279 and that pushes analysts to rely on physical and electrical properties. 342 00:17:13.799 --> 00:17:18.200 Let's look at colloids. Graphometric analysis usually involves forcing a 343 00:17:18.279 --> 00:17:21.559 chemical to precipitate into a solid, filtering it out, and 344 00:17:21.640 --> 00:17:25.400 just weighing it on a scale. But colloidal suspensions ruin that. 345 00:17:25.559 --> 00:17:28.599 They absolutely ruin it, mainly because of their physical size. 346 00:17:29.079 --> 00:17:33.160 Colloidal particles are incredibly tiny, between one and zero point 347 00:17:33.240 --> 00:17:37.319 one micrometers. They will pass right through the microscopic pores 348 00:17:37.400 --> 00:17:38.680 of standard filter paper. 349 00:17:38.839 --> 00:17:41.720 And you can actually see why they're so stubborn by 350 00:17:41.720 --> 00:17:44.359 shining a powerful beam of light through the liquid the 351 00:17:44.400 --> 00:17:48.359 tinderal effect. The light hits these tiny particles and scatters, 352 00:17:48.599 --> 00:17:50.759 making the beam visible, and if you put that liquid 353 00:17:50.799 --> 00:17:53.720 under an ultra microscope you see something called Brownian movement. 354 00:17:54.079 --> 00:17:57.720 Yes, the particles are so small that the ambient thermal 355 00:17:57.839 --> 00:18:00.240 energy of the water molecules themselves is an have to 356 00:18:00.279 --> 00:18:03.519 keep kicking them around. They're constantly vibrating, bouncing off each other, 357 00:18:03.720 --> 00:18:06.599 and repelling each other due to static electrical charges. 358 00:18:06.680 --> 00:18:08.640 They're like microscopic bumper. 359 00:18:08.359 --> 00:18:12.079 Cars exactly because they never stop moving. They never settle 360 00:18:12.119 --> 00:18:12.599 to the bottom. 361 00:18:12.680 --> 00:18:14.559 So how do you weigh them If you can't filter 362 00:18:14.680 --> 00:18:16.079 them and they won't settle. 363 00:18:15.839 --> 00:18:18.960 You have to force them to clump together into larger, 364 00:18:19.039 --> 00:18:22.880 heavier masses. You have to coagulate them. An analyst might 365 00:18:22.920 --> 00:18:27.039 apply intense heat or vigorous stirring, but the most reliable 366 00:18:27.039 --> 00:18:32.039 method is adding a specific electrolyte. The electrolyte chemically neutralizes 367 00:18:32.119 --> 00:18:35.319 the static electrical charges that are causing the particles to 368 00:18:35.400 --> 00:18:38.799 repel each other. Once the friction is gone, they clump 369 00:18:38.839 --> 00:18:41.559 into a filterable mass and you can finally weigh them. 370 00:18:41.839 --> 00:18:46.240 Wow. Okay, what about complex organics like a massive vat 371 00:18:46.359 --> 00:18:49.160 of fixed oils or fats. You aren't going to get 372 00:18:49.160 --> 00:18:50.880 a color change tight trading castor oil. 373 00:18:51.039 --> 00:18:54.720 No, definitely not. For functional group analysis of organics, analysts 374 00:18:54.839 --> 00:18:58.440 use specific chemical values. The iodine value is a primary one. 375 00:18:58.480 --> 00:19:02.200 It measures the degree of unsatur in an oil, specifically 376 00:19:02.200 --> 00:19:04.920 how many double bonds exist in the long carbon chains. 377 00:19:05.039 --> 00:19:08.319 And they use iodine because halogens like iodine are chemically aggressive, right. 378 00:19:08.400 --> 00:19:12.200 Right, exactly, Well, yes, that is the mechanism The halogen region, 379 00:19:12.440 --> 00:19:16.680 like iodine bromide literally slices into the double bonds of 380 00:19:16.680 --> 00:19:20.200 the carbon chain and attaches itself. By measuring exactly how 381 00:19:20.279 --> 00:19:23.480 much iodine was consumed by the reaction, the analysts can 382 00:19:23.480 --> 00:19:26.119 map out exactly how many double bonds were present in 383 00:19:26.160 --> 00:19:26.559 the oil. 384 00:19:26.880 --> 00:19:30.599 Another way to measure those double bonds is through catalytic hydrogenation. 385 00:19:30.960 --> 00:19:33.759 You literally pump hydrogen gas into the oil to force 386 00:19:33.839 --> 00:19:36.960 those bonds to break. But to make that reaction happen 387 00:19:36.960 --> 00:19:40.279 at room temperature, you need a catalyst, something like incredibly 388 00:19:40.319 --> 00:19:41.920 expensive platinum. 389 00:19:41.519 --> 00:19:43.599 Or rainy nickel rainy bigel. 390 00:19:43.440 --> 00:19:45.359 Yeah, which is a highly active form of nickel that 391 00:19:45.400 --> 00:19:48.440 has to be stored meticulously or it simply degrades and 392 00:19:48.480 --> 00:19:49.400 becomes useless. 393 00:19:49.480 --> 00:19:52.279 And then there's the estra value, which tells an analyst 394 00:19:52.319 --> 00:19:56.000 how much potassium hydroxide is required to neutralize the acids 395 00:19:56.039 --> 00:19:59.640 that are liberated when you completely hydrolyze or break down 396 00:19:59.680 --> 00:20:01.680 one of the oil sample. 397 00:20:01.799 --> 00:20:06.359 These are highly specific, deeply physical methods, but modern analysis 398 00:20:06.440 --> 00:20:09.759 is increasingly moving away from just measuring volume in weight 399 00:20:09.880 --> 00:20:16.319 and moving toward measuring electricity instrumental methods. Let's talk about electrogrivymmetry. 400 00:20:15.519 --> 00:20:16.480 Oh, this is a great one. 401 00:20:16.519 --> 00:20:18.839