Vinum resinatum: Scientists and unintended consequences.

The art of winemaking has a long history. The methods and techniques changed over millennia as did the consumers taste and habits. Improving the taste of the wine and preventing conversion to vinegar required fantasy and creativity. The principal substances employed as conditurae were seawater, turpentine, either pure, or in the form of pitch (pix), tar (pix liquida), or resin (resina); lime, in the form of gypsum, burnt marble, or calcined shells; inspissated must, aromatic herbs, spices, and gums, and these were used either singly, or cooked up into a great variety of complicated confections. Turpentine exposure (oral. dermal. or respiratory) confers urine the scent of violets. It is generally assumed that turpentine's effect on urine was noticed subsequent to its use as medicine, as a component of various remedies popular in antiquity and thereafter. The high price of such elaborate concoctions would have made however such means available to only a privileged few. Furthermore, the high number of components would also have made association of a particular ingredient with a specific effect difficult if not impossible. We examined the possibility that the effect of turpentine on urine was noticed due to its presence in wines and therefore to the likely widespread exposure of the population to its effects. We review the literature supporting this possibility and provide biographic data on some of the pharmacists, chemists, and physicians involved.

Possible metabolic conversion of pinene to ionone.

The unintended consequence of the ingestion of certain foods to alter the scent or color of urine is well known. Less awareness exists regarding the practice of ingestion of natural products or drugs with the intended purpose of conferring urine the scent of violets. The resin of the terebinth tree and the derived turpentine were widely used in antiquity in wine-making, both as taste enhancer and conserving agent, so the effect on urine was possibly noticed due to the presence in wines. It is also possible that turpentine's effect on urine was noticed subsequent to its use as medicine, as a component of various remedies popular in those days. The scent altering effect requires metabolic conversion of pinene, the main turpentine component to ionone, the molecule mainly responsible for the scent of violets. The metabolic pathway (in humans or otherwise) was (to our knowledge) not yet described. We here propose a possible metabolic pathway for the conversion of pinene to ionone, explaining the scent altering effect of turpentine. We also provide calculated pharmacokinetic (pK) data for the mentioned substances.

Veilchenduft, ectopic olfactory receptors and endogenous volatile compounds: Who was Paul Krüger (1859-1916)?

The pharmacist and chemist Ferdinand Tiemann (1848-1899) having succeeded in the synthesis of vanillin, is considered to be the father of Geschmackstoff-Chemie (flavor chemistry). Tiemann, together with Paul Krüger (1859-1916) and then with Friedrich-Wilhelm Semmler (1860-1931), developed a method to obtain with a good yield Veilchenduft (violet scent); they condensed citral with di-methyl-ketone (acetone) thus generating an intermediate which upon exposure to an acidic environment cyclizes to ionone. By doing so the fragrance chemistry was born. Ionone (the compound responsible for the violet scent) was produced on an industrial scale at the factory of Wilhelm Haarmann (1847-1931) in Holzminden, factory renamed 1876 Haarmann & Reimer, after Karl Reimer (1845-1881) joined the group of owners. While a number of chemists and pharmacists were involved in the synthesis of Ionone (Veilchenduft; violet scent) and irone (iris scent), with few exceptions, their biographies are pretty well documented. In contrast, very little transpired about Dr. Paul Krüger, who spent some seven years trying to iron out the difficulties of ionone synthesis. The purpose of this short contribution is to shed some light on the life and work of Paul Krüger while providing an overview on the status of ionone pharmacology and to highlight the historical significance of ionone synthesis.

Cleopatra: from turpentine and juniper to ionone and irone.

Cleopatra VII (69-30 BC), the last Ptolemaic ruler of Egypt, is probably best known for her love affairs with Julius Caesar (100-44 BC) and Marcus Antonius (83-30 BC). Rightly or wrongly she became the epitome of shrewd seduction, leading brave Roman commanders on a path to debauchery and destruction. Among the seductive strategies attributed to her is the ingestion of small amounts of turpentine [the resin of the terebinth tree (Pistacia terebinthus)] or of derived oil (Oleum terebinthinae) with the purpose of conferring to her urine a more pleasant scent reminding of violets. Turpentine components are metabolized among other compounds to ionones and irones, which - renally excreted - are responsible for the flowery scent. Having obviously worked with great generals, the strategy is said to have been embraced for everyday use by many affluent Roman women. Complicating the issue somewhat is the fact that juniper berries (Fructus juniperi) and derived oil (Oleum juniperi) containing many of the same terpenoids as turpentine have a similar effect on urine. The purpose of this contribution is to briefly review the pharmacology of turpentine and juniper derived compounds assumed to be responsible for altering the scent of urine and to examine the origin and veracity of the mentioned habit. While the effect of ingested turpentine on the scent of urine is well documented our attempts at identifying Greek or Latin authors mentioning its intentional use for this explicit purpose (by Cleopatra or anybody else) failed.

The pharmacy in Gersfeld/Rhoen and Veit Jakob Metz (1792-1866).

A "Privileg" for a pharmacy in Gersfeld (Rhoen) was issued October 26, 1788 by the ruler of Gersfeld, the Reichsfreiherr Amand von Ebersberg (1747- 1803), to Peter Franz Wilhelm Feuchter (1766-1835). Feuchter was not only a dedicated pharmacist but also scholarly active both by publishing and by serving on various journal editorial boards. Vitus Jacobus Metz (1792-1866) was accepted 1808 as an apprentice in the pharmacy and later enjoyed private lessons from Feuchter (until around 1813, when he gave up the study of pharmacy to pursue medical studies in Würzburg). This major decision was possibly influenced by Metz experiencing the outcome of a dispute between pharmacist Feuchter and the physician Andreas Laubreis (*1778), dispute with an outcome favoring the physician. As a physician Metz great achievement was to establish 1830 the Mariannen-Institut, the lying-in asylum in Aachen, Bendelstrasse, the first such institution in Germany. How revolutionary and way ahead of its time the Mariannen-Institut really was can only be understood considering that it took over half a century until a similar institution, the second one in Germany, opened in Düsseldorf. With this short contribution we attempt to shed some light on the life and family of Veit Jakob Metz from Römershag (Bad Brückenau) and on the Gersfeld pharmacy, the place that played such a major role in shaping his personality.

Neuropathic organophosphates: from Scrugham, Heim and Lorot to Jake leg paralysis.

Henry Scrugham (1811-1898), the father of triphenyl-phosphate, was a student of Alexander Williamson (1824-1904), Professor of analytical and practical chemistry at the University College London. Williamson using the approach perfected by Scurgham reacted phosphorus pentachloride with cresol (a mixture of ortho, para and meta isomers) thus obtaining tricresyl phosphate (TCP). The triesters of phenol, cresol and naphtol were prepared with a higher yield by Rudolf Heim (1861-1919) by their respective reaction with phosphorus oxychloride (POCl3). Heim is also the first one to obtain pure tri-o-cresyl phosphate (TOCP). In the meantime French pharmacist Jules Brissonnet (1859-1915) synthesized creosote phosphate (containing i.a. TOCP) and popularized its use in the treatment of pulmonary phthisis (tuberculosis). Camille Lorot (1872-1951) and others in France and Germany recognized the ability of creosote phosphate to induce polyneuropathies but this knowledge did not prevent the Ginger Jake epidemic (Jake leg) of the 1930s in the US. The Jake induced neuropathy was first recognized and described in Oklahoma City by a General Practitioner, Ephraim Goldfain (1894-1983). Soon thereafter Maurice Isadore Smith (1887-1951), a pharmacologist, and chemist Elias Elvove (1883-1962) identified TOCP in Jamaican ginger extract as the causative agent. We attempt to shed some light on the life and family of the less known chemists, pharmacists and physicians associated with the synthesis of neuropathic organophosphates and with the recognition of their toxicity.

Inhibitory actions of bisabolol on α7-nicotinic acetylcholine receptors.

Bisabolol is a plant-derived monocyclic sesquiterpene alcohol with antinociceptive and antiinflammatory actions. However, molecular targets mediating these effects of bisabolol are poorly understood. In this study, using a two-electrode voltage-clamp and patch-clamp techniques and live cellular calcium imaging, we have investigated the effect of bisabolol on the function of human α7 subunit of nicotinic acetylcholine receptor (nAChR) in Xenopus oocytes, interneurons of rat hippocampal slices. We have found that bisabolol reversibly and concentration dependently (IC50 = 3.1 µM) inhibits acetylcholine (ACh)-induced α7 receptor-mediated currents. The effect of bisabolol was not dependent on the membrane potential. Bisabolol inhibition was not changed by intracellular injection of the Ca(2+) chelator BAPTA and perfusion with Ca(2+)-free solution containing Ba(2+), suggesting that endogenous Ca(2+)-dependent Cl(-) channels are not involved in bisabolol actions. Increasing the concentrations of ACh did not reverse bisabolol inhibition. Furthermore, the specific binding of [(125)I] α-bungarotoxin was not attenuated by bisabolol. Choline-induced currents in CA1 interneurons of rat hippocampal slices were also inhibited with IC50 of 4.6 µM. Collectively, our results suggest that bisabolol directly inhibits α7-nAChRs via a binding site on the receptor channel.

Synthesis of tetraethyl pyrophosphate (TEPP): from physician Abbot and pharmacist Riegel to chemist Nylen.

Tetraethyl pyrophosphate (TEPP) made history not only as the first man-made organophosphate cholinesterase inhibitor but also as a most successful commercial product traded under a good number of names. The substance was first synthesized by a Russian chemist, Wladimir Petrovich Moshnin, while studying in Paris as an eleve (student) of Wurtz. The synthesis was soon thereafter repeated and reported to the Academy of Sciences by Philippe de Clermont, another student of Wurtz, who acknowledged the earlier work of Moshnin. Holmstedt in his chapter dealing with the beginnings of organophosphate chemistry in Koelle's Textbook Cholinesterases and Anticholinesterase Agents concluded his remarks by noting that after the initial synthesis by Moshnin and de Clermont, over the years, a good half-a-dozen of other pharmacists and chemists also managed the feat (of synthesizing TEPP). This led to my attempts at identifying those involved in the synthesis of TEPP. The compiled list turned out to be quite long: Abbot (1879), Riegel (1896), Cavalier (1906), Rosenheim A, Stadler & Jacobsohn (1906), Rosenheim & Pritze (1908), Balareff (1914), Nylen (1930), Arbusow & Arbusow (1931), Schrader (1938), Woodstock (1946) and Toy (1948). This report while summarizing the synthetic approach used in obtaining TEPP by the respective scientists mainly attempts to shed light on the life of the less known pharmacists and chemists involved in the synthesis of TEPP. The focus is on the pre-industrial synthesis period ending with Nylen largely because details on the Arbusow family, as well as on Schrader and Toy are fairly well known or have recently been described.

Pharmacists Adolf Schall and Ernst Ratzlaff and the synthesis of tabun-like compounds: a brief history.

The history of the synthesis of organophosphate inhibitors of cholinesterase starting with the synthesis of tetraethyl-pyrophosphate by Moschnin(e) and de Clermont and leading to the recognition about half a century later of the toxicity of the phosphor ester by Lange and von Krueger has been told in great detail previously. An almost parallel history -described originally by Bo Holmstedt--exists for organophosphonate inhibitors of cholinesterase starting with the synthesis (1898) in Rostock of diethylamido-ethoxy-phosphoryl-cyanide by the pharmacist Adolph Schall (1870-1957), a graduate student of August Michaelis (1847-1916), the re-examination of the chemical structure of the Schall compound (1903) by Michaelis, recognition (1937) of the toxicity of class by Gerhard Schrader (1903-1990) and confirmation (1951) of the structure by Bo Holmstedt (1919-2002). This short report attempts to shed some light on the life of the pharmacists and chemists involved in the synthesis of the first P-CN organophosphonate inhibitor of cholinesterase, focusing on the two less known pharmacists, the graduate students of Professor Michaelis Adolph Schall and Ernst Ratzlaff (1870-1948).

Study on medicinal chemistry of K203 in wistar rats and beagle dogs.

K203 is an experimental bis-pyridinium mono-aldoxime type cholinesterase reactivator of potential use in organophosphate/ organophosphonate poisoning. Pharmacokinetics of K203 were examined in Wistar rats and beagle dogs using ion-pair HPLC. Serum and cerebrospinal fluid concentrations of K203 were determined using ion-pair reversedphase chromatography on octadecyl silica column. HPLC with ultraviolet detection was used for determination of serum concentration of K203 higher than 0.1 µg/mL while its low concentrations in cerebrospinal fluid required electrochemical detection (0.015 through 4 µg/mL range). In rats the serum levels of K203 followed zero order pharmacokinetics from 15 to 120 minutes post administration. Zero order pharmacokinetics was also observed in beagle dogs after low dose (15 µmol/kg) of K203 administration. High dose administration (250 µmol/kg) led to subsequent hindered elimination from both cerebrospinal fluid and serum.

The history of pyridinium oximes as nerve gas antidotes: the British contribution.

Irwin B. Wilson, working in the laboratory of David Nachmansohn at Columbia, demonstrated the ability of hydroxylamine to reactivate cholinesterase inhibited by organophosphates. Soon thereafter Wilson and Ginsburg reacted pyridine-2-aldoxime with methyl iodide to synthesize the first pyridinium aldoxime reactivator of clinical relevance, 2-PAM (pralidoxime). Independently, and at the same time, similar work was conducted in Britain at the Chemical Defence Experimental Establishment in Porton by Green leading also to the synthesis of 2-PAM and the recognition of its reactivating properties. While the American contribution is well known, the British achievements were less publicized. The present contribution attempts to shed some light on the life and work of the people who contributed to the early development of cholinesterase reactivators, the pyridinium aldoximes at Porton.

History of arsenic ethers: who was Felix D'Arcet?

Williamson serendipitously discovered (1851) a new and efficient way to produce esters using ethyl iodide and potassium salts and in doing so elucidated the molecular mechanism behind ether formation. Lassaigne (1820) made the analogy between sulphovinic and phosphovinic acids and demonstrated the existence of phosphovinic acid, while Pelouze (1833) synthesised monoethyl phosphovinic acid. Finally 1848 Voegeli produced diethyl phosphovinic acid and the first neutral ester of phosphoric acid, the triethyl phosphate (TEP). The successes of Lassaigne and Pelouze in producing phosphovinic acids and Mitscherlich's theory of isomorphism fuelled the search for the vinic acids of arsenic, phosphorus neighbor in the periodic system. This short report attempts to identify the (less known) pharmacists and chemists involved in the quest for both arsenovinic acids and the neutral esters of arsenic and pyroarsenic acids.

The history of cholinesterase reactivation: hydroxylamine and pyridinium aldoximes.

Hydroxylamine (NH2OH) the substance which will turn out to be of importance to those interested in the treatment of organophosporus cholinesterase inhibitor exposure, was synthesized by Wilhem Clemens Lossen in 1865 while working in Halle as an assistant in the laboratory of Wilhelm Heinrich Heintz. The Lossen synthesis generated hydroxylamine in aqueous solution. Anhydrous hydroxylamine was prepared almost simultaneously by Lobry de Bruyn and Crismer (1891). Using hydroxylamine as a starting point Meyer synthesized aldoximes and ketoximes (1897). Lange, a PhD student of Ladenburg, isolated 2-methyl-pyridine (alpha-picoline). Some fifty years later Wilson, working in the laboratory of Nachmansohn, demonstrated the ability of hydroxylamine to reactivate cholinesterase inhibited by organophosphates. Finally Wilson and Ginsburg using 2-methyl-pyridine as a starting point synthesized the first pyridinium aldoxime reactivator of clinical relevance, pralidoxime (1955).

In an animal model nephrogenic systemic fibrosis cannot be induced by intraperitoneal injection of high-dose gadolinium based contrast agents.

AIM AND OBJECTIVE: Nephrogenic systemic fibrosis (NSF) has been reported in humans to be most likely induced by gadolinium based contrast agents (GBCA), namely by gadodiamide, gadopentetate dimeglumine, and gadoversetamide, rarely by other GBCA. The pathogenesis of NSF remains unclear; different hypotheses are under discussion. The objective of the study is to assess if in the animal model human-like NSF changes can be induced by high-dose, intraperitoneal GBCA injections over four weeks. MATERIALS AND METHODS: After approval by the institutional animal ethics committee, six rats each were randomly assigned to groups, and treated with seven different GBCA. Intraperitoneal (IP) injections - proven in the animal model to be effective - were chosen to prolong the animals' exposure to the respective GBCA. GBCA doses of previous intravenous (IV) animal studies were applied. After five weeks all rats were sacrificed. Sham controls were treated with IP saline injections, employing the same regimen. RESULTS: No findings comparable with human NSF were observed in all animals after IP treatment with all seven GBCA at daily doses of 2.5 and 5.0 mmol/kg body weight (BW). No histopathological abnormalities of all examined organs were noted. Weight loss was stated in weeks three and four with GBCA injections at doses of 5.0 mmol/kg BW, but rats regained weight after cessation of GBCA treatment. CONCLUSIONS: NSF-comparable pathological findings could not be induced by high dose intraperitoneal injection of seven GBCA.

Pharmacist Theodor Salzer (1833-1900) and the discovery of bisphosphonates.

Herbert Fleisch, the father of the therapeutic use of bisphosphonates in modern medicine, repeatedly stated in his numerous reviews that bisphosphonates were first synthesized 1865 in Germany by the Russian chemist Menschutkin. He was wrong on two counts. Had Menschutkin synthesized bisphosphonates, as he was a student of Wurtz at the time of the "synthesis", the birthplace of the substances would have been France and not Germany; but he did not. By reacting phosphorous acid with acetyl-chloride he obtained derivatives of pyro-phosphorous and pyro-phosphoric acids (P-O-P backbone) and not bisphosphonates (P-C-P backbone). The discovery of the first bisphosphonate occurred indeed in Germany but some thirty years later and not without some drama. First 1894 the pharmacist Theodor Salzer (1833-1900) described an impurity contained in commercially available phosphoric acid but failed to identify it as acetodiphosphoric acid, a bisphosphonate. 1896, an undergraduate student, Hans von Baeyer working in Munich at the Royal Academy of Sciences in the chemical laboratory of his father Adolf (the 1905 Nobel Prize laureate and discoverer of the barbiturates) synthesized an unknown substance which his famous father summarily rejected as some "Dreck" or impurity. Only due to the tenacity of young Hans work on the matter was continued and the paper describing the synthesis published a year later. The correctness of the chemical structure of the compound as assumed by von Baeyer (and his Ph.D. supervisor Hofmann) was confirmed 1901 by Heidepriem, a Ph.D. student of Hofmann. This short report attempts to shed some light on the life of the lesser known pharmacists and chemists involved in the synthesis of the first bisphosphonate, focusing on Salzer, Heidepriem and von Baeyer.

Toxicity of phosphor esters: Willy Lange (1900-1976) and Gerda von Krueger (1907-after 1970).

In 1851 Williamson serendipitously discovered a new and efficient way to produce ethers using ethyl iodide and potassium salts. Based on this new synthetic approach, the Frenchman Philippe de Clermont and the Muscovite Wladimir Moschnin, both élèves of Adolphe Wurtz in his Paris School of Chemistry, achieved the synthesis of the first ester of pyrophosphoric acid (TEPP). de Clermont "tasted" the new compound and although TEPP is a potent cholinesterase inhibitor he failed to recognize its toxicity. Almost a century later, in 1932, Willy Lange (1900-1976) and his graduate student Gerda v. Krueger (1907-after 1970) described the toxicity of organophosphonates. While the classic paper of the two "Uber Ester der Monofluorphosphorsäure." is cited by almost everybody working in the field, little is known about Lange and almost nothing about v. Krueger. This brief communication attempts to shed some light on the life of both.

Differential expression of presynaptic genes in a rat model of postnatal hypoxia: relevance to schizophrenia.

Obstetric complications play a role in the pathophysiology of schizophrenia. However, the biological consequences during neurodevelopment until adulthood are unknown. Microarrays have been used for expression profiling in four brain regions of a rat model of neonatal hypoxia as a common factor of obstetric complications. Animals were repeatedly exposed to chronic hypoxia from postnatal (PD) day 4 through day 8 and killed at the age of 150 days. Additional groups of rats were treated with clozapine from PD 120-150. Self-spotted chips containing 340 cDNAs related to the glutamate system ("glutamate chips") were used. The data show differential (up and down) regulations of numerous genes in frontal (FR), temporal (TE) and parietal cortex (PAR), and in caudate putamen (CPU), but evidently many more genes are upregulated in frontal and temporal cortex, whereas in parietal cortex the majority of genes are downregulated. Because of their primary presynaptic occurrence, five differentially expressed genes (CPX1, NPY, NRXN1, SNAP-25, and STX1A) have been selected for comparisons with clozapine-treated animals by qRT-PCR. Complexin 1 is upregulated in FR and TE cortex but unchanged in PAR by hypoxic treatment. Clozapine downregulates it in FR but upregulates it in PAR cortex. Similarly, syntaxin 1A was upregulated in FR, but downregulated in TE and unchanged in PAR cortex, whereas clozapine downregulated it in FR but upregulated it in PAR cortex. Hence, hypoxia alters gene expression regionally specific, which is in agreement with reports on differentially expressed presynaptic genes in schizophrenia. Chronic clozapine treatment may contribute to normalize synaptic connectivity.

The effect of Δ9-tetrahydrocannabinol on 5-HT3 receptors depends on the current density.

The effects of Δ(9)-tetrahydrocannabinol (THC), the psychoactive component of cannabis, on the function of 5-HT type 3 (5-HT(3)) receptors were investigated using a two-electrode voltage clamp technique in Xenopus oocytes, and a whole-cell patch clamp technique in rat nodose ganglion neurons. In oocytes injected with 3 ng cRNA of 5-HT(3A) receptor, THC reversibly inhibited currents evoked with 5-HT (1 µM) in a concentration-dependent manner (IC(50)=1.2 µM). The extent of THC inhibition was inversely correlated with the amount of cRNA injected and the mean 5-HT(3A) receptor current densities. Pretreatment with actinomycin D, which inhibits transcription, decreased the mean 5-HT(3) receptor current density and increased the extent of THC inhibition on 5-HT(3) receptor-mediated currents. The IC(50) values for THC increased from 285 nM to 1.2 µM in oocytes injected with 1 and 3 ng of 5-HT(3A) cRNA, respectively. In radioligand binding studies on membrane preparations of oocytes expressing 5-HT(3A) receptors, THC did not alter the specific binding of a 5-HT(3A) receptor antagonist, [(3)H]GR65630. In the presence of 1 µM THC, the maximum 5-HT-induced response was also inhibited without a significant change in 5-HT potency, indicating that THC acts as a noncompetitive antagonist on 5-HT(3) receptors. In adult rat nodose ganglion neurons, application of 1 µM THC caused a significant inhibition of 5-HT(3) receptors, extent of which correlated with the density of 5-HT-induced currents, indicating that the observed THC effects occur in mammalian neurons. The inhibition of 5-HT(3) receptors by THC may contribute to its pharmacological actions in nociception and emesis.

History of organophosphate synthesis: the very early days.

The early days of ether chemistry can be divided into two periods: before and after Williamson serendipitous discovery (1851) of a new and efficient way to produce ethers using ethyl iodide and potassium salts. In the early part of the 19th century however, before Williamson, the direct reaction between the "spirit of wine" (ethanol) and acids was the only method of generating the elusive "ethers". This brief report looks at the pharmacists-chemists involved in the quest to produce phosphoric acid ether in the pre-Williamson period (Boudet, Boullay, Lassaigne, Pelouze), paving the way for Voegeli's synthesis of triethyl phosphate (TEP) in 1848.

In vitro assessment of the antibiotic efficacy of contrast media and antibiotics and their combinations at various dilutions.

Discography is a controversial diagnostic procedure involving the injection of radiographic contrast medium (RCM) into the intervertebral disc. Iatrogenic bacterial discitis is a rare but serious complication. The intervention has been increasingly performed in our patients here in the United Arab Emirates. Prophylactic intravenous antibiotic administration can reduce post-interventional discitis; however, this may favour the development of bacterial resistance. Direct intradiscal injection of an antibiotic together with the RCM is a potential alternative. To date, there has been only one study on the efficacy of antibiotics added to an RCM. Equally, there are only limited data regarding the potential direct effect of RCM on bacterial growth. The purpose of this study was to determine whether the efficacy of antibiotics is affected when RCM are added. In an in vitro study, the effect of non-ionic RCM on the growth of five laboratory bacterial strains, alone and in combination with three broad-spectrum antimicrobials, was tested. Bacterial growth was assessed in the absence and the presence of RCM, antibiotics and their combinations. All three RCM alone demonstrated some inhibition of bacterial growth at high concentrations. In the presence of the RCM, all three antibiotics retained their inhibitory effect on bacterial growth. In conclusion, our in vitro experiments did not reveal any changes in the antimicrobial efficacy of the three antibiotics in the presence of the three tested RCM. Subsequent clinical trials will need to assess whether intradiscal antibiotic administration may be a suitable substitute for, or a supplement to, prophylactic systemic antibiotics before discography.