Not all inorganic salts are suitable as substrates for reduction reactions. Inorganic salts that can undergo reduction usually contain a central element in a relatively high oxidation state, such as N(+5) in NO₃⁻, S(+6) in SO₄²⁻, Cl(+7) in ClO₄⁻, Cr(+6) in CrO₄²⁻, as well as Fe(III), Mn(IV), Se(IV/VI), and Te(IV/VI). These substances can accept electrons through microbial electron transport chains, plant assimilatory metabolism, enzymatic systems, or chemical reducing agents, and are converted into products with lower oxidation states.

Keywords: inorganic salt reduction reaction; nitrate reduction; nitrite reduction; sulfate reduction; thiosulfate reduction; perchlorate reduction; iron reduction; metal salt reduction

1 Logic for Determining Whether an Inorganic Salt Can Undergo Reduction

1.1 Common Features of Reducible Inorganic Salts

(1) The central element is in a high oxidation state

Reducible inorganic salts are usually not defined by stable cations such as Na⁺, K⁺, Ca²⁺, or Mg²⁺, but by the high-valent state of the anion or metal center. For example, NO₃⁻, NO₂⁻, SO₄²⁻, SO₃²⁻, ClO₄⁻, ClO₃⁻, CrO₄²⁻, SeO₃²⁻, and TeO₃²⁻ all have the potential to accept electrons.

(2) Clear lower-valent products can be formed

To determine whether an inorganic salt can serve as a reduction substrate, it is necessary to assess whether it can form stable or detectable lower-valent products. For example, NO₃⁻ can be converted into NO₂⁻, NO, N₂O, N₂, or NH₄⁺; SO₄²⁻ can be converted into H₂S or S²⁻; Fe(III) can be converted into Fe(II); and SeO₃²⁻ can be converted into elemental selenium.

(3) The reaction requires a suitable electron donor

Reduction of inorganic salts requires an electron source. In microbial systems, electrons may come from organic carbon, hydrogen, lactate, acetate, or reductive metabolic intermediates. In chemical systems, electrons may be provided by reducing agents such as zinc powder, ascorbic acid, sulfite, or borohydride. Without an electron donor, even a theoretically reducible substrate may not show an obvious reaction in experiments.

Table 1 Typical Reducible Inorganic Salts and Their Reduction Products

1.2 Inorganic Salts Usually Not Used as Reduction Substrates

(1) Stable alkali metal salts

Na⁺ and K⁺ in NaCl, KCl, and Na₂SO₄ are usually not used as biological reductive electron acceptors. They are more commonly used to regulate osmotic pressure, ionic strength, or to provide basal salts in culture media.

(2) Common divalent metal salts

MgCl₂ and CaCl₂ are commonly used in enzyme reactions, cell culture, or buffer systems. In conventional aqueous and biological systems, Mg²⁺ and Ca²⁺ usually do not undergo further reduction.

(3) Salts already in a low oxidation state

NH₄⁺ salts, Fe²⁺ salts, and S²⁻ salts are often reduced products or components associated with electron donors, and are generally not treated as typical “reduction substrates.”

Table 2 Common Inorganic Salts Usually Not Used as Reduction Substrates

2 Reduction Reactions of Nitrogen-Containing Inorganic Salts

2.1 Nitrate Reduction

(1) Reduction of NO₃⁻ to NO₂⁻

Nitrogen in nitrate is in the +5 oxidation state and can be converted into NO₂⁻ by nitrate reductase. This reaction is commonly used in microbial nitrate reduction tests, plant nitrate assimilation, wastewater denitrification, and soil nitrogen cycle research.

(2) Further reduction of NO₂⁻

NO₂⁻ can be further converted into NO, N₂O, and N₂, or converted into NH₄⁺ through DNRA. If NO₃⁻ decreases but NO₂⁻ does not accumulate in an experiment, NH₄⁺, N₂O, or N₂ should be measured simultaneously to avoid misinterpreting deep reduction as no reduction.

(3) Color development and zinc powder interpretation

In the microbial nitrate reduction test, a red color after adding the color reagent indicates NO₂⁻ accumulation. No red color after adding the color reagent cannot be directly interpreted as negative; zinc powder must be added to determine whether NO₃⁻ remains.

Table 3 Typical Interpretation of the Nitrate Reduction Test

2.2 Nitrite Reduction

(1) Denitrification pathway

Nitrogen in NO₂⁻ is in the +3 oxidation state and can be further reduced to NO, N₂O, and N₂. This pathway is used to evaluate denitrification capacity and is common in low-oxygen soils, sediments, wastewater treatment systems, and facultative anaerobic microorganisms.

(2) DNRA pathway

NO₂⁻ can also be reduced to NH₄⁺. This process does not remove nitrogen from the system in gaseous form; instead, nitrogen is retained as ammonium. It is suitable for studying nitrogen retention in sediments, anaerobic sludge, and organic-carbon-rich environments.

(3) Assimilatory reduction

Plants, algae, and some microorganisms can convert NO₃⁻/NO₂⁻ into NH₄⁺, which then enters amino acid and nucleotide synthesis. This pathway is usually not characterized by large accumulation of free NH₄⁺, and should be analyzed together with biomass, protein nitrogen, and amino acid content.

3 Reduction Reactions of Sulfur-Containing Inorganic Salts

3.1 Thiosulfate Reduction

(1) Substrate for H₂S production tests

Thiosulfate can be reduced by some microorganisms to generate H₂S. H₂S reacts with iron or lead salts in the medium to form black precipitates, so it is commonly used to interpret H₂S production in media such as TSI, KIA, and SIM.

(2) Interpretation of black precipitate

A black precipitate indicates sulfide formation, but does not directly prove that the substrate must be sulfate. Different media contain different sulfur sources. Interpretation should consider the medium formulation, inoculation method, and incubation time.

(3) Significance in strain differentiation

H₂S positivity can help differentiate Salmonella, Proteus, Citrobacter, and some anaerobes, but it cannot serve as definitive evidence for species identification on its own. In practice, it should be interpreted together with sugar fermentation, indole, urease, lysine decarboxylase, motility, and other results.

3.2 Sulfite and Sulfate Reduction

(1) Sulfite reduction

Sulfur in sulfite is in the +4 oxidation state and can be further reduced to S⁰ or H₂S. This reaction is common in studies of anaerobes, food microorganisms, and environmental sulfur cycling, and is suitable for evaluating sulfide production by strains.

(2) Sulfate reduction

Sulfur in sulfate is in the +6 oxidation state. Reduction to H₂S requires strongly reducing conditions and a complete sulfate reduction enzyme system. This reaction commonly occurs in strictly anaerobic or strongly reducing environments, and is central to studies of sulfate-reducing bacteria, oilfield corrosion, black-odor water bodies, and sediment sulfur cycling.

(3) Dependence on electron donors

Sulfate reduction usually requires lactate, acetate, hydrogen, or other reductive substrates as electron donors. If electron donors are insufficient, obvious sulfide production may not be observed even when SO₄²⁻ is present in the system.

Table 4 Types of Reduction Reactions of Sulfur-Containing Inorganic Salts

4 Reduction Reactions of Halogen-Containing Inorganic Salts

4.1 Chlorate and Perchlorate Reduction

(1) Perchlorate reduction

Chlorine in ClO₄⁻ is in the +7 oxidation state and can be reduced stepwise by specific microorganisms to ClO₃⁻, ClO₂⁻, and finally Cl⁻. This reaction is commonly used in biological remediation of perchlorate pollutants and anaerobic respiration research.

(2) Chlorate reduction

Chlorine in ClO₃⁻ is in the +5 oxidation state and can be further reduced to ClO₂⁻ or Cl⁻. This reaction requires corresponding chlorate reductases and electron donors, and is commonly used for screening chlorate-reducing bacteria.

(3) Experimental safety and by-products

Chlorate, perchlorate, and their intermediates are strongly oxidizing. Experiments should control concentration, avoid improper mixing with strong reducing agents or organic matter, and pay attention to waste disposal.

4.2 Bromate and Iodate Reduction

(1) Bromate reduction

Bromine in BrO₃⁻ is in the +5 oxidation state and can be reduced to Br⁻. In water treatment systems, bromate reduction is often used to evaluate pollutant removal, reducing materials, and microbial reduction capacity.

(2) Iodate reduction

Iodine in IO₃⁻ is in the +5 oxidation state and can be reduced to I₂ or I⁻. This reaction is associated with the marine iodine cycle, iodine migration in soil, and iodine speciation in reducing environments.

(3) Key interpretation points

Reduction of halogen oxyanion salts should not be judged solely by substrate decrease. Halide generation, intermediates, and abiotic reduction background should also be measured. In complex systems, sterile and inactivated controls are required.

Table 5 Reducible Halogen Oxyanion Salts

5 Reduction Reactions of Metal Salts and Metalloid Salts

5.1 Iron and Manganese Salt Reduction

(1) Fe(III) reduction

Fe(III) can serve as an electron acceptor for some microorganisms and be reduced to Fe(II). Iron reduction is commonly studied in sediments, groundwater, anaerobic sludge, and iron-reducing bacteria. Fe(II) color development or quantification is a common criterion.

(2) Mn(IV) reduction

Mn(IV) oxides can be reduced to Mn(II). This process is usually accompanied by changes in the color, solubility, and morphology of solid-phase minerals. Dissolved Mn(II) and solid-phase transformation should be monitored simultaneously.

(3) Abiotic reduction interference

Fe(III) and Mn(IV) can be abiotically reduced by reducing medium components, sulfide, ascorbic acid, or natural organic matter. Sterile and inactivated controls must be included in the experiment.

5.2 Reduction of Chromium, Selenium, Tellurium, and Arsenic Salts

(1) Cr(VI) reduction

Cr in CrO₄²⁻ and Cr₂O₇²⁻ is in the +6 oxidation state and can be reduced to Cr(III). This reaction is commonly used in studies of hexavalent chromium pollution remediation. Because Cr(VI) is highly toxic, experimental concentration, protection, and waste disposal must be strictly controlled.

(2) Se(VI/IV) reduction

SeO₄²⁻ and SeO₃²⁻ can be reduced to elemental selenium or lower-valent selenium. Selenite reduction often produces red or orange-red deposits, making it suitable for screening selenium-transforming bacteria and preparing biogenic nano-selenium.

(3) Te(VI/IV) reduction

TeO₄²⁻ and TeO₃²⁻ can be reduced to black elemental tellurium. Tellurite is commonly used for selective culture and resistant strain screening, with black colonies serving as a common preliminary screening signal.

(4) As(V) reduction

AsO₄³⁻ can be reduced to As(III). This reaction changes arsenic mobility and toxicity, and is commonly studied in environmental arsenic cycling and arsenic-reducing microorganisms.

Table 6 Types of Reduction Reactions of Metal Salts and Metalloid Salts

6 Experimental Interpretation and Detection Methods

6.1 Interpretation Principles

(1) First determine whether the substrate is reducible

Before the experiment, confirm whether the key element in the inorganic salt is in a high oxidation state. If the salt is only a stable background salt, such as NaCl, KCl, or MgCl₂, it should not be designed as the main reduction substrate.

(2) Then confirm whether the reduction product is detectable

Reducible substrates require matching product detection methods. NO₂⁻ can be detected by Griess color development, Fe(II) by Ferrozine color development, H₂S by black precipitation or sulfide colorimetry, and Se⁰ and Te⁰ by precipitate color and elemental analysis.

(3) Distinguish reduction, adsorption, and precipitation

A decrease in substrate concentration does not necessarily indicate reduction. Metal salts, selenates, tellurates, and chromates are especially prone to adsorption, precipitation, or matrix binding. Lower-valent products should be detected simultaneously, and sterile controls should be included.

Table 7 Common Detection Indicators for Inorganic Salt Reduction Reactions

6.2 Control Setup

(1) Sterile control

Used to exclude abiotic reduction caused by the medium, reducing agents, or environmental conditions. If substrate decrease or color change also occurs in the sterile control, the chemical background should be addressed first.

(2) Inactivated control

Used to distinguish live-cell metabolism from cell adsorption, release after lysis, or non-enzymatic reduction. Inactivated controls are especially important in environmental samples, sludge, and sediment experiments.

(3) Electron donor control

Reduction reactions are usually limited by electron donors. A no-electron-donor control can determine whether the reaction depends on organic carbon, hydrogen, or other exogenous reductive substrates.

(4) Positive and negative systems

Microbial identification and functional strain screening should use known positive and negative strains. If quality-control strains behave abnormally, test sample results should not be directly interpreted.

7 Reagents and Materials Related to Inorganic Salt Reduction Reactions

Table 8 Key Substrates and Color Reagents for Inorganic Salt Reduction Reactions

Table 9 Detection Products and Culture Materials for Inorganic Salt Reduction Reactions

8 Common Questions

8.1 Which inorganic salts most commonly undergo reduction reactions?

The most common examples include nitrate, nitrite, thiosulfate, sulfite, sulfate, perchlorate, chlorate, bromate, iodate, Fe(III) salts, Mn(IV) oxides, Cr(VI) salts, Se(VI/IV) salts, Te(VI/IV) salts, and As(V) salts. The central elements in these substances are in relatively high oxidation states and can accept electrons.

8.2 Can salts such as NaCl, KCl, and MgCl₂ undergo reduction reactions?

In conventional aqueous systems, biological culture systems, and microbial identification systems, they are usually not used as reduction substrates. Na⁺, K⁺, Mg²⁺, and Ca²⁺ have stable valence states and are mainly used to regulate ionic strength, osmotic pressure, or enzyme reaction conditions.

8.3 Are sulfate reduction and thiosulfate reduction the same?

No. Sulfur in sulfate is in the +6 oxidation state, and its reduction to H₂S usually requires strictly anaerobic conditions and a complete sulfate reduction enzyme system. Thiosulfate is more commonly used in H₂S production tests. The reaction pathways and identification significance are different.

8.4 Why can a decrease in some inorganic salts not be directly interpreted as reduction?

Substrate decrease may result from adsorption, precipitation, volatilization, complexation, or detection interference. Only when lower-valent products such as NO₂⁻, NH₄⁺, H₂S, Fe(II), Se⁰, or Te⁰ are detected simultaneously can reduction be more reliably demonstrated.

8.5 What are the most commonly used inorganic salt reduction reactions in microbial identification?

The most commonly used tests are the nitrate reduction test and the H₂S production test. The former determines whether NO₃⁻ has been reduced, while the latter often identifies sulfur-containing substrate reduction by detecting black sulfide precipitates. Both require interpretation together with controls and other biochemical indicators.

8.6 Why must sterile controls be included in metal salt reduction experiments?

Metal and metalloid salts are easily affected by medium components, reducing substances, pH, and light, leading to abiotic reduction or precipitation. Sterile controls can exclude chemical background effects and are necessary for determining the microbial contribution to reduction.

Inorganic salts capable of undergoing reduction reactions are mainly concentrated among oxyanion salts containing high-valent central elements and high-valent metal or metalloid salts. Experimental design should first confirm whether the substrate is reducible, then select detection methods that can indicate lower-valent products. A decrease in salt concentration or a color change alone is insufficient to prove a complete reduction pathway.

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