Photosensitized Oxidative Damage from a New Perspective: The Influence of Before-Light and After-Light Reaction Conditions

Photooxidative damage is heavily influenced by the presence of bioactive agents. Conversely, bioactive agents influence the local environment, which in turn is perturbed by photooxidative damage. These sorts of processes give rise to a version of the “chicken-and-egg” quandary. In this Perspective, we probe this issue by referring to photooxidative damage in one direction as the light-dark (L-D) sequence and in a second direction as the dark-light (D-L) sequence with a reversed cause and effect. The L-D sequence can lead to the downstream production of reactive molecular species (RMS) in the dark, whereas the D-L sequence can be a pre-irradiation period, such as an additive to limit cellular iron levels to enhance biosynthesized amounts of a protoporphyrin sensitizer. A third direction comes from L-D or D-L sequences, or both simultaneously, which can also be useful for optimizing photodynamics. Photodynamic optimization will benefit from understanding and quantitating unidirectional L-D and D-L pathways, and bidirectional L-D/D-L pathways, for improved control over photooxidative damage. Photooxidative damage, which occurs during anticancer photodynamic therapy (PDT), will be shown to involve RMS. Such RMS include persulfoxides (R2S+OO–), NO2•, peroxynitrate (O2NOO–), OOSCN–, SO3•–, selenocyanogen [(SeCN)2], the triselenocyanate anion [(SeCN)3–], I•, I2•–, I3–, and HOOI, as well as additives to destabilize membranes (e.g., caspofungin and saponin A16), inhibit DNA synthesis (5-fluorouracil), or sequester iron (desferrioxamine). In view of the success that additive natural products and repurposed drugs have had in PDT, a Perspective of additive types is expected to reveal mechanistic details for enhanced photooxidation reactions in general. Indeed, strategies for how to potentiate photooxidations with additives remain highly underexplored.


■ INTRODUCTION
This Perspective describes work from the authors' laboratories and literature on the proliferation of photooxidative reactions in the presence of adjuvants.Reactive oxygen species (ROS) are formed in photosensitized oxidation reactions; however, pathways to optimize photodynamics are needed for improved damage control.A synopsis of this subject could unveil factors that control negative side effects of photosensitized oxidation reactions.
−5 In the first case, free radical species arise from Type-I electron and/or H atom transfer from triplet photoexcited sensitizer to ground state oxygen (e.g., HO • , O 2

•−
, HO 2 • , and H 2 O 2 ).In the second case, Type-II energy transfer leads to the formation of the nonradical, singlet molecular oxygen ( 1 O 2 ).−11 As will be discussed later, additives in photooxidation reactions can lead to reactive molecular species (RMS, where M are species containing not only oxygen, but also nitrogen, sulfur, selenium, and iodine).RMS can range from nitrite radical and sulfite radical anion to iodide radical and iodine radical anion.We will also see that additives can induce phototoxicity by membrane disruption, DNA intercalation, enzyme inhibition, and/or sequestration.
Assessing additives' influence is based on consideration of photooxidative damage in one direction [light-dark (L-D) sequence] and in a second direction [dark-light (D-L) sequence] (Figure 2).In the first case, the light event leads to the production of RMS, then subsequent reactions take place thermally (in the dark).In the second case, a dark event is a postirradiation period, but also can be a pre-irradiation period such as compound incubation prior to the photoproduction of RMS.An example of the D-L sequence is an additive in the dark limiting the cellular iron level to enhance the amount of protoporphyrin (PpIX) sensitizer biosynthesized for an increased photochemical step.Another example of the D-L sequence is to inhibit the export of PpIX.This is also relevant for pre-existing sensitizers, where inhibiting their export will lead to an increased photochemical step in the D-L sequence.
The "chicken and egg" problem of photooxidative damage and the influence of additives or adjuvants in this regard is the subject of this Perspective.The Perspective attempts to uncover how compounds can elicit increased photooxidative damage upon their addition, and includes mechanistic considerations.No previous Review or Perspective exists on this topic.Previous literature was mainly limited to photodynamic thereapy (PDT) for cancer or inactivation of microbes with adjuvants such as natural products, 12 chemotherapeutic agents, 13,14 immunoadjuvants, 15 metal−organic frameworks and dyes for thermal effects, 16,17 plant extracts and essential oils, 18 vitamin D and other differentiation-promotion compounds, 19,20 and even hyperbaric oxygen. 21Reviews on PDT have also described iron-catalyzed reduction of lipid hydroperoxides and nitric oxide scavenging of peroxyl and alkoxy radicals. 22,23There are reviews on the use of PDT itself as an adjuvant in surgical resection 24 and in fluorescence guided resection. 25Previous literature reviews have focused on photooxidation reactions, especially the organic chemistry of singlet oxygen. 26Thus, we believe that a Perspective is needed on photooxidation enhancement in the presence of additives/adjuvants with attention paid to the reactive species formed.
−39 There is less consideration of the two together, and also sparse consideration of RMS subsequent or prior to Type-I and Type-II photooxidation reactions.This Perspective is aimed to foster further research on the subject, and is largely a comparison of species that amplify RMS, which includes natural compounds and repurposed drugs in the coverage as mechanistic insights are sought.

■ SCOPE
In this Perspective, we will discuss sensitized photooxidations in the presence of additives/adjuvants.The first section of the Perspective will include organic 1 O 2 reactions with additives that lead to oxidants that are more powerful than 1 O 2 itself.Subsequent sections of the Perspective will describe the reactive species generated, where data are available.The additives in these sensitized photooxidation reactions include organic sulfides, amino acids, lipids, natural products, repurposed drugs, and inorganic salts.Some of these additives are shown in Figure 3.
We will next compare sequential L-D and D-L pathways, where data are available.Different levels of confidence exist in understanding additive effects, where L-D vs D-L cycles have not yet been deconvoluted.Additives have displayed utility in significantly amplifying sensitized photooxidation reactions, giving us reason to embark on this Perspective.
Amplified Reactivity from Post-irradiation Erythrocyte Lipid and Organic Sulfide ROS.What follows is a discussion of the boosting of reactivity with additives that arises in chemical systems after photooxidation reactions by an L-D reaction sequence (Figures 4 and 5, and Table 1).Here, we summarize instances of amplified photooxidative activity with amino acid additives (Trp, His, Met, Ph 2 S or Ph 2 SO).
In 1984, Krieg and Whitten 40 reported that photooxidation of protoporphyrin (PpIX) was enhanced by the inclusion of certain amino acids and erythrocyte membrane (ghost) lipids (Table 1, entry 1).PpIX underwent self-sensitized photooxidation, producing hydroxyaldehydes (via [4 + 2] 1 O 2 cycloadditions), and formyl products (via [2 + 2] 1 O 2 cycloadditions) (lower arrow, Figure 4).The PpIX photooxidation was enhanced 1.6-, 1.8-, 2.0-, and 1.8-fold by the presence of tryptophan, histidine, methionine, and ghost lipids, respectively.These additives formed peroxy intermediates that converted the porphyrin into biliverdins by subsequent dark reactions.The enhancement was thought to arise from initial 1 O 2 reactions, but even more importantly from subsequent dark reactions from persulfoxides, endoperoxides, and hydroperoxides. 41,42These intermediate    The Journal of Organic Chemistry pubs.acs.org/jocPerspective stabilities vary; for example evidence for the existence of persulfoxides comes from indirect trapping studies, whereas endoperoxides and hydroperoxides are often detected by NMR.−51 Alkyl sulfides [e.g., diethyl sulfide Et 2 S or thietane (CH 2 ) 3 S] react with singlet oxygen to produce sulfoxides (Et 2 SO) and small amounts of sulfone (Et 2 SO 2 ) (Figure 5).For the sulfide− 1 O 2 reaction in the presence of diphenyl sulfoxide (Ph 2 SO) or diphenyl sulfide (Ph 2 S), Ph 2 SO 2 or Ph 2 SO are formed, respectively.Because these aromatic compounds react very slowly with 1 O 2 , the oxidation of Ph 2 SO and Ph 2 S was attributed to a peroxysulfoxide (A), a nucleophilic oxidant, and an S-hydroperoxysulfonium-ylide (B), an electrophilic oxidant), both shown to be stronger oxidants than 1 O 2 itself.In the following section, the use of a cholesterol-derived additive is discussed in the photooxidative eradication of cancer cells.
Amplified Membrane Damage by PDT-Generated Lipid Hydroperoxides Such as Cholesterol Hydroperoxides (ChOOHs).What follows is a discussion of lipid-specific L-D reaction sequence that can occur in cancer cells after a PDT challenge.Relatively low polarity photosensitizers such as PpIX can localize in lipoproteins or cellular membranes making them highly susceptible to photooxidative insults.Although damage to associated proteins can occur, unsaturated lipids such as phospholipids, glycolipids, and cholesterol are more prominent targets due to their overall prevalence.Lipid photooxidation or peroxidation (LPO) can be triggered by a free radical ROS such as hydroxyl radical (HO • ) or by 1 O 2 , the former generated by type I photodynamic reactions and the latter by type II reactions. 9,10Lipid hydroperoxide (LOOH) intermediates are generated in the process (Figure 6).A photogenerated radical can initiate LPO via allylic hydrogen abstraction from an unsaturated lipid, e.g.sn-2 fatty acyl hydrogen in a phospholipid or carbon-7 hydrogen in cholesterol.The resulting lipid radical reacts rapidly with O 2 to give a peroxyl radical (LOO • in general, but 7-OO • for cholesterol specifically).The LOO • then triggers propagative LPO by abstracting hydrogen from another lipid, thereby becoming a hydroperoxide species [designated LOOH/ ChOOH in general or 7α/β-OOH (7-OOH) as a specific positional ChOOH ]. 9,52 Photogenerated 1 O 2 adds directly to an unsaturated lipid to give LOOH with hydrogen retention and double bond allylic shift. 10For cholesterol, the major LOOH/ ChOOH generated from 1 O 2 is 5α-OOH. 52n the presence of suitably ligated Fe 3+ and a reductant such as superoxide or ascorbate, LOOHs can undergo 1-electron reduction to oxyl radical (LO • ) intermediates which, either directly or after rearrangement with O 2 addition to give epoxyallylic peroxyl radicals (OLOO • ), 9,52 abstract hydrogens from other lipids, thereby initiating chain LPO (Figure 6).This is an important example of a post-irradiation, light-independent process that can exacerbate the membrane-damaging effects of photodynamic action alone. 9,10,52hain LPO induced by primary LOOHs can be suppressed by natural lipophilic antioxidants such as α-tocopherol and βcarotene, which intercept free radicals.However, primary and downstream LOOHs are usually susceptible to enzymatic 2electron reduction, which prevents deleterious 1-electron turnover.The only enzyme known to catalyze redox inactivation of ChOOHs as well as PLOOHs in membrane environments is selenoperoxidase GPx4 (∼20 kDa), which uses glutathione (GSH) as a reducing cofactor. 53GPx4 is found in several compartments of mammalian cells, including cytosol and mitochondria, protecting against LPO-inflicted damage/dysfunction 54 or a recently discovered form of cell death called ferroptosis. 54amaging LPO due to 1-electron turnover of photogenerated cellular LOOHs is not necessarily restricted to their membranes of origin.It is now clear that ChOOHs, for example, can translocate to other membranes, and more rapidly than parent cholesterol due to greater polarity of the hydroperoxides. 55ntracellular translocation rate of ChOOHs is increased by trafficking proteins such as sterol carrier protein-2 (SCP-2) and proteins of the steroidogenic acute regulatory (StAR) family. 56,57SCP-2 is a nonspecific lipid transporter, whereas StAR proteins are specific for sterol-based lipids.Mammalian cells overexpressing SCP-2 internalized liposomal 7-OOH more rapidly than controls and died faster by apoptosis due to mitochondrial LPO and loss of membrane potential. 56More recent studies showed that StAR-mediated transport of 7-OOH along with cholesterol to mitochondria in steroidogenic cells caused damage/dysfunction that significantly reduced progesterone output. 58Chain peroxidation of mitochondrial lipids was again found to be mainly responsible.In the following section,  2) is followed by enhanced PpIX formation and greater photodynamic killing.This is a D-L reaction sequence.
Reports have appeared for methotrexate leading to enhancements of PpIX concentrations of 2−4 fold. 59,60−66 Reports have also appeared for desferrioxamine leading to enhancements of PpIX concentrations of 1.2− 1.9 fold. 67,68EDTA also led to a 1.4-fold enhancement, 69 11−17 fold for the HPO dendrimer, 70 1.5−5.0fold for CP94 as adjuvants in ALA-PDT. 71Also, a 2.3−12.4fold enhancement of PpIX was shown for AP2−18. 72Lastly, a report appeared showing a 2.0 fold enhancement PpIX with CP94 as an adjuvant in methyl aminolevulinate (MAL)-PDT. 72The cell types studied in the in vitro and in vivo are also shown in Table 2.
The adjuvants in this section (Amplified 5-Aminolaevulinic Acid (ALA)-PDT in the Presence of Additives) led to 1.5−17 fold PpIX concentration increase in cells, in which corresponding cell photokilling increased from a low of 1.0 to a high of 49fold.In a similar vein, Girotti et al., using ALA-PDT in vitro, have shown photokilling increases of 1.6 to 2.7-fold with adjuvants such as (i) N- (3-(aminoethyl)benzyl)acetamidine (1400W) and [2-[(1-iminoethyl)amino]ethyl]-L-homocysteine (GW271450), which are specific inhibitors of iNOS enzymatic activity, (ii) L-N G -nitroargininemethyl ester (L-NAME), a nonspecific NOS activity inhibitor, or (iii) 2−4-carboxyphenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO), a NO scavenger. 73,74The proposed mechanisms of adjuvant effects in ALA-PDT are often based not only on the effects of iNOS inhibitors or NO traps, but also on cell differentiation and proliferation, upregulation of coproporphyrinogen oxidase (CPO) and downregulation of ferrochelatase (FC), iron sequestration, and sometimes p53 protein expression to increase PpIX concentrations for enhanced cell photokilling.In the following section, the use of inorganic additives are discussed in regard to photooxidative eradication of microbes.
Amplified Photodynamic Killing of Bacteria and Fungi in the Presence of Inorganic Additives.This section summarizes additives that enhance the photooxidized killing where L-D and/or D-L sequences may arise.The additives discussed are NaNO 2 , KSCN, KSeCN, and KI (Table 3).
In 2019, it was reported that the rose bengal (RB)-sensitized photoinactivation of E. coli and S. aureus was substantially enhanced by NaNO 2 (Table 3, entry 1). 75The photokilling enhancement was up to 6 log for E. coli and 2−3 log for S. aureus.The proposed mechanism involved Type-I and Type-II ROS (eq 1−3).Furthermore, 3 RB is thought to react with nitrite (NO 2 − ) to produce nitrogen dioxide radical (NO In 2013, St. Denis et al. 76 described the photodynamic inactivation of E. coli and S. aureus in the presence of KSCN (Table 3, entry 2).The enhancement in photokilling by KSCN was 2.5 log for in S. aureus and 3.5 log for in E. coli.The mechanism for heightened activity was proposed to be a combined effect from 1 O 2 and SO 3 •− , the latter being generated by the singlet oxygenation of SCN − and further oxidation of HSO 3 − anion (eq 4−8).
More recently, Yuan et al. described the MB-sensitized oxidation of E. faecalis in the presence of KI (Table 3, entry 4). 78he enhancement in photokilling was 2−7 log for E. faecalis.The combination of MB and KI was found to possess killing effects in an L-D sequence, where postillumination incubation was carried out under hypoxic conditions.The proposed mechanism involves the production of I 3 − from the reaction with 1 O 2 with I − ; the total quenching rate constant (k T ) of 1 O 2 for I − is 9.1 x 10 7 M −1 s −1 . 79Other participating active agents likely included I 3 − and H 2 O 2 , and possibly I 2 •− and HOO • (eq 11−15).Continuing interest in the use of iodide in photo-dynamic inactivation of bacteria has been reported. 80 Amplified Photodynamic Killing of Bacteria and Fungi in the Presence of Drugs or Surfactant Additives.This section summarizes additives that enhance the sensitized log killing by L-D and/or D-L sequences, such as integrity of the cell membrane compromised for greater cell permeability.The additives discussed are OPE−TTAB complex (oligo-p-phenylene ethynylene tetradecyltrimethylammonium cation), saponin A16, caspofungin, miconazole, and 8-methylnon-7-ene-1 sulfonate (Table 4).
About ten years ago, it was reported that oligo-p-phenylene ethynylene (OPE)-sensitized photodynamic inactivation of E. coli and S. aureus could be significantly enhanced by tetradecyltrimethylammonium bromide (TTAB) as an OPE− TTAB complex (Table 4, entry 1, eq 16−18). 81The enhancement in photokilling was 3 log in E. coli and 2 log in S. aureus.The proposed mechanism involved a combination of the membrane-disrupting effect of anionic OPE and the photogenerated ROS.The formation of the positively charged OPE-TTAB complex allows anionic OPE to enter the bacterial cell membrane.The OPE-TTAB complex dissociates in the membrane due to the stronger interaction of TTAB with lipids.The now exposed anionic OPE experiences strong electrostatic repulsion with the negatively charged lipid bilayer resulting to membrane disruption and leakage.The repulsion may also cause the OPE to have greater access to the bacterial cytoplasm or periplasm, where it can cause further damage through ROS photogeneration.
TTAB OPE OPE TTAB complex An earlier study described the polyethylenimine (PEI)chlorin-e6-sensitized photinactivation of Cyrptococcus neoformans in the presence of caspofungin (Table 4, entry 2). 82A 4.0 log enhancement of cell kill was observed with caspofungin.The The Journal of Organic Chemistry pubs.acs.org/jocPerspective mechanism was thought to be due to Type-I and Type-II ROS along with caspofungin's inhibitory effect on (1,3)β-D-glucan synthase, leading to the reduction of (1,3)β-D-glucan, thus compromising cell wall integrity, leading to a greater uptake of PEI-chlorin e6.
In 2010, a report appeared on the RB, chlorin-e6, and polycationic polyethylenimine/chlorin e6 conjugate (PEIchlorin-e6)-mediated photodynamic inactivation of Candida albicans in the presence of the natural product, saponin A16 (Table 4, entry 3). 83The enhancement in the photokilling was 2.0−5.0 log with RB and chlorin-e6 with saponin A16.The proposed mechanism involved greater production of Type-I and Type-II ROS due to cell membrane disruption by saponin A16, thereby increasing RB and chlorin-e6 uptake.The polycationic charge of PEI-chlorin e6 facilitated its entry into the fungal cell; thus no enhancement in photokilling was observed in the presence of saponin A16. 83 report in 2010 described 5,10,15,20-tetrakis(1-methyl-4pyridinio)porphyrin tetracation (TMPyP)-sensitized photodynamic inactivation of fungus C. albicans in the presence of miconazole.Inclusion of the latter led to a 0.3 log enhancement in fungal killing (Table 4, entry 4). 84Mechanistically, it was proposed that miconazole's effect was due to its ability to inhibit mitochondrial ATPase and mitochondrial respiration, resulting in the formation of additional ROS and thereby enhancing the oxidative stress of PDT.
More recently, there were two reports dealing with airborne 1 O 2 -mediated inactivation of E. coli in the presence of alkene surfactant 1 by an L-D reaction sequence (Figure 8, Table 4, entry 5). 85,86The enhancement in photokilling was 0.23−0.30log for E. coli.The proposed mechanism was based on the toxicity of airborne 1 O 2 itself (k T for 1 O 2 with alkene surfactant 1 at the air−water interface is 1.1 × 10 6 M −1 s −1 ), and in the dark subsequent toxicity from ROS due to allylic hydroperoxide decomposition likely forming hydrotrioxide, methyl radical, additional 1 O 2 .This is thought to be a 1 O 2 priming reaction and subsequent contribution from ROS in the dark due to the oxidized additive decomposition.Interestingly, hydroperoxide products 2 and 3 are of very low toxicity on their own.Rather, they become toxic only after the bacteria are pretreated with 1 O 2 singlet oxygen.Thus, there is a "one-two" punch, where the alkene surfactant has a strong secondary toxic effect, due to its light-independent (dark) reaction.

■ MECHANISMS
After photooxidation forming HO

The Journal of Organic Chemistry
arises not only in L-D but also D-L sequences causing greater photooxidative damage (Figure 9).It is valuable to highlight how this L-D and D-L reaction sequence with additives boosts photooxidative activity.We highlight examples in which (1) amino acids and lipids enhance the net oxidizing power in photooxidation reactions, mainly attributed to peroxy intermediates formed downstream some with greater destructive power than 1 O 2 itself.Such post-irradiation organic sulfide ROS included persulfoxide R 2 S + OO − and hydroperoxy-sulfonium ylide RS(=CH 2 R)OOH.(2) Cholesterol-enhanced PDT arises by post-irradiation membrane damage/dysfunction caused by PDT-generated lipid hydroperoxides.While primary membrane LOOHs may be deleterious on their own (depending on levels attained), they could induce secondary or after-light LPO which is potentially more damaging and cytotoxic.This may be due to light-independent redox turnover of primary photogenerated LOOHs. 87This turnover, which results in damaging LPO, could occur in originating membranes or in others to which primary LOOHs may have been transferred.(3) For ALA-PDT, a variety of adjuvants have shown success, including repurposed drugs, vitamins, and iron chelating agents.This is clearly due perturbation of heme biosynthesis by a D-L path increasing CPO and decreasing FC, leading to increased amounts of PpIX.Enhancement in PpIX concentrations were up to 17-fold in vitro and 5.8-fold in vivo.(4) Table 3 shows that inorganic compounds lead to amplified photodynamic of up to 6 or 7 log killing of bacteria or fungi.This amplification of the photosensitized activity is attributed to RMS including NO 2 − , and HOOI.These RMS will of course have a varying range of lifetimes and reactivities.Thus, compounds and cell constituents otherwise unreactive to 1 O 2 may be decomposed.
(5) Drugs can react by destabilizing cell walls and membranes (caspofungin and saponin A16), inhibiting DNA synthesis (5fluorouracil), or sequestering iron (desferrioxamine and HPO dendrimer) to amplify damage pre-or postphotodynamically. Amplified photodynamic killing enhanced PDI is found with surfactant additives, where dark toxicity from ROS due to allylic hydroperoxide decomposition likely forming hydrotrioxide, methyl radical, additional 1 O 2 based on the proposed mechanism in Figure 8.The state-of-science is currently limited to classes of additives that have been covered here.Future screening of additives that are, e.g., susceptible to peroxidation, could allow for a rational selection with downstream RMS mechanistic pathways in mind.Broader classes of additives could be tested for deeper insight into predicting wellperforming additives.
■ CONCLUSIONS An importance of this Perspective is emphasized, namely that not only for organic chemists, but also photobiologists, that there is a need to know more about underlying mechanisms to assist in therapies like PDT.We now report on progress in understanding of mechanistic details of additives to amplify photooxidation reactions.Greater oxidizing power is sometimes caused by the additives.The generation of RMS to enhance photooxidation was discussed, where the RMS are often tentatively assigned.A challenge is the direct detection of RMS arising from additives, which form in downstream reactions.
Challenging problems that await exploration include: (i) the design of photoreactions to deconvolute the light and dark paths, to have better amplification control, (ii) further development of techniques to monitor RMS in cells and microbes, (iii) examination of wider range of additives, including sensitizers as oxidizable substrates themselves, (iv) to assess the amplification of photooxidation based on RMS vs drugs to provoke different mechanisms, (v) establishing additive roles both as adjuvant and converse, e.g., Fe chelators where low amount (promotes radical generation and oxidative stress via Fenton-like reactions), and high amount (starves the organism of this essential metal making it weaker and more susceptible to photodynamic stress).Efforts are needed to capitalize further on additives for an understanding to amplify photooxidative reactions.Thus, we believe that exploring additive-based photodynamics and mechanisms of action has a bright future in light as well as dark reactions.

■ ASSOCIATED CONTENT Data Availability Statement
The data underlying this study are available in the published article.Alexander Greer is a professor of chemistry at Brooklyn College of the City University of New York (CUNY) with more than 25 years of experience in photochemistry, organic chemistry of singlet oxygen, peroxy intermediates, and photodynamic therapy.He co-founded SingletO2 Therapeutics LLC and is a past president of the American Society for Photobiology, associate editor of Photochemistry and Photobiology, and co-chair of the Committee of Concerned Scientists.

Figure 1 .
Figure 1.Upon exposure to light, the sensitizer is excited, which in the presence of oxygen forms oxygen radicals and radical ions by Type-I processes or singlet oxygen ( 1 O 2 ) by a Type-II process leading to oxidized and oxygenated products.

Figure 2 .
Figure 2. Light-dark (L-D) reaction sequence with ROS from Type-I or Type-II photosensitized oxidation followed by RMS production for overall amplification of reactivity.In the L-D sequence, various RMS initially form; an example of sens-OO • is the peroxyl radical formed in the self-sensitized photooxidation of toluidine blue O.An opposite reaction sequence can occur, namely a dark-light (D-L) reaction sequence.Middle ground where L-D and/or D-L sequences may arise, such as the integrity of the cell membrane being compromised for greater cell permeability, where in turn sensitizer uptake is higher for greater photooxidative damage.

Figure 3 .
Figure 3. Structures of some additives discussed in this Perspective.

Figure 5 .
Figure 5. Reaction of diethyl sulfide with 1 O 2 in the presence of an additive (diphenyl sulfoxide or diphenyl sulfide).Under the reaction conditions, Ph 2 SO and Ph 2 S are themselves not reactive with 1 O 2 .

Figure 6 .
Figure 6.Formation and possible fates of lipid hydroperoxides (LOOHs).Selected free radical and nonradical initiators of lipid peroxidation are shown.The translocation of the LOOH to an acceptor mebrane or transesterification to an acceptor lipid is represented.Also shown is the turnover of lipid hydroperoxide (LOOH) intermediates via (a) one-electron reduction, which results in damage enhancement, and (b) two-electron reduction, which results in damage containment.

Figure 7 .
Figure 7. Topical administration of ALA that in the presence of an adjuvant (listed in Table2) is followed by enhanced PpIX formation and greater photodynamic killing.This is a D-L reaction sequence.

Figure 8 .
Figure 8. Percent of E. coli killed by airborne 1 O 2 alone (yellow bars) and the additional percent of E. coli killed upon adding hydroperoxides 2 and 3 in the dark as a follow-up treatment (gray bars).This is an L-D reaction sequence.

Figure 9 .
Figure 9. Mechanistic summary of L-D and D-L reaction sequences leading to overall greater photooxidative damage.

Biographies
Lloyd Lapoot received his B.S. degree and M.S. degree in agricultural chemistry from the University of the Philippines, Los Banõs.In 2024, he received his Ph.D. in biochemistry as a member of Professor Alexander Greer's research group at Brooklyn College and The Graduate Center of the City University of New York (CUNY).His research interests are in singlet oxygen reactions and their application in the photodynamic killing of cancer cells.Shakeela Jabeen received her B.S. from Brooklyn College.In 2022, she received her Ph.D. in chemistry as a member of Professor Alexander Greer's research group at Brooklyn College and The Graduate Center of the City University of New York (CUNY).She is now a chemist at the Food and Drug Administration (FDA).Ryan O'Connor received his B.S. degree in chemistry from Brooklyn College of the City University of New York (CUNY) as a member of Professor Alexander Greer's research group.He received his M.S. degree in chemistry from New York University as a member of Professor Marvin Parasram's research group.He is currently a New York City Teaching Fellow for the New York City Department of Education.Witold Korytowski is a professor of biophysics at Jagiellonian University.His research interests are focused on the role of lipid hydroperoxides in human pathology.He has contributed extensively to the field of photodynamic therapy by investigating the role of physiological nitric oxide in regulation of cancer cell proliferation, migration, and metastasis.Albert Girotti is an emeritus professor of biochemistry at the Medical College of Wisconsin.He has long-standing interests in the pathologic and therapeutic effects of oxidative stress, including photodynamic stress used in antitumor photodynamic therapy (PDT).His work has focused on generation, translocation, signaling action, and detoxification of lipid oxidation products, particularly lipid hydroperoxides (LOOHs).His recent work has focused on biochemical factors that influence PDT efficacy, e.g., the antagonistic effects of (i) LOOHdetoxifying GPx4 and (ii) iNOS-generated nitric oxide (NO), both effects being discovered in his laboratory.The Journal of Organic Chemistry pubs.acs.org/jocPerspective

Table 1 .
Additives That Amplify the Activity of 1 O 2

Table 2 .
2 • ) and RB radical anion (RB •− ), reaction of the latter with O 2 giving Additive Boosting of Photodynamic Action by Increasing the PpIX Concentration in Cells

Table 3 .
Additive Boosting of Photodynamic Inactivation of Bacteria and Fungi In the following section, more examples of additives are discussed in photooxidative eradication of microbes.

Table 4 .
Additive Boosting of Photodynamic Inactivation of Bacteria and Fungi •, and H 2 O 2 in Type-I reactions, and 1 O 2 in the Type-II reaction, the presence of additives can lead to subsequent RMS.The formation of RMS Department of Biochemistry, Medical College of Wisconsin, Milwaukee, Wisconsin 53226, United States; Email: agirotti@mcw.eduAlexander Greer − Department of Chemistry, Brooklyn College of the City University of New York, Brooklyn, New York 11210, United States; Ph.D. Program in Biochemistry and Ph.D. Program in Chemistry, The Graduate Center of the City University of New York, New York, New York 10016, United States; orcid.org/0000-0003-4444-9099;Email: agreer@ brooklyn.cuny.edu