When Operation Barbarossa started, the IAR 80 equipped Esc. 41, 59 and 60 of Grupul 8 Vânátoare, part of the Grupul Aerian de Lupta (GAL), that were tasked to support the Romanian 3rd and 4th Armies deployed at the southern flank of the Eastern Front.
Grupul 8 was the only unit assigned a pure fighter role, while Grupul 5 and Grupul 7, equipped with German superior aircraft (Heinkel He 112s and Messerschmitt Bf 109s) were employed primarily as fighter-bombers and bomber escorts.
On 22 June 1941, during the first day of the offensive, the IAR 80 patrols had their baptism of fire, achieving a single aerial victory (claimed by Sublocotenent aviator Ioan Miháilescu of Esc 60 van, a future ace) during four separate air combats. However, at least four IARs force landed with battle damage, while another two suffered engine trouble. By the end of 1941, 20 IAR 80/81s had been lost in combat in accidents. During 1942 the Romanian aviation industry reached its highest output so that the Royal Romanian Air Force could be re-equipped as follows: Esc. 47, 48 and 52 (Grupul Vânátoare), Esc. 43, 44 and 50 (Grupul 3 Vânátoare) and Esc. 41, 42 and 60 (Grupul 8 Vânátoare) received the new IAR 80A. Esc. 53 also replaced its Hurricanes with the IAR 80A, while Grupul 6 Bopi re-equipped with the IAR 81.
In June 1942, the operational IAR fighter forces on the eastern front, combined into the Flotilla 2 Vânátoare consisted of Grupul 8 Vânátoare, commanded by Cdr. Lt Col E. Pirvulescu, and included Escadrila 41, Escadrila 42 and Escadrila 60 with 12 IAR 80As each. During the Battle of Stalingrad, on 12 September, Grupul 8 Vânátoare IAR 80Bs (along with Grupul 7 Vânátoare’s Bf 109s) claimed to have shot down seven Yaks but they lost two IARs. Grupul 8 moved at the end of September, to Karpovka, joining Grupul 7, equipped with Bf 109s. On 12 and 13 December, Grupul 6 used its IAR 81s to support the German counterattack by the Panzergruppe Hoth of the Heeresgruppe Don, from Kotelnikovo towards Stalingrad. In the summer of 1943 the FARR's IAR-80s were transferred to Romania for air defense duties, where they were used in combat against the USAAF. USAAF attacks were directed at the oil refinery installation at Ploiesti, in particular. On 1 August 1943 the IAR 80 faced the Consolidated B-24 Liberator heavy bomber for the first time. There were 178 B-24s from 9th USAAF, part of the Operation Tidal Wave. The IAR 80Bs of Escadrila 61 and 62 of Grupul 6 Vânátoare, as well as IAR 80Cs from the newly formed Escadrila 45 of Grupul 4 Vânátoare, together with the Bf 109Gs from Esc. 53 and Bf 110s from the Romanian night fighter squadron, dived on the low-flying, four-engined bombers, belonging to five USAAF bomber groups (the 44th, 93rd, 98th, 376th and 389th). The Americans lost – in combat or on the way back – 51 bombers. Only 89 reached their bases, of which only 31 were serviceable for a mission the next day. The Romanian pilots claimed 25 certain and probable victories for just two losses, one IAR 80 B and one Bf 110C. According to Romanian statistics, IARs and Messerschmitts were confirmed as having shot down ten B-24s, with two probables.
On 10 June 1944, IAR 80s took part in a major air battle when the USAAF attacked Ploiesti with 36 P-38 Lightnings of the 82nd Fighter Group carrying one bomb each, escorted by 39 Lightnings of the 1st and 82 FGs. The IAR 81Cs from Grupul 6, as well as the German fighters from I./JG 53 and 2./JG 77, intercepted the large American formation. Romanian pilot Dan Vizanty, commander of Grupul 6, recalled later:
"Our Lightning attack came as a complete surprise to the Americans. Our attack was so quick that not one of the 100 (sic) American aircraft managed to fire a single shot at our aircraft parked on the ground. Everything happened between ground level and about 2,000 meters (6,600 feet), and was total confusion. I was excited and proud of my "mills", the IAR 80s, which, thanks to their extraordinary agility, remained victorious in the air. I saw their crazy dives, quick rolls, reverse turns and inverted flying, always with just brief burst of fire to save ammunition. It was an incredible sight, but also a drama for the Lightning pilots, who, at this low altitude, were inferior to the ever-present, nimble IAR 80s".
The USAAF lost 22 or 23 P-38s on that day. Eight were claimed by Grupul for themselves – the rest were claimed by the Luftwaffe and by anti-aircraft fire. The Americans claimed 23 victories, although the Romanians and Germans each reported only one aircraft lost on that day.
The American account of this battle conflicts significantly with the Romanian one. Fighter pilot Herbert "Stub" Hatch, who took part in the dogfight, wrote that his flight of 16 P-38s, the 71st Fighter Squadron, was challenged by a large formation of Romanian IAR 81C fighters that he misidentified as Focke-Wulf Fw 190s. According to Hatch, the fight took place at and below 300 feet (100 m) in a narrow valley. Hatch saw two IAR 81Cs hit the ground after taking fire from his guns, and his fellow pilots confirmed three more kills from his guns, making Hatch an ace in a day. However, the outnumbered 71st Fighter Squadron lost nine aircraft. The Americans never again repeated the P-38 dive-bombing mission profile over Romania. But during 1944 USAAF aircraft appeared over Romania in more significant numbers. Many air combats occurred and by the time of their last encounter with the USAAF on 3 July 1944, pilots of Grupul 6 vanatoare had submitted 87 confirmed and ten unconfirmed claims. Casualties among the Romanian fighter pilots quickly mounted too. The three IAR 80/81 groups (the 1st, 2nd and 6th) in a period of less than four months – known as the "American Campaign" – had at least 32 IAR pilots killed in action, including 11 aces. These losses exceeded the number of casualties suffered in the previous two and a half years of fighting against the Soviets. Because of heavy losses, all IAR 80/81 units were withdrawn from combat against Americans in July 1944 and IAR pilots started to convert to the more modern Bf 109G-6s.